Australian Biochemist the Magazine of the Australian Society for Biochemistry and Molecular Biology Inc

ISSN 1443-0193 Australian Biochemist The Magazine of the Australian Society for Biochemistry and Molecular Biology Inc. Volume 47 APRIL 2016 No.1

SHOWCASE ON RESEARCH Stem Cells in Development and Cancer

THIS ISSUE INCLUDES

Showcase on Research Regular Departments  Haematopoietic Stem Cells  SDS (Students) Page  Repairing the Damaged Heart  Competition with Cardiac Progenitor Cells  Sydney Protein Group SIG  Defining Prostate Stem Cells  Our Sustaining Members  Generating Kidney Tissue  Forthcoming Meetings from Pluripotent Stem Cells  Directory

INSIDE Profiles of ASBMB Medallists and Awardees Vol 47 No 1 April 2016 AUSTRALIAN BIOCHEMIST Page 1 ‘Lemberg Medal’ Competition

We have another competition for the readers of the Australian Biochemist. All correct entries received by the Editor (email [email protected]) before 16 May 2016 will enter the draw to receive a gift voucher. With thanks to Rebecca Lew. Fill in the blanks around the letters in the words LEMBERG MEDAL with the names of previous medallists. L E M B E R G M E D A L

The Australian Academy of Science Awards for 2017 are now open

Please note that the closing date for nominations for honorific awards is 30 April 2016 and that nominations/ applications for research conferences, research awards and travelling fellowships are due by 15 June 2016. Nomination schemes and awards that may be of particular interest to you are also highlighted below:

Gottschalk Medal (biomedical sciences) Nancy Millis Medal for Women in Science (any branch of the natural sciences) Ruth Stephens Gani Medal (human genetics including clinical, molecular, population and epidemiological genetics and cytogenetics) Gustav Nossal Medal (global health) Jacques Miller Medal (experimental biomedicine)

Boden Research Conferences

The Moran Award for history of science research

Oxford Nuffield Medical Fellowship Selby Travelling Fellowship (all fields of science)

Further information on the awards (including eligibility criteria and nomination/application procedures and forms) visit the website: www.science.org.au/awards or email: Dominic Burton (Awards Officer) [email protected]

Page 2 AUSTRALIAN BIOCHEMIST Vol 47 No 1 April 2016 SHOWCASE ON RESEARCH

EDITORIAL Stem Cells - What Makes Them Special? Stem cell biology is a rapidly moving field with transplantation and to expand blood stem cells ex vivo. profound implications for regenerative medicine and In their review of cardiac progenitor cells, Le and cancer. Stem cells can be broadly divided into two Chong show that the isolation and characterisation groups: 1. embryonic stem (ES) cells, which include of stem and progenitor cells in the normal heart is induced pluripotent stem cells (iPSC), and 2. tissue- crucial for developing strategies to replenish damaged specific stem cells (adult stem cells). The hallmark cardiac cells. Cardiovascular disease remains a major feature of stem cells is extensive self-renewal capacity. cause of morbidity worldwide. While it is still unclear ES cells are undifferentiated, pluripotent cells that can if implantation of progenitor cells into injured hearts give rise to every type of cell in the body. After birth, leads to myocardial repair by direct or indirect means, distinct populations of adult stem cells (very small in the delineation of paracrine signalling pathways number) are found in most organs and cellular systems. that influence the growth of these cells following In contrast to ES cells, adult stem cells are multipotent, transplantation remains an important area of research. as the differentiation capacity of these cells is limited In the third article, Taylor describes stem and to the organ or tissue in which they reside. These progenitor cells in prostate tissue and emphasises the tissue-specific stem cells maintain tissue homeostasis importance of dissecting heterogeneity to understand and repair damaged cells throughout the life of the the cell populations that drive prostate tumour growth. individual. Cell fate choices must be tightly regulated to Identification of ‘cells of origin of cancer’ may reveal ensure the correct supply of mature cells, otherwise cell biomarkers for prognosis and improved treatment. transformation and carcinogenesis can result. Questions Future challenges include the identification of cancer of paramount importance in stem cell biology are: 1. how stem cell populations that drive tumour growth and can we use our knowledge of normal tissue to identify eradication of regenerating tumour cells that are better markers and therapies for cancer patients, and 2. resistant to hormonal therapy. how can one harness the regenerative potential of stem The discovery that a fully mature, specialised somatic cells to replace diseased or damaged tissue? cell could be induced to mimic embryonic stem cells This issue of the Australian Biochemist features four under appropriate conditions has opened a new field articles from leading Australian stem cell researchers. of iPSC biology. In the fourth review, Little explores In the first, Ng and Alexander provide an overview the recent advances in regenerating kidney tissue from of haematopoietic stem cells. For more than 50 years, pluripotent stem cells. Kidney organoids can now be the haematopoietic system has paved the way for formed ex vivo after directed differentiation of human understanding stem cells and lineage hierarchies iPSCs. These and other redirected iPSCs have important and for deciphering the cellular constituents of solid implications for disease modelling and patient-specific organs. Blood stem cells, although very rare, are drug screening to identify better druggable targets. The capable of sustaining haematopoiesis in bone marrow next era remains a very exciting one for Australian stem transplantation owing to their robust self-renewing cell biologists seeking to build on their past discoveries potential. Future efforts are focussed on advancing stem and implement improvements in clinical practice through cell transplantation therapy to enable efficient allogeneic tissue regeneration and novel therapeutic targets in cancer. Jane Visvader Walter and Eliza Hall Institute of Medical Research, 1G Royal Parade, Parkville, VIC 3052 visvader@wehi.edu.au

Cover Illustration Stem Cells in Development and Cancer A kidney organoid generated from Guest Editor: Jane Visvader a human pluripotent stem cell line (16). This organoid has been cultured 4 Haematopoietic Stem Cells for 18 days as an aggregate post- Ashley Ng and Warren Alexander induction of intermediate mesoderm. 7 Repairing the Damaged Heart with Cardiac Progenitor Cells Immunofluroescence displays the Thi Y.L. Le and James J.H. Chong presence of differentiating nephrons 10 Defining Prostate Stem Cells: Clues to Improving Prostate comprised of glomeruli (NPHS1, yellow), Cancer Treatment proximal tubules (LTL, pink) and distal Renea Taylor tubules/collecting ducts (CDH1, green). 13 Generating Kidney Tissue from Pluripotent Stem Cells Image courtesy of Dr Minoru Takasato Melissa Little (Murdoch Childrens Research Institute ).

Australian Biochemist – Editor Chu Kong Liew, Editorial Officer Liana Friedman © 2016 Australian Society for Biochemistry and Molecular Biology Inc. All rights reserved. Vol 47 No 1 April 2016 AUSTRALIAN BIOCHEMIST Page 3 SHOWCASE ON RESEARCH

Haematopoietic Stem Cells Ashley Ng1,2,3* and Warren Alexander1,2 1Walter and Eliza Hall Institute of Medical Research, Parkville, VIC 3052 2Department of Medical Biology, University of Melbourne, Parkville, VIC 3010 3Department of Clinical Haematology and Bone Marrow Transplantation, Royal Melbourne Hospital, Parkville, VIC 3050 *Corresponding authors: [email protected] Introduction they were eventually identified. Haematopoietic stem cells (HSCs) are the architects of The identification and characterisation of HSCs required definitive haematopoiesis, that is, blood cell production strategies to separate these rare bone marrow cells from that occurs continuously throughout the life of an organism. other more numerous cellular bone marrow components. Each HSC is programmed to allow the efficient production Functional competitive repopulating unit assays estimated of the cellular components of blood with a manifest purpose the frequency of these rare cells in bone marrow as 1 in that has been shaped by evolution. This can be observed 10,000 cells (13). Like the proverbial search for the needle in from red cells that have evolved to allow efficient carriage a haystack, HSCs were eventually isolated with increasing of oxygen, megakaryocytes and their platelet progeny that purity based on physical properties, such as Hoescht 33342 interact with blood vessels and soluble coagulation factors supravital dye exclusion (14), resistance to 5-fluorouracil to regulate clotting, to the cells of the innate and acquired (15) or gamma-irradiation (16). Ultimately, however, it immune systems that act against microbial attack. HSCs was the advent of flow cytometry and the use of specific are defined by their pluripotentiality: the capacity for a surface antigen markers (17) that led to the prospective single HSC to generate any and all of the diverse mature identification of cell populations able to reconstitute functional haematopoietic cell types, with key genes and multiple lineages upon transplantation into primary select genetic programmes invoked for the maintenance or recipients, and capable of self-renewal as judged by serial self-renewal of HSCs and for the formation of the specific transplantation assays. These cell populations enriched for haematopoietic lineages (1–5). HSCs, were notable for their lack of mature lineage antigen Definitive haematopoiesis in the embryo begins with expression, and expression of antigens such as cKit, the the emergence of the first identifiable HSCs in the cellular receptor for the cytokine stem cell factor (SCF) aorto-gonado-mesonephros region (6,7). Thereafter, (18,19). haematopoiesis shifts to the fetal liver, and then to the bone HSCs were found to possess unique properties that marrow, where HSCs reside for the life of the mammal appeared to set them apart from other blood-forming (8,9) (Fig. 1, left panel). progenitor cells. In addition to the properties of pluripotency and self-renewal, long-term HSCs in the Identification of Haematopoietic Stem Cells adult were found to reside in a specific niche environment It became apparent, initially through work that sought in the bone marrow that was closely associated with the to characterise radiation sensitivity, that donor adult bone endosteum (20), where they existed in conditions of relative marrow transplanted into syngeneic irradiated murine hypoxia (21). Here, HSCs predominantly existed in a non- recipients was capable of protecting the recipients from replicative and quiescent state (22), in which signalling by lethal doses of irradiation by regenerating (reconstituting) the cytokine thrombopoietin (TPO) (23,24) and the presence the ablated haematopoietic system. This observation was of megakaryocytes appear to play an important role (25– crucial in the development of the concept of HSCs as cells 27). In contexts that place the haematopoietic system under in the bone marrow that were capable of generating the stress, such as in chronic infection, these quiescent stem complete blood cell system, even though the specific cell cells are recruited into the cell cycle by regulatory pathways had yet to be isolated and characterised. that include interferon signalling, resulting in an increase In these early experiments, donor-derived clonogenic in downstream progenitor cells (28). Evidence increasingly colonies of multiple haematopoietic lineages were able to suggests that lineage specification occurs very early in be macroscopically identified in the spleen of transplanted the haematopoietic hierarchy in immunophenotypically recipient mice (10,11). These spleen colony forming units defined ‘stem cell’ populations (29–32), supporting (CFU-S), although not definitive HSCs, were nevertheless findings that self-renewing lineage-restricted progenitors useful in allowing characterisation of progenitor cells may emerge directly from HSCs (33). A significant degree responsible for haematopoietic reconstitution. Specific of complexity may therefore exist in the pathways by which progenitor cells appeared to possess the ability to mature haematopoeitic lineages develop from HSC and form multiple haematopoietic lineages within the one progenitor populations. More direct pathways from HSCs colony (pluripotency) while others appeared able to to mature cells may coexist alongside traditional models form daughter cells that retained the characteristics of that involve a hierarchy of progenitor populations. Recent the original parental cell (self-renewal) (12). These two evidence from clonal dynamic studies that traced the origin important characteristics of pluripotency and self-renewal of blood cells over time has suggested that the process came to be recognised as essential features of HSCs when of normal blood cell maintenance does not incessantly Page 4 AUSTRALIAN BIOCHEMIST Vol 47 No 1 April 2016 SHOWCASE ON Haematopoietic Stem Cells RESEARCH

Fetal to Adult Haematopoiesis Adult Stem Cell Markers Lineage- cKit+Sca1+

Platelet biased Yolk Sac Stem Cells Long-term Blood Islands CD150hi stem cell CD150+ CD48- CD48- CD41- Vwf+ AGM Rho123lo PsP CD34-/lo Myeloid biased E9.0 Flt3R- Stem Cells E8.25 Emergence CD49lo CD150+ of HSCs Yolk Sac CD244- CD48- CD229- CD41+ AGM Embryo Haemopoietic Fetal Liver E7.5 Progenitors Intermediate-term Rho123lo Primitive E10.5 stem cell CD34-/lo Erythrocytes De nitive Flt3R- Haemopoiesis CD150lo CD49hi

Short-term CD34hi stem cell Flt3R-/+ Rho123hi CD150-

“Adult” quiescent HSCs Lymphoid Myeloid > 4 weeks post partum Stem Cells Bone marrow Cycling HSCs CD150- Flt3R+ < 3-4 weeks post partum CD34+ CD27+ Bone marrow Flt3R+ Fig. 1. Left panel: the journey from fetal to adult haematopoiesis in the mouse, adapted from (8). PsP Para-aortic- splanchnopleura, AGM Aor-to-gonado-mesonephros. Right panel: Immunophenotypic markers of adult murine HSCs and ‘lineage restricted’ HSC populations. See references in the main text. call upon the quiescent HSCs to enter into cell cycle, but markers of human HSCs (lineage- CD34+CD38-) differ rather, the successive recruitment of long-lived progenitor from functionally similar murine counterparts (40). Unlike populations, which appear to primarily maintain blood inbred genetically and immunologically identical mouse cells at steady-state (34). strains, successful allogeneic transplantation therapy in In the setting of bone marrow transplantation, which humans requires significant immunological barriers to is dependent upon HSCs to sustainably reconstitute be overcome. Identification of the HLA system of MHC haematopoiesis, further refinement of cell surface markers class I and II receptors that engage T-cell antigen receptors has identified specific subsets of HSCs that have more allows appropriate matching of donors and recipients, limited capacity for self-renewal, yet are important for and the use of immunosuppression during and after the maintaining haematopoiesis in the short and intermediate transplantation of allogeneic stem cells from volunteer terms after transplantation (35,36). Stem cells capable (related and unrelated) donors into patients is essential of long-term reconstitution with durable self-renewal to prevent graft rejection and graft versus host disease. potential appear to be a very rare but crucial cell population Advancement of stem cell transplantation therapy has required for haematopoietic engraftment over long periods focussed on research to broaden the availability of donors of time (37,38,39) (Fig. 1, right panel). to patients. The use of cord blood units as a source of stem cells (41,42) and development of new conditioning Bone Marrow Transplantation as Therapy regimens have allowed haploidentical transplantation Although murine haematopoiesis reflects human (43) to become a therapeutic reality whilst limiting the haematopoiesis in many ways, the immunophenotypic immunological consequence of graft versus host disease. Vol 47 No 1 April 2016 AUSTRALIAN BIOCHEMIST Page 5 SHOWCASE ON Haematopoietic Stem Cells RESEARCH

Increasingly, these approaches are making allogeneic 15. Hodgson, G.S., and Bradley, T.R. (1979) Nature 281, transplantation an option for patients who otherwise do 381-382 not have a traditional source of HLA matched stem cells 16. Ploemacher, R.E., van Os, R., van Beurden, C.A., and available from related or volunteer unrelated donors. Down, J.D. (1992) Int. J. Radiat. Biol. 61, 489-499 17. Spangrude, G.J., Heimfeld, S., and Weissman, I.L. The Future (1988) Science 241, 58-62 Understanding the nature and regulation of HSCs 18. Okada, S., Nakauchi, H., Nagayoshi, K., et al. (1991) continues to be a very active and relevant field of research. Blood 78, 1706-1712 Delineating of how cell populations arise from HSCs and 19. Morrison, S.J., and Weissman, I.L. (1994) Immunity 1, the factors that regulate blood cell production has allowed 661-673 pathways and regulators of blood cells in ways that have 20. Morrison, S.J., and Scadden, D.T. (2014) Nature 505, revolutionised the treatment of blood disorders and the 327-334 use of stem cell therapy. The findings arising from stem 21. Nombela-Arrieta, C., Pivarnik, G., Winkel, B., et al. cell research continue to have a profound influence on our (2013) Nat. Cell Biol. 15, 533-543 understanding of blood disorders and will hopefully open 22. Wilson, A., Laurenti, E., Oser, G., et al. (2008) Cell 135, new doors for treatment of these conditions. 1118-1129 Important challenges remain. These include the 23. Qian, H., Buza-Vidas, N., Hyland, C.D., et al. (2007) Cell development of robust methods to maintain HSCs in vitro Stem Cell 1, 671-684 and expand cell numbers for therapy. These aims should 24. Yoshihara, H., Arai, F., Hosokawa, K., et al. (2007) Cell be able to be achieved through developing a deeper Stem Cell 1, 685-697 understanding of the HSC niche and the intrinsic and 25. Zhao, M., Perry, J.M., Marshall, H., Venkatraman, A., extrinsic regulators of HSCs, as well as developing the Qian, P., He, X.C., Ahamed, J., and Li, L. (2014) Nat. future capacity to ‘reprogram’ cells to HSCs. In addition, Med. 20, 1321-1326 the ability to correct genetically defective haematopoiesis 26. Bruns, I., Lucas, D., Pinho, S., et al. (2014) Nat. Med. 20, via transplantation of ‘corrected’ syngeneic patient cells is 1315-1320 an area of intensive ongoing research. This could potentially 27. Nakamura-Ishizu, A., Takubo, K., Kobayashi, H., lead to the correction of genetically defective HSCs in order Suzuki-Inoue, K., and Suda, T. (2015) J. Exp. Med. 212, to generate large numbers of HSCs for transplantation of 2133-2146 ‘corrected’ syngeneic patient cells. 28. Baldridge, M.T., King, K.Y., Boles, N.C., Weksberg, D.C., and Goodell, M.A. (2010) Nature 465, 793-797 References 29. Adolfsson, J., Mansson, R., Buza-Vidas, N., et al. (2005) 1. Ivanova, N.B., Dimos, J.T., Schaniel, C., et al. (2002) Cell 121, 295-306 Science 298, 601-604 30. Serwold, T., Ehrlich, L.I., and Weissman, I.L. (2009) 2. Mercer, E.M., Lin, Y.C., and Murre, C. (2011) Semin. Blood 113, 807-815 Immunol. 23, 317-325 31. Sanjuan-Pla, A., Macaulay, I.C., Jensen, C.T., et al. 3. Novershtern, N., Subramanian, A., Lawton, L.N., et al. (2013) Nature 502, 232-236 (2011) Cell 144, 296-309 32. Pietras, E.M., Reynaud, D., Kang, Y.A., et al. (2015) Cell 4. Moignard, V., Macaulay, I.C., Swiers, G., et al. (2013) Stem Cell 17, 35-46 Nat. Cell Biol. 15, 363-372 33. Yamamoto, R., Morita, Y., Ooehara, J., et al. (2013) Cell 5. Riddell, J., Gazit, R., Garrison, B.S., et al. (2014) Cell 157, 154, 1112-1126 549-564 34. Sun, J., Ramos, A., Chapman, B., et al.(2014) Nature 514, 6. Ivanovs, A., Rybtsov, S., Welch, L., Anderson, R.A., 322-327 Turner, M.L., and Medvinsky, A. (2011) J. Exp. Med. 35. Yang, L., Bryder, D., Adolfsson, J., et al. 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Repairing the Damaged Heart with Cardiac Progenitor Cells Thi Y.L. Le and James J.H. Chong* Centre for Heart Research, Westmead Institute for Medical Research, University of Sydney, and Department of Cardiology, Westmead Hospital, Westmead, NSW 2145 *Corresponding author: [email protected]

Introduction preclinical models of myocardial infarction (MI) and Cardiovascular disease continues to be a major heart failure (HF). These findings promised a paradigm cause of morbidity and mortality worldwide. Despite shift in cardiac biology and new opportunities for future considerable progress in revascularisation techniques treatment. However, 15 years after these initial reports, and pharmacotherapy, many patients progress to a consensus on the biological role of these populations heart failure after acute myocardial infarction. Current still remains elusive. In this review, we provide a brief therapies are unable to replace dead cardiomyocytes overview of CPCs currently being considered for cardiac and largely irreversible cardiac dysfunction ensues. The repair and the potential mechanisms of action of CPCs in discovery of multiple classes of stem cells has generated the damaged heart. hope for their use as therapeutic agents in heart failure. This may involve the transplantation of pluripotent stem Cardiac Progenitor Cells cells or multipotent adult progenitor cells into the area CPCs are a heterogeneous group of cells that are of the infarcted myocardium, to promote regeneration distributed throughout the heart, in areas including the of new functioning myocytes and vascular cells, thus atria, ventricles, and epicardium or pericardium. Under improving heart function. normal physiological conditions, CPCs are thought The adult mammalian heart was previously considered a to be inactive and do not contribute significantly to post-mitotic organ, without the capacity for self-renewal. cardiomyocyte renewal (3,4). After injury however, CPCs However, recent evidence suggests that the adult heart is can be activated and may differentiate into new myocytes capable of cardiomyocyte turnover, possibly from cardiac or vascular cells (3,4). Unlike other adult cell types such as stem cells (CSCs)/cardiac progenitor cells (CPCs) (1,2). bone marrow cells (BMCs), for which surface markers have In a genetic fate-mapping study, it was shown that CPCs been extensively characterised, resident CPCs show mixed contribute to the replenishment of adult mammalian and overlapping expression of stem cell markers. Several cardiomyocytes lost after injury, throughout adult life CPC populations have been reported in the developing (3,4). Numerous other studies have also identified and and adult heart including: c-Kit+ CPCs; cardiospheres/ isolated endogenous CPC populations in the hearts cardiosphere-derived cells (CDCs); epicardium-derived from multiple species including mouse, rat, dog, porcine cells (EPDCs); cardiac side population cells (CSPCs, and human (5–10). These data suggest that CPCs are identified by their ability to exclude Hoechst dye (8)); capable of differentiating into multiple cell types, such Stem cell antigen-1 (Sca-1+) CPCs; Islet-1 (Isl-1+) expressing as cardiomyocytes (CM), vascular smooth muscle cells CPCs and PDGFRa+ expressing CPCs, (interested readers (VSMCs), and endothelial cells (ECs). After transplantation are directed to other reviews for further details (11,12)). of CPCs, reduction in the infarct area and improvement A timeline for major events in studies of cardiac stem/ of left ventricular (LV) function were demonstrated in progenitor cells is shown in Fig. 1. Although considerable

Fig. 1. Timeline for major events in studies of the cardiac stem/ progenitor cells. CSC cardiac stem cell, CDC cardiosphere-derived cell, CPC cardiac progenitor cell, CSPC cardiac side population cell, c-CFU-F cardiac colony-forming unit fibroblast, CADUCEUS CArdiosphere-Derived aUtologous Stem CElls to Reverse ventricUlar dySfunction, SCIPIO Stem Cell Infusion in Patients with Ischemic cardiomyopathy. Randomised trials of CPCs as therapy are indicated in blue.

Vol 47 No 1 April 2016 AUSTRALIAN BIOCHEMIST Page 7 Repairing the Damaged Heart SHOWCASE ON with Cardiac Progenitor Cells RESEARCH overlap between these populations is likely, there are potential in vivo. However, recent evidence has suggested insufficient data to specifically address this possibility. that Isl-1 is not a marker of endogenous CPCs (20). CPCs were first identified in 2003 through the expression Furthermore, using genetic fate-mapping approaches, of the tyrosine kinase receptor, c-Kit and the absence of Isl-1 has been shown to mark not only progenitors from common hematopoietic lineage markers (such as CD45-, the second heart field, but also from the cardiac neural Lin-) in the adult mammalian heart (13). c-Kit+ cardiac crest (21). Recent reports suggest that resident cardiac cells possess prolonged self-renewing, clonogenic and c-Kit+ cells in the mouse are not a source of myocytes multipotent characteristics. Encouraging results whereby but are predominantly a source of endothelial cells after c-Kit+ CPCs improved LV dysfunction and remodelling in cardiac injury (22,23). Therefore, as previously mentioned, various pre-clinical models of post-MI cardiomyopathy the exact biological role of these various progenitor have paved the way for Cardiac Stem cell Infusion in populations in the injured heart remains unknown. Patients with Ischemic Cardiomyopathy (SCIPIO), the We recently demonstrated that expression of the first clinical trial of CSCs. Here, c-Kit+ cells were isolated tyrosine kinase Platelet-derived Growth Factor Receptor- from the right atrial appendage of patients undergoing alpha (PDGFRa+) identifies a resident cardiac progenitor open heart surgery for coronary artery bypass grafting population (named colony-forming unit fibroblasts; (CABG). Harvested CPCs underwent expansion in vitro c-CFU-Fs) in the murine (6) and human heart (24). These and were then infused back into the donor heart via the cells possess prolonged self-renewal and multipotent coronary arterial circulation. Although the SCIPIO results potential in vitro. CRE lineage-tracing studies suggest a appeared to support the preclinical work by the same proepicardial/epicardial origin for c-CFU-Fs (6). Both group (improvement in LV systolic function and reduced cardiac fibroblasts and c-CFU-Fs originate from the infarct size) (14,15), it is important to note that editors of proepicardium, and undergo epithelial-mesenchymal the prestigious Lancet journal have taken the unusual step transition (EMT) before populating the subepicardium of expressing concern over the integrity of data published and myocardial interstitium, where they adopt a in the SCIPIO trial (16). perivascular location (6). What defines the cardiac Another well-characterised CPC population is the fibroblast pool is currently poorly understood (25). cardiosphere-derived cell (CDC) population. CDCs are a Under specific conditions, c-CFU-Fs are able to give rise mixture of stromal, mesenchymal and progenitor cells that to vascular cell types, differentiated fibroblasts and to a are derived from cultures of atrial or ventricular biopsy much more limited degree, cardiomyocyte-like cells. We specimens. When cloned in suspension, they develop are currently continuing further studies in rodent models multicellular clusters known as cardiospheres (17). From of cardiac injury that will lead to a better understanding of these cardiospheres, millions of proliferative cells that the pathophysiological role of these progenitors. express markers of mesenchymal and progenitor cell- related antigens can be harvested (17). In vitro CDCs are Potential Mechanisms of Cardiac Progenitor Cell clonogenic and have multilineage potential. The safety and Action in the Damaged Heart efficacy of CDC transplantation has also been demonstrated Implantation of CPCs in the injured heart may lead to in pre-clinical studies (10). In a murine model of MI, the myocardial repair via direct and indirect mechanisms functional outcome of CDC transplantation was superior (Fig. 2). These include transplanted cells that directly (in terms of ischaemic tissue preservation, anti-remodelling transdifferentiate into cardiomyocytes and vascular effects and functional benefits) to the transplantation of cells, paracrine effects inducing hyperplasia of resident bone marrow mesenchymal stromal cells (BM-MSCs) and cardiomyocytes, induction of resident endogenous CPCs adipose-derived regenerative cells (ADRCs) (18). In the to myocardial differentiation and cell fusion between CArdiosphere-Derived aUtologous stem CElls to reverse transplanted cells and resident cardiomyocytes. After many ventricUlar dySfunction (CADUCEUS) study, patients years of pre-clinical and clinical studies, the current view with acute MI were randomised to receive standard is that the positive effects of adult stem cell therapy (such medical therapy or autologous CDCs (19). This phase I as BMC and adipose-derived cells) is mediated through randomised clinical trial showed no difference between paracrine release of anti-apoptotic, immunomodulatory, treatment groups in terms of left ventricular ejection proangiogenic host- and cell-derived factors (26). Therefore, fraction (LVEF)-change after six months (19). However, the moderate positive effects of adult stem cell delivery infarct size was significantly reduced in the cell treated to impaired myocardium is more likely due to enhanced group. Patients treated with CDCs showed a reduction neovascularisation or favourable changes in the cardiac in scar mass, increased viable heart mass and improved scar (which is itself contractile and not inert) rather than the regional contractility (19). formation of new cardiomyocytes. Cell fusion between the Several other CPC populations have also been identified transplanted cells and resident myocytes or direct trans- and characterised using different membrane markers (Sca- differentiation into cardiomyocytes (as robustly seen in 1, Abcg-2, Flk-1, CD34, CD90, CD105) and transcription current pluripotent stem cell differentiation protocols) is factors (Isl-1, Nkx2.5, MEF2C, GATA4) in the embryonic considered unlikely. and adult heart. Again, these cells are clonogenic, self- Many of the pre-clinical studies and the SCIPIO clinical renewing and multipotent both in vitro and in vivo. trial (discussed above) have used c-Kit+ cells as the primary They express several markers characteristic of stem cells source of cardiac regeneration after injury. Furthermore, (Oct3/4, Bmi-1, Nanog) and have significant regenerative recent reports have used powerful genetic fate mapping

Page 8 AUSTRALIAN BIOCHEMIST Vol 47 No 1 April 2016 SHOWCASE ON Repairing the Damaged Heart RESEARCH with Cardiac Progenitor Cells experiments to show cardiomyocyte formation from c-Kit+ progenitors in a rodent catecholamine-induced injury model (27). However, van Berlo and colleagues recently used rigorous genetic fate mapping experiments to show that minimal cardiomyocytes were generated from c-Kit+ cells in vivo (22). In contrast, abundant cardiac endothelial cells appeared to be derived from c-Kit+ CPCs (22,23). Taken together, these new findings suggest that improvements in cardiac function after injury may be due to c-Kit+ CPC vascularisation of the injured heart and subsequent favourable effects on hibernating myocardium rather than the generation of new cardiomyocytes.

Future Directions Despite the publication of Fig. 2. Potential mechanisms of action of transplanted cardiac progenitor cells. several studies in different CM cardiomyocyte, CDC cardiosphere-derived cell, CPC cardiac progenitor species, the physiological and cell, CSPC cardiac side population cell, EC endothelial cell, EPDC epicardium pathophysiological functions of derived cell, VSMC vascular smooth muscle cell. the various CPC populations in the heart remain unclear. Utilisation of CPCs (either J.J.H. (2015) Biophys. Rev. 7, 127-139 exogenously delivered or harnessing CPCs residing within 12. Chong, J.J., Forte, E., and Harvey, R.P. (2014) Stem Cell the injured heart) has repeatedly been shown to induce Res. 13, 592-614 favourable reparative or regenerative effects. Nevertheless 13. Beltrami, A.P., Barlucchi, L., Torella, D., et al. (2003) Cell the molecular mechanisms underpinning such changes 114, 763-776 are not completely understood. Thus, the application of 14. Bolli, R., Chugh, A.R., D’Amario, D., et al. (2011) Lancet CPCs as a clinical treatment for cardiovascular disease will 378, 1847-1857 remain difficult until these limitations are appropriately 15. Bolli, R., Tang, X.L., Sanganalmath, S.K., et al. (2013) addressed. In addition, attention should be focused on Circulation 128, 122-131 gaining a better understanding of the cardiac fibroblast 16. The Lancet Editors (2014) Lancet 383, 1279 population, which is known to aid repair and regeneration 17. Messina, E., De Angelis, L., Frati, G., et al. (2004) Circ. Res. of cardiomyocytes in addition to producing cardiac scar. 95, 911-921 18. Li, T.S., Cheng, K., Malliaras, K., et al. (2012) J. Am. Coll. References Cardiol. 59, 942-953 1. Anversa, P., and Kajstura, J. (1998) Circ. Res. 83, 1-14 19. Makkar, R.R., Smith, R.R., Cheng, K., et al. (2012) Lancet 2. Bergmann, O., Bhardwaj, R.D., Bernard, S., et al. (2009) 379, 895-904 Science 324, 98-102 20. Weinberger, F., Mehrkens, D., Friedrich, F.W., et al. 3. Hsieh, P.C., Segers, V.F., Davis, M.E., et al. (2007) Nat. (2012) Circ. Res. 110, 1303-1310 Med. 13, 970-974 21. Engleka, K.A., Manderfield, L.J., Brust, R.D., Li, L., 4. Senyo, S.E., Steinhauser, M.L., Pizzimenti, C.L., et al. Cohen, A., Dymecki, S.M., and Epstein, J.A. (2012) Circ. (2013) Nature 493, 433-436 Res. 110, 922-926 5. Bearzi, C., Rota, M., Hosoda, T., et al. (2007) Proc. Natl. 22. van Berlo, J.H., Kanisicak, O., Maillet, M., Vagnozzi, R. Acad. Sci. USA 104, 14068-14073 J., Karch, J., Lin, S.C., Middleton, R.C., Marban, E., and 6. Chong, J.J.H., Chandrakanthan, V., Xaymardan, M., et Molkentin, J.D. (2014) Nature 509, 337-341 al. (2011) Cell Stem Cell 9, 527-540 23. Sultana, N., Zhang, L., Yan, J., et al. (2015) Nat. Commun. 7. Oh, H., Bradfute, S.B., Gallardo, T.D., et al. (2003) Proc. 6, 8701 Natl. Acad. Sci. USA 100, 12313-12318 24. Chong, J.J.H., Reinecke, H., Iwata, M., et al. (2013) Stem 8. Martin, C.M., Meeson, A.P., Robertson, S.M., et al. (2004) Cells Dev. 22, 1932-1943 Dev. Biol. 265, 262-275 25. Furtado, M.B., Costa, M.W., Pranoto, E.A., et al. (2014) 9. Laugwitz, K.L., Moretti, A., Lam, J., et al. (2005) Nature Circ. Res. 114, 1422-1434 433, 647-653 26. Gnecchi, M., Zhang, Z., Ni, A., and Dzau, V.J. (2008) 10. Johnston, P.V., Sasano, T., Mills, K., et al. (2009) Circ. Res. 103, 1204-1219 Circulation 120, 1075-1083 27. Ellison, G.M., Vicinanza, C., Smith, A.J., et al. (2013) Cell 11. Ge, Z., Lai, S., Le, T.Y.L., dos Remedios, C., and Chong, 154, 827-842 Vol 47 No 1 April 2016 AUSTRALIAN BIOCHEMIST Page 9 SHOWCASE ON RESEARCH

Defining Prostate Stem Cells: Clues to Improving Prostate Cancer Treatment Renea Taylor* Biomedicine Discovery Institute, Department of Physiology, Monash University, Clayton, VIC 3800 *Corresponding author: [email protected]

Introduction cells showed an activation of stem-like genes specifically Prostate cancer is one of the leading causes of death in associated with a set of genes targeted by the E2F family Australian men. Despite the clinical significance of this of transcription factors (10). Nevertheless, a rate-limiting disease, our understanding of the cellular origins of prostate step for basal cells in the initiation of prostate cancer is that cancer and the cells within tumours that propagate growth they must differentiate into luminal cells, demonstrating and survival are relatively limited. Many laboratories the importance of the luminal phenotype in prostate cancer have sought to isolate prostate stem cells to understand progression. their hormone-dependent and -independent growth and As such, the recent focus in the field has been on differentiation, with the hope that this will ultimately lead subpopulations of luminal cells in the normal prostate gland to the development of new therapies for prostate cancer. that may contribute to cancer initiation and propagation. Recent evidence supports the concept that there are Luminal cells have been traditionally regarded as multiple cells from which prostate cancer can arise, and that terminally differentiated cells with secretory functions and features of the molecular signature of normal stem cells can thus were unlikely to possess stem/progenitor activity. be detected in prostate cancer tissues. More importantly, However, a recent study identified a distinct population of there are regenerating cells within tumours that are luminal epithelial progenitor cells in mice, marked by Sca- resistant to hormonal therapy and continue to propagate 1 expression, that can propagate multilineage organoids tumour progression. One of the most important challenges in culture and in an in vivo prostate regeneration assay in cancer biology is to understand the cell populations (11). Importantly, this luminal cell population can survive that drive tumour growth and how they can be effectively androgen deprivation, one of the hallmarks of prostatic targeted. stem cells. The study provides new evidence that a Sca1+ fraction of prostatic luminal cells may be the preferred cell Epithelial Stem Cells in the Prostate of origin of prostate cancer, although an equivalent human The prostate gland is a relatively slow growing, androgen- population is yet to be identified. dependent organ. In normal prostatic epithelium, there are three primary cell types: luminal secretory cells, the Stem/Progenitor Cells in Prostate Cancer underlying basal support cells, and rare neuroendocrine Androgen-deprivation therapy is the mainstay treatment cells (Fig. 1). Lineage tracing studies consistently for advanced prostate cancer and relies on the exquisite demonstrate that prostate basal cells and luminal cells sensitivity of the prostate to androgens. Clinically, most are independently sustained in adult mice, but may give tumours respond and up to 95% of the cancer bulk is rise to other cell types under certain conditions (1–3). The removed. However, tumour regrowth is inevitable, leading advances in 3D organoid culture have provided conditions to a lethal form of the disease referred to as castration- under which multiple prostate cell types can survive and resistant prostate cancer (Fig. 1). The scientific goal is to proliferate (4,5). Studies using this approach have provided identify the prostate cancer cells that survive androgen- further support of the existence of respective multipotent deprivation therapy, because this population offers the stem cells or progenitors within both the basal and luminal potential for disease progression. The ability to eliminate cell lineages (3,6,7). The discovery of multiple stem cells in these cells therapeutically will lead to increased survival normal tissue has expanded the intrigue surrounding the for men with prostate cancer. cell(s) of origin of prostate cancer. Using a human patient-derived xenograft model (12), our laboratory identified a subpopulation of luminal prostate Cells of Origin of Prostate Cancer cancer cells that survive androgen withdrawal in vivo Prostate adenocarcinomas represent the vast majority of (13). The subset of human cancer cells that survive in the prostate cancer and are composed of cells with a luminal androgen-depleted environment show stem-like features phenotype, largely devoid of basal cells, Thus it was long including expression of the stem cell markers Oct-4, Sox2, presumed that prostate cancer is derived from a luminal NANOG and ALDH1, as well as the ability to actively cell within the normal epithelium. However, it is clear that repopulate tumours upon androgen readministration. basal cells also can be genetically transformed to give rise to The demonstration that a population of ‘castrate-tolerant’ prostate tumours (3,8,9). In fact, comprehensive molecular cancer cells pre-exist in hormone-dependent prostate studies on sorted populations of human prostate cancer cells cancer, prior to hormonal treatment, was a major discovery. showed that the transcriptional profiles of aggressive forms The finding now provides an opportunity to determine of prostate cancer share common features with human the mechanisms by which they survive and continue to basal cells. Specifically, metastatic human prostate cancer propagate tumour progression.

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Complementary studies in a Pten/TP53-null mouse advanced castrate-resistant metastatic prostate cancer model of prostate cancer revealed a luminal progenitor specimens, where continuously propagated organoid population in prostate tumours (14). Agarwal and lines were established from metastatic biopsies at a rate colleagues used organoid cultures to demonstrate tumour- of 15%–20%. Whilst this method has proven successful initiating activity that exists in both the luminal and basal for advanced tumours, adaptation to organ-confined, fractions. Using FACS cell sorting strategies, two distinct hormone-dependent tumours is yet to be optimised. PROM1+ mouse luminal progenitors were identified; a Nonetheless, the approach will provide a reliable in vitro minor population within mouse tumours that gives rise culture system for high throughput drug screening of to multilineage organoids (multipotent progenitors) and clinical samples with significant molecular and phenotypic a major population producing luminal-only organoids heterogeneity. Advances in fractionating cell populations (luminal committed progenitors). Similar to the findings from within tumours will also generate new knowledge from castration of human patient-derived xenografts, a about the role of stem cells in propagating prostate cancer, significant proportion of mouse luminal progenitor cells and provide a new approach for testing novel therapeutic survived in vivo castration. A comparison of the molecular agents. features that are shared between human and mouse luminal progenitor cells within tumours may reveal key Conclusions and Future Directions signalling pathways that offer new therapeutic potential. The prostate epithelial hierarchy is complex, with A major advance in the field was the adaptation of the multiple stem/progenitor cells residing in both the basal mouse 3D organoid culture technique to human prostate and luminal lineages. The nature of the perturbations in cancer tissues. Gao and colleagues successfully established these stem cells remains largely unknown, and therefore long-term (more than six months) cultures of patient- the implications for disease are still not clear. Despite the derived prostate tumour organoids from dissociated cells best efforts of researchers, the molecular profiles of prostate taken from metastatic biopsy specimens and circulating epithelial stem cells have not been definitively characterised tumour cells (4). This technique is relatively efficient for at the single cell level. In prostate cancer, there is an

Epithelium Normal prostate

Prostate cancer Basal and luminal epithelial cells can act as cells of origin of prostate cancer

Models to study prostate cancer stem cells: 1. Patient-derived xenografts (in vivo) Tumour 2. Patient-derived organoids (in vitro)

Androgen withdrawal reveals ‘castrate-tolerant’ prostate cancer cells

Fig. 1. In the normal prostate gland, the epithelium is composed of luminal secretory cells (blue), basal cells (red) and rare neuroendocrine cells (green). Multipotent stem cells have been reported in basal and luminal populations. In addition, each of these populations has the potential to give rise to prostate adenocarcinomas, thereby acting as cells of origin of prostate cancer. Recent advances in studying human prostate cancer using patient-derived xenografts or patient-derived organoids have revealed key subpopulations of cancer cells that can propagate tumours in vivo and in vitro. Using these approaches, a subset of prostate cancer cells (red nuclei) have been identified that are therapy resistant (i.e. survive androgen withdrawal therapy) and repopulate tumours given growth stimuli. These ‘castrate-tolerant’ prostate cancer cells are the focus of new therapeutic targets that may be effective in preventing progression to lethal prostate cancer.

Vol 47 No 1 April 2016 AUSTRALIAN BIOCHEMIST Page 11 Defining Prostate Stem Cells: Clues to SHOWCASE ON Improving Prostate Cancer Treatment RESEARCH important subpopulation of tumour cells that can survive 4. Gao, D., Vela, I., Sboner, A., Iaquinta, P.J., Karthaus, androgen withdrawal and potentially lead to incurable W.R., Gopalan, A., et al. (2014) Cell 159, 176-187 prostate cancer. Until now, a deeper understanding of the 5. Drost, J., Karthaus, W.R., Gao, D., Driehuis, E., Sawyers, biology of these castrate-tolerant cells has been hindered C.L., Chen, Y., et al. (2016) Nat. Protoc. 11, 347-358 by a lack of biomarkers and cell surface markers that 6. Chua, C.W., Shibata, M., Lei, M., Toivanen, R., Barlow, can be used to prospectively isolate them from human L.J., Bergren, S.K., et al. (2014) Nat. Cell Biol. 16, 951-961 prostate cancer tissue and potentially apply them to novel 7. Karthaus, W.R., Iaquinta, P.J., Drost, J., Gracanin, A., van platforms such as patient-derived organoids. Whether Boxtel, R., Wongvipat, J., et al. (2014) Cell 159, 163-175 or not the cells that survive castration are bona fide stem 8. Xin, L., Lawson, D.A., and Witte, O.N. (2005) Proc. Natl. cells that exist within tumours, or a transient population Acad. Sci. USA 102, 6942-6947 that is induced in response to androgen withdrawal is 9. Goldstein, A.S., Huang, J., Guo, C., Garraway, I.P., and yet to be determined. Nonetheless, this subpopulation of Witte, O.N. (2010) Science 329, 568-571 cancer cells should be considered as a cellular target for 10. Smith, B.A., Sokolov, A., Uzunangelov, V., Baertsch, R., prostate cancer therapeutics, perhaps in combination with Newton, Y., Graim, K., et al. (2015) Proc. Natl. Acad. Sci. androgen-deprivation therapy to maximise the control of USA 112, E6544-E6552 cancer progression. 11. Kwon, O.J., Zhang, L., and Xin, L. (2016) Stem Cells 34 , 191-202 References 12. Lawrence, M.G., Taylor, R.A., Toivanen, R., Pedersen, 1. Liu, J., Pascal, L.E., Isharwal, S., Metzger, D., Ramos J., Norden, S., Pook, D.W., et al. (2013) Nat. Protoc. 8, 836- Garcia, R., Pilch, J., et al. (2011) Mol. Endocrinol. 25, 1849- 848 1857 13. Toivanen, R., Frydenberg, M., Murphy, D., Pedersen, 2. Lu, T.L., Huang, Y.F., You, L.R., Chao, N.C., Su, F.Y., J., Ryan, A., Pook, D., et al. (2013) Sci. Transl. Med. 5, Chang, J.L., et al. (2013) Am. J. Pathol. 182, 975-991 187ra71 3. Wang, Z.A., Mitrofanova, A., Bergren, S.K., Abate-Shen, 14. Agarwal, S., Hynes, P.G., Tillman, H.S., Lake, R., Abou- C., Cardiff, R.D., Califano, A., et al. (2013) Nat. Cell Biol. Kheir, W.G., Fang, L., et al. (2015) Cell Rep. 13, 2147-2158 15, 274-283

ELECTION OF COUNCIL 2017 Nominations are called for the following positions on the Council of the Australian Society for Biochemistry and Molecular Biology Inc for 2017: Secretary, Treasurer, Editor, Secretary for Sustaining Members and State Representatives for ACT, NSW, Qld, SA, Tas, Vic and WA.

The Council for the President M. Ryan period 1 January 2016 President Elect L. Tilley to 31 December 2016 Secretary B. Forbes# is composed of the Treasurer T. Piva# following members: Editor C.K. Liew# Education Representative J. Macaulay# Secretary for Sustaining Members S. Jay# Representatives for: ACT Y.P. Mabbitt# NSW K. Michie# Vic D. Stojanovski* * Retiring member, not Qld D. Ng# eligible for re-election SA S. Polyak# # Eligible for re-election Tas A. Holloway* WA N. Taylor#

Nomination forms are available on the ASBMB website. Nominations for all vacant positions must be signed and seconded by members of the Society. The nominations must be signed by the nominee to indicate his/her willingness to stand. If more than one nomination is received for any position, a ballot will be held to determine the successful candidate. All members may vote for all positions except those of State Representatives where election is by members in the State concerned.

NOMINATIONS MUST REACH THE SECRETARY 19 SEPTEMBER 2016 (14 DAYS BEFORE THE ANNUAL GENERAL MEETING TO BE HELD 3 OCTOBER 2016)

Page 12 AUSTRALIAN BIOCHEMIST Vol 47 No 1 April 2016 SHOWCASE ON RESEARCH

Generating Kidney Tissue from Pluripotent Stem Cells Melissa Little1,2* 1Murdoch Childrens Research Institute, Parkville, VIC 3052 2Department of Pediatrics, University of Melbourne, VIC 3010 *Corresponding author: [email protected]

Introduction mesoderm and forms through interactions between two A pluripotent stem cell is, by definition, capable of major cell types; an epithelial duct called the ureteric differentiating into all possible cell types. The pluripotent bud that forms the collecting ducts required for urine to state within a developing embryo includes the inner cell leave the kidney and a metanephric mesenchyme, which mass of the preimplantation embryo and the subsequently gives rise to all the different cell types of the epithelial derived epiblast. While pluripotency in the embryo is filtration units called nephrons (3,4). Our understanding regarded as a transient state, the isolation and continuous of the formation of these cell types in other organisms, culture of such cells in a pluripotent state has been possible particularly the mouse, has guided protocols for the in mice for more than three decades. The derivation of the directed differentiation of human pluripotent stem cells first such human pluripotent stem cell line in 1998 (1) and to kidney cell types. Some of the earliest approaches for the subsequent identification of key transcription factors directing differentiation of stem cells to kidney cells used a able to convert an adult somatic cell into an equivalent combination of growth factors known to be important either pluripotent state (2), the induced pluripotent stem cell for early kidney formation or the specification of individual (iPSC), has totally transformed stem cell biology, opening kidney cell types, to look for differentiation into these cell the door for many options in regenerative medicine. It is types. In this way, groups have reported the formation of easy to understand how pivotal these findings have been, proximal tubular epithelium (5) and glomerular podocytes given the fact that in theory, you should now be able to (6) from human embryonic stem cells. Other approaches make a renewable, expandable and patient-specific stem have more systematically monitored the progression of cell that can be directed to form the required cell type for the differentiating cells in culture through intermediate treatment, which can then be delivered back to the patient stages of development, including the primitive streak, as an autologous treatment (Fig. 1). This is the potential intermediate mesoderm and into either the collecting of induced pluripotent stem cells. In reality, there remain duct epithelium (7) or nephrogenic mesenchyme (8-11). many obstacles, not least of all the ability to direct the Not surprisingly, most of these approaches have focussed differentiation of these stem cells to the cell type that is on the addition of similar recombinant growth factors desired. While early progress has been made with respect or small agonists, with an initial induction of primitive to in vitro directed differentiation of pluripotent stem cells streak usually involving canonical Wnt signalling and/ to ectodermal endpoints (particularly specific neuronal or activin/nodal and BMP signalling. This is followed by subtypes), the generation of kidney cell types (derivatives the addition of a fibroblast growth factor (FGF; either FGF2 of the intermediate mesoderm) has been slower to or FGF9) and frequently the inclusion of the signalling come. However, the last two years have seen substantial protein BMP7 to support the formation of the nephrogenic advances. mesenchyme. Some approaches have generated these tissues from monolayers of starting cells (7,10), while others Directing Differentiation to Kidney have used embryoid body culture or even formation of Like the muscles and the blood, the kidney is derived an intermediate epiblast stage within matrigel (9,12). The from the definitive mesoderm of the embryo (3). More more convergence of differentiation protocols for the formation specifically, it arises like the gonad, from the intermediate of early nephrons is striking in that a number of groups

Fig. 1. Potential applications for directing the differentiation of human pluripotent stem cells into kidney cells (adapted from 25).

Vol 47 No 1 April 2016 AUSTRALIAN BIOCHEMIST Page 13 Generating Kidney Tissue SHOWCASE ON from Pluripotent Stem Cells RESEARCH now show clear evidence for the formation of segmented agreement with this, an unbiased comparison of the and patterned nephrons with individual segments expression profile of organoids with that of human fetal showing clear functional differentiation into the proximal tissue indicates kidney organoids most closely resemble tubule, loop of Henle, distal tubule and the epithelial cell trimester 1 human kidney (16). What remains to be types of the glomeruli (9–14). Transplantation of such determined is how mature such in vitro organoids can structures under the kidney capsule of a recipient animal become. have also been demonstrated to facilitate vascularisation of the glomeruli as would normally occur during nephron formation (12,14).

Formation of Complex Multicellular Kidney Organoids from Human Pluripotent Cells As efforts to enable differentiation of pluripotent stem cells often attempt to replicate development, one might think that within a dish you might get multiple cellular outcomes rather than a single endpoint. This is in fact the case. Studies over the last few years have demonstrated the formation of complex multicellular organoids comprised of interacting cell types arranged in an organotypic fashion. In this way, human pluripotent stem cells have been shown to form organoids of the developing eye (optic cup), the cerebral cortex, stomach and intestine (reviewed in 15). In each of these cases, the progenitors of the organ of interest self-organise in three dimensions as they might during normal development, to form a model of the organ. We have recently demonstrated the formation of kidney organoids after the in vitro directed differentiation of human iPSCs cultured as a pellet at an Fig. 2. A kidney organoid generated from a human air media interface (16) (Fig. 2). This is the same culture pluripotent stem cell line (16). method that has long been applied to the ex vivo culture This organoid has been cultured for 18 days as of mouse embryonic kidneys (17). Indeed, it is possible an aggregate, post-induction of intermediate to completely dissociate an embryonic mouse kidney, mesoderm. Immunofluroescence indicates the reform an aggregate of the component cells and have the presence of differentiating nephrons comprised of cells self-organise to reform the epithelial elements of the glomeruli (NPHS1, yellow), proximal tubules (LTL, original organ (18,19). Presumably, therefore, the culture pink) and distal tubules/collecting ducts (CDH1, of the differentiated iPSCs in this format facilitates a green). Image by Minoru Takasato. similar self-organising environment. Within these human kidney organoids, there is evidence of more than 8 distinct cell types, including the formation of appropriately Nephrotoxicity Screening segmenting nephrons comprised of distal tubule, loop With such progress in the generation of kidney cell of Henle, proximal tubule and Bowman’s capsules types, the options for use of these cells has significantly containing parietal epithelial cells and podocytes (Fig. 2). widened (Fig. 1). One early application has been the Simultaneously, the collecting duct epithelium forms and evaluation of iPSC-derived kidney cells for the screening connects to the nephrons. Surrounding these epithelial of drugs to evaluate nephrotoxicity. The use of pre-clinical elements there is a stromal population that expresses pharmaceutical exposure is currently the Gold standard key transcription factors known to mark the cortical for nephrotoxicity screening. However, the mouse does stroma of the developing kidney, including Meis1. More not always predict outcome in human. The use of in vitro surprisingly, an extensive endothelial capillary network drug screening has been attempted using primary or arises, with an accompanying perivascular compartment. immortalised proximal tubule epithelial cells, the principle There is even evidence that a subset of the glomeruli cell type targeted by nephrotoxicants. However, accepted begin to draw in these endothelial and pericytic in vivo biomarkers do not appear to be induced in such progenitors to form the glomerular capillaries. The origin screens and hence these are of low predictive value. The of the vasculature of the kidney has long been proposed use of iPSC-derived proximal tubule cells has now been to include both vasculogenic and angiogenic progenitors, evaluated (20) with results suggesting greater accuracy in however the origin of each of these progenitors types predicting toxicity than primary human proximal tubule has been unclear. The presence of vasculature in such cells (20). However, these were not evaluated for the kidney organoids suggests that at least the vasculogenic induction of Kim1, the biomarker most widely regarded component also arises from the intermediate mesoderm, as an early and accurate predictor of nephrotoxic injury. as does the nephron progenitor population (16). The With the generation of protocols for the creation of more histological features of kidney organoids appear to complex kidney structures, including nephrons and represent relatively early kidney development. In whole organoids, has come early evidence that these Page 14 AUSTRALIAN BIOCHEMIST Vol 47 No 1 April 2016 SHOWCASE ON Generating Kidney Tissue RESEARCH from Pluripotent Stem Cells respond to known nephrotoxicants via either specific fluids, amino acids, electrolytes and other nutrients proximal tubular apoptosis or increased production of requires a particular histological topology as well as an Kim1 protein (12,13,16). Hence, these endpoints may also integrated collecting duct network with a viable exit path act as viable screens for nephrotoxic injury. to the bladder. This has not yet been achieved. Hence, generating an entire functionally competent replacement Disease Modelling Using Patient-derived Stem Cells organ remains a major challenge. One possible approach While the majority of chronic kidney disease in adults is using iPSC-derived kidney cells might be the delivery of accepted as the consequence of accumulated insult, it is these cells back into decellularised scaffolds generated estimated that 50% of children reaching end stage renal from human kidneys. Certainly, approaches for the failure have an inherited form of kidney disease. The de- and recellularisation of human kidney have been most common genetic cause of renal failure is autosomal reported (21). It is possible that the generation of specific dominant polycystic kidney disease (ADPKD). However, renal cell types for delivery back into the injured kidney there are many other heritable cystic kidney diseases – cellular therapy – may also provide some value. Early (nephronophthisis, tubulointerstitial kidney disease), data suggests that this might be the case, with the delivery glomerulopathies and tubulopathies. Improvements in of human-derived renal progenitors into mouse models Next Generation Sequencing over the last five years has of injury showing evidence of reduced damage (22, 23). led to the identification of many novel gene mutations However, it remains to be seen whether such cells can in such conditions. However, as with many other functionally integrate long term and whether they will do diseases, there is significant variation in penetrance so in the face of chronic renal injury. What may be more and expressivity within such families, likely due to feasible is the use of kidney cell types generated from iPSC accompanying variations in their genome. This makes the in microfluidics-based organ-on-a-chip technology (24). validation of any novel gene variant challenging. With the Such options remain in the future. However, identifying newly developed protocols for directed differentiation of the methodology to move a pluripotent stem cell state iPSCs, it is now feasible to generate patient-specific lines to a kidney endpoint has opened the door to all of these for disease modelling and functional genomics (Fig. 1). possibilities. It also opens the door, for the first time, to The capacity to scale up directed differentiation protocols a better understanding of the molecular basis of normal may also facilitate patient-specific drug screening for kidney development in the human and it is here that there the identification of new treatments. The first reports is much to be learned. illustrating the proof of concept of kidney disease modelling have come with the demonstration of cystic References epithelia within kidney tissue derived from an ADPKD 1. Thomson, J.A., Itskovitz-Eldor, J., Shapiro, S.S., et al. iPSC line (12). There remain many hurdles to these studies. (1998) Science 282, 1145-1147 The identification of genuinely disease-specific changes 2. Takahashi K., Tanebe, K., Ohnuki, M., et al. (2007) Cell in gene expression or developmental potential in vitro 131, 861–872 will need to be carefully distinguished from differences 3. Takasato, M., and Little, M.H. (2015) Development arising due to variations between individual iPSC clones 142,1937-1947 and even between individual differentiation experiments 4. Little, M.H., and McMahon, A.P. (2012) Cold Spring (technical variation). It will also be incredibly important Harb. Perspect. Biol. 4, a008300 to ensure that any ‘control’ comparison is performed 5. Narayanan, K., Schumacher, K.M., Tasnim, F., et al. with lines from a closely related individual, preferably an (2013) Kidney Int. 83, 593-603 isogenic clone from the patient themselves, corrected for 6. Song, B., Smink, A.M., Jones, C.V., et al. (2012) PLoS the mutation of interest. The advent of efficient and more One 7, e46453 accurate gene editing technologies, such as CRISPR, are 7. Xia, Y., Nivet, E., Sancho-Martinez, I., et al. (2013) Nat. making this possible. Cell Biol. 15, 1507-1515 8. Mae, S., Shono, A., Shiota, F., et al. (2013) Nat. Options for Regenerative Medicine: Future Directions Commun. 4, 1367 While cultures of kidney cells, or even the more complex 9. Taguchi, A., Kaku, Y., Ohmori, T., et al. (2014) Cell kidney organoids, are likely to deliver results with disease Stem Cell 14, 53-67 modelling and drug screening in the short term, the longer 10. Takasato, M., Er, P.X., Becroft, M., et al. (2014) Nat. Cell term goal of regenerative medicine is some way off. At Biol. 16, 118-126 present, kidney organoids reach approximately 8mm 11. Lam, A.Q., Freedman, B.S., Morizane, R., et al. (2014) J. in diameter after approximately three weeks in culture, Am. Soc. Nephrol. 25, 1211-1225 with each organoid containing up to 100 nephrons (16). 12. Freedman, B.S., Lam, A.Q., Sundsbak, J.L., et al. (2013) Each human kidney contains, on average, 1 million J. Am. Soc. Nephrol. 24, 1571-1586 nephrons (4). After renal failure, dialysis provides renal 13. Morizane, R., Lam, A.Q., Freedman, B.S., Kishi, function equivalent to 10–15% of glomerular function, S., Valerius, M.T., and Bonventre, J.V. (2015) Nat. but to even replicate this suggests the need to generate Biotechnol. 33, 1193-1200 a kidney replacement with >100,000 nephrons. The 14. Sharmin, S., Taguchi, A., Kaku, Y., et al. J. Am. Soc. challenge does not end with scaling up nephron number Nephrol. In press as the capacity of the kidney to appropriately reclaim 15. Ader, M., Tanaka, E.M. (2014) Curr. Opin. Cell Biol. 31, 23-28

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16. Takasato, M., Er, P., Chiu, H., et al. (2015) Nature 526, 21. Caralt, M., Uzarski, J.S., Iacob, S., Obergfell, K.P., et al. 564-568 (2015) Am. J. Transplant. 15, 64-75 17. Saxen, L., and Lehtonen, E. (1987) Differentiation 36, 22. Toyohara, T., Mae, S., Sueta, S., Inoue, T., et al. (2015) 2-11 Stem Cells Transl. Med. 4, 980-992 18. Unbekandt, M., and Davies, J.A. (2010) Kidney Int. 77, 23. Imberti, B., Tomasoni, S., Ciampi, O., et al. (2015) Sci. 407-416 Rep. 5, 8826 19. Hendry, C.E., Vanslambrouck, J.M., Ineson, J., et al. 24. Jang, K.J., Mehr, A.P., Hamilton, G.A., et al. (2013) (2013) J. Am. Soc. Nephrol. 24, 1424-1434 Integr. Biol. 5, 1119-1129 20. Li, Y., Kandasamy, K., Chuah, J.K., et al. (2014) Mol. 25. Takasato, M., Maier, B., and Little, M.H. (2014) Pediatr. Pharm. 11, 1982-1990 Nephrol. 29, 543-552

Sydney Protein Group: an ASBMB Special Interest Group

The Sydney Protein Group (SPG) consists of protein The SPG also offers several prestigious prizes aimed at scientists and students from academia, hospitals and PhD/Honours students: industry that meet regularly to hear updates from local (1) The annual Thompson Prize is in recognition of the and international speakers on the latest developments in eminent local protein scientist, E.O.P. (Ted) Thompson, protein science. The group started up in the early 1980s for his many contributions to the field of protein structure and today includes members from Woollongong and and function in Sydney from the 1950s until his retirement Canberra. The current organising committee includes Liza in 1990. The Thompson Prize was inaugurated in 1992 and Cubeddu (Western Sydney University) as President, Ben is awarded for the best oral presentation by a young local Crossett (University of Sydney) as Treasurer, and Jason scientist in the field of protein structure and function. Low (University of Sydney) as recently elected Secretary. The 2015 Thompson Prize was held at the Garvan Institutional representatives continue to function to Institute, hosted by Tara Christie. Jacob Lewis (University disseminate SPG information and increase the visibility of of Wollongong) kicked off the talks by describing his SPG events. efforts to characterise DNA polymerase dynamics at the E. coli replication fork using single molecule techniques. SPG Meetings and Early Career Researcher Support Quill Bowden (University of New South Wales) spoke The SPG organises and supports several annual events, about her use of fluorescence microscopy to visualise with the help of generous funding from the ASBMB and the dynamics and interactions of molecular chaperones dedicated trade partners. and alpha-synuclein and how these experiments can An important SPG ethos is to provide opportunities for help us understand the progress of neurodegenerative young researchers working in the field of protein science diseases where protein misfolding/aggregation occurs. to attend high-quality scientific meetings, where they can Next, Laura McCaughey (University of Technology hear about recent research, interact with guest speakers Sydney) described her PhD research done mainly at and other researchers, and present their own work. An the University of Glasgow, where she discovered and initiative of the SPG in recent years has been to host an characterised a new protein antibiotic from Pseudomonas Early Career Researcher (ECR) Symposium to choose the aeruginosa and its potential as a new target in the fight SPG-sponsored ECR for a guaranteed speaking slot at against drug resistant bacteria. Mana Liao (Garvan ComBio. In 2015, the SPG-sponsored ECR was chosen at Institute) introduced us to how modulation of hepatic the East Coast Protein Meeting (ECPM) at Coffs Harbour insulin action and lipid metabolism is affected by different in July. The ECPM is a joint initiative with the Queensland protein kinase C isoforms. Ruvini Kariawasam (Western Protein Group held every second year; the QPG organised Sydney University) gave us an interesting insight into another excellent meeting in 2015. The focus of this meeting the recognition of single-stranded DNA by simple was on ECRs, who present their work alongside a few single-stranded binding (SSB) proteins from archaea keynote speakers (Juliet Gerrard, University of Auckland; and humans, and how the structure/function of these Mark Kvansakul, La Trobe University; Jenny Martin, evolutionarily related proteins has been conserved. The Institute of Molecular Bioscience). At this meeting, the last speaker of the evening, Joshua Hamey (University SPG chose Alistair Edwards (University of Sydney) as the of New South Wales) told us about how his mass best ECR speaker, to represent the SPG at ComBio2015 in spectrometry approach led him to discover a novel protein Melbourne. Alastair talked about structural and functional methyltransferase that catalyses conserved N-terminal insights into the evolution and stress adaptation of type II and lysine methylation on translational elongation factor chaperonins. 1A. In the end, Laura McCaughey was awarded the 2015 Thompson Prize, but all the finalists should hold their

Page 16 AUSTRALIAN BIOCHEMIST Vol 47 No 1 April 2016 Sydney Protein Group: an ASBMB Special Interest Group heads high as they gave excellent presentations. Thanks to the judging panel consisting of Nick Dixon (University of Wollongong), Mitchell Guss (University of Sydney) and Kate Mitchie (University of New South Wales). Thanks Below: 2015 also to Thermofisher Scientific for their support of the Thompson Prize host Thompson Prize evening. and judges, from left: The SPG, along with ATA Scientific, also award three (2) Tara Christie, Kate annual Lorne Travelling Scholarships to help the best Mitchie, Nick Dixon and brightest students attend the Lorne conferences. The and Mitchell Guss. two Lorne Travel awards went to two University of New South Wales PhD students Daniel Winter (ATA Scientific Above: 2015 Thompson Travel Scholarship) and Joshua Hamey. The Greg Ralston Prize finalists, from Memorial Award, which helps an Honours student who is left: Jacob Lewis, Joshua continuing on to do a PhD attend Lorne, was awarded to Hamey, Quill Bowden, Athina Manakas (University of Sydney). Thanks to all the Ruvini Kariawasam, Laura students who applied for the Thompson Prize and Lorne McCaughey and Mana Liao. Scholarships; the standard of applications continues to be very high, so consider entering in 2016 if you were not successful this time around.

Website Liza Cubeddu, University of Western Sydney More information on SPG happenings can be found on Website: www.mmb.usyd.edu.au/spg/ the SPG website maintained by Roland Gamsjaeger. Short Discussions SDS Page for Students Page Supervisor types – which one is your match? The relationship with your postgraduate supervisor is the supervisor is never in the office. one of the most important ones you will have and will Advantages: Supervisor probably has a strong profile with be one of the major determinants in the success of your an international reputation whose name carries weight studies. It is essential that you both communicate well to when writing a reference. Make sure an absent supervisor build a good relationship to become a successful team. is not an unreliable one. There are a number of different types of supervisor and it Disadvantages: May not be there when you need them. is important that you have a good match – just as you may Also, frequent absence doesn’t necessarily correlate with opt for a certain type of partner – otherwise your journey success – make sure that the absent supervisor really is might not be as enjoyable as you want it to be. one who can give your career a boost by association. Compatible with: Someone who is independent and does Type 1: The Know-it-all not need much guidance. A person who is not afraid to ask Telltale signs: You are too familiar with the phrase: ‘I questions to other students, postdocs and co-supervisors. see your point of view but I think you should do the Also compatible with someone who takes the initiative following…’ to schedule face-to-face or online meetings to discuss the Advantages: They probably are a walking encyclopedia project. In this day and age, you don’t have to be both and have a lot of experience in your field. physically in the same place to have meetings. Disadvantages: Their way is not always the only right way or may not be the right way at all. Type 3: The Perfectionist Compatible with: A student who listens and takes on Telltale signs: Someone who criticises every aspect of feedback. your work. Someone who, while using Track Changes to provide you feedback, turns your document more red Type 2: The Absent Supervisor than black. Usually someone who is hard to impress. Telltale signs: Always invited to give seminars worldwide, Advantages: Will be able to help you achieve the best part of a million committees and even when on campus, results possible.

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Disadvantages: Can be overly critical and destroy Disadvantages: Don’t forget that your research should still confidence. be your main focus. Compatible with: Students who are also perfectionists. Compatible with: Someone who wants to have an all-round Students who would like a large amount of guidance experience during their studies. They want to be doing before developing their own independence. cutting edge research in the lab but also be involved in other activities to expand their skill set. A multitasker Type 4: The Very Hands-on Supervisor would be preferable. Telltale signs: You get calls/messages after hours from your supervisor. Your supervisor writes you emails late at There you have it. Which type is your perfect match? It night that need to be answered within 12 hours. You feel is important to find someone who is compatible because like they are always breathing down your neck. they are not only going to be your supervisor for a few Advantages: They tend to be very involved with every years while you are studying; they may also be your aspect of your project, which may be useful when you mentor for the rest of your life. Importantly, they will encounter any problems. probably be the first person to be called when you are Disadvantages: May stifle independence. Remember, applying for your dream job after you graduate. It is postgraduate studies are an evolving process towards critical to establish good working practices early and independence as a researcher, so by the end you need to manage your expectations. Make sure you are both on be in control. the same page about your milestones and direction of Compatible with: Someone who likes to have constant your project, and make necessary changes to the working guidance. Also suitable with a student who can set relationship if needed. But if you have managed to land boundaries and is not afraid to let the supervisor know of yourself a supervisor that is not your match, all is not lost. their working hours. Even though postgraduate studies Most students have more than one supervisor these days are not a 9 to 5 job, you should still be entitled to some for a reason, utilise them! You can also get useful guidance time off. Remind your supervisor that you have a life and mentorship from other people in your circle. There outside of work. may be a co-supervisor, a postdoc or a more senior lab member that you can draw upon for advice/guidance. Type 5: The Pessimist And remember that sometimes opposites attract! Telltale signs: Supervisor starts every meeting with ‘This will probably not work but you should try it anyway’. The Student’s Page is coordinated by Dr Tatiana Soares da Advantages: At least they are not going to be disappointed Costa, who is an NHMRC Early Career Fellow at the La if something does not go according to plan. They certainly Trobe Institute for Molecular Science (T.SoaresdaCosta@ won’t fill you with false hope! latrobe.edu.au). Disadvantages: Can really erode your confidence and enthusiasm, particularly if things aren’t going well in the first place. Compatible with: An optimist.

Type 6: The Friend Dr Tatiana Telltale signs: Someone who will tell you all the things you Soares want to hear. da Costa Advantages: Supervisor will understand life events will and her impact on your work, and will be very sympathetic and NHMRC talk through practical solutions. Early Disadvantages: May sometimes have difficulty delivering Career bad news or much needed criticism. Fellowship Compatible with: Students who need emotional support. supervisor, Associate Type 7: The Coach Professor Telltale signs: Supervisor is involved in the growth of Matthew the student in all aspects of their life. Supervisor will Perugini, encourage you to go on training courses, to present your who work and get you involved in community/departmental describes his engagements. supervisory Advantages: You will be involved in different aspects of style as the academia that will help expand your resume. You will Coach. also acquire other transferrable skills that will be useful in non-academic fields.

Page 18 AUSTRALIAN BIOCHEMIST Vol 47 No 1 April 2016 Australia Day Honours for ASBMB Members

Professor Marilyn Anderson was awarded an Officer of Professor Philip Kuchel was awarded a Member of the the Order of Australia (AO) for ‘distinguished service to Order of Australia (AM) for ‘significant service to science science, and to higher education, particularly to biochemistry in the field of biochemistry, as an academic, author and and molecular biology, as an academic and researcher, and researcher, and to professional organisations.’ to professional associations.’ Philip is currently Professor Emeritus at the School of Professor Anderson’s work is based on plant immunity, Molecular Bioscience at the University of Sydney. He was focusing on antifungal and insecticidal molecules, circular originally appointed to one of the two ‘establishment’ peptides, cell walls and plant biotechnology, to help fight chairs in the Department of Biochemistry at the University diseases that cause major crop losses worldwide. of Sydney in 1980. He alternated Headship of the Together with Professor David Craik from the University Department with Professor Gerry Wake in two-year of Queensland, she has recently been in the limelight periods until the mid-1990s. Philip also spent a number of for work that aims to turn plants into pharmaceutical years as Executive Director of the Singapore Bioimaging ‘factories’ to produce potent next-generation medicine for Consortium. He has been a Member of ABS/ASBMB for human diseases more cheaply. 41 years, and was awarded the Boehringer Mannheim After gaining her BSc Honours from the University of Medal in 1983 and the Lemberg Medal in 1999. He was Melbourne and her PhD in Biochemistry from La Trobe on the ASBMB Council from 1993–1997 and was President University, she spent seven years in the United States from 1994–1996. He chaired the Organising Committee working on diabetes and cancer genes at the University for the ~1000-delegate joint FAOBMB/ASBMB meeting of Miami and Cold Spring Harbor Laboratory on Long at Darling Harbour, Sydney, in 1995, and chaired the Island. ~1000-delegate ComBio2002 meeting also at Darling She returned to Australia in 1982 to establish a molecular Harbour. He was involved in founding the Biochemical biology group at the newly established Plant Cell Biology Education Special Interest Group of ASBMB. He co- Research Centre at the University of Melbourne. In 1995 edited and authored with Greg Ralston Schaum’s Outline she set up a laboratory in the Department of Biochemistry of Biochemistry (1988; third edition 2009), and Biochemistry at La Trobe studying defence molecules produced by Through Questions (1993). He was elected as a Fellow of plants to protect them against insect pests and disease. the Australian Academy of Science in 1997 and has been Hexima Limited, of which Professor Anderson involved in many committees of the Australian Academy has been Chief Science Officer since 2009, is now of Science, including serving a four-year term as Secretary developing applications of this work for agricultural and for Science Policy, 2005–2009. pharmaceutical industries worldwide. His research is classified as Physical Biochemistry and it Founded by Professor Anderson and La Trobe Chancellor revolves around applying NMR spectroscopy to cellular and former CSIRO head, Professor Adrienne Clarke, systems. Hexima is located on La Trobe’s Melbourne Campus, Philip was awarded a Leverhulme Visiting Professorship where it has a series of glasshouses for making transgenic to the University of Southampton in 2014 and 2015; he was crops with increased tolerance against fungal disease. elected a Fellow of the International Society of Magnetic A senior scientist at the La Trobe Institute for Molecular Resonance in 2014; and his research in NMR-based studies Science (LIMS), Professor Anderson is the Charles La of red blood cells is funded by the ARC. Trobe Fellow at the University, as well as a Fellow of both A large number of former BSc(Med), BSc(Hons), PhD the Australian Academy of Science and the Australian students and postdocs have gone on to leading scientific/ Academy of Technological Sciences and Engineering. academic appointments in Australia and overseas. His favourite moment at a ComBio meeting was in 2005 when he won an HP digital camera from Sapphire Biosciences in the Passport Competition.

Vol 47 No 1 April 2016 AUSTRALIAN BIOCHEMIST Page 19 Medallist and Awardee Profiles

cancer xenografts has begun to serve as excellent preclinical The Lemberg Medal is awarded to a distinguished Australian biochemist or molecular biologist who will present the Lemberg models for testing new therapeutic drug combinations for Lecture at the ComBio meeting. The Medal is presented in the treatment of breast cancer. memory of Emeritus Professor M.R. Lemberg, who was the Jane’s work, supported by her close collaborator Geoff Society’s first President and Honorary Member. Nominees must Lindeman, many outstanding postdoctoral fellows and PhD have been members of the Society for at least five years before the year in which the Medal nomination is to be considered. An students, and funding from the NHMRC, NBCF, VBCRC honorarium is provided by ASBMB. and other funding bodies, has been published in over 150 papers. She is a Fellow of the Australian Academy of Science and the recipient of a NHMRC Australia Fellowship. She has received awards including the Tschira Stiftung Lectureship (German Cancer Centre), the GlaxoSmithKline Award for The Lemberg Research Excellence (joint award) and the Royal Society of Victoria Medal for Excellence in Research. Medal: The Merck Research Medal is awarded to an outstanding Jane Visvader Australian biochemist or molecular biologist with less than 15 years postdoctoral experience. The successful candidate will present the Merck Medal Lecture at the ComBio meeting. Nominees must have been members of the Society for at least two years before the year in which the Medal nomination is to be considered. An honorarium is provided through the generosity of Merck Pty Ltd. Professor Jane Visvader is joint head of the Division of Stem Cells and Cancer at the Walter and Eliza Hall Institute. She obtained her PhD in molecular biology under the late molecular virologist Robert Symons in the Department of The Merck Biochemistry at the University of Adelaide. She then carried Research out postdoctoral work at the Salk Institute with Professor Inder Verma, and at WEHI with Professor Jerry Adams. Her Medal: contributions to defining regulators of haematopoiesis were Brett Collins recognised by appointment as a Faculty member at WEHI in 1993. This was followed by an appointment as a Harvard Medical School Instructor in Boston, where she continued to study molecular regulators in the blood compartment with Professor Stuart Orkin. In 1998, Jane was recruited to Victoria by the prestigious Victorian Breast Cancer Associate Professor Brett Collins is a group leader Research Consortium (VBCRC) to establish a Breast Cancer and a National Health and Medical Research Council Laboratory at WEHI jointly with Professor Geoff Lindeman. (NHMRC) Career Development Fellow at the University of Over the past decade, Jane and her team have made Queensland’s (UQ) Institute for Molecular Bioscience (IMB). important contributions to the mammary biology and breast He is a structural biologist and heads IMB’s Molecular cancer fields by isolating mammary stem cells, defining Trafficking laboratory. master regulators of mammary gland development and Brett completed his undergraduate studies in biochemistry identifying genetic lesions that drive oncogenesis. In 2006, at the University of New South Wales, and a PhD at Jane and colleagues published a milestone study in Nature Macquarie University’s School of Chemistry in 2001. Under describing the successful isolation of the long-sought the guidance of Professor Bridget Mabbutt, his research mouse mammary stem cell. In other work, it was revealed aimed to use NMR spectroscopy to determine structures of that breast stem cells are highly responsive to steroid small proteins involved in RNA splicing. However, he soon hormone signalling, despite lacking hormone receptors, discovered these proteins formed larger complexes, which thus explaining the long-established epidemiological link required him to learn X-ray crystallography with help from between hormone exposure and breast cancer. Several Professor Paul Curmi. It seemed fate had intervened and set master regulators that orchestrate cell fate decisions in the him on the path towards being a crystallographer. mammary gland have also been defined, providing an In 2001, Brett moved to the Cambridge Institute for Medical indispensable framework for understanding mammary Research in the UK under the mentorship of Professor David lineage commitment and differentiation. Jane and colleagues Owen. It was during his postdoctoral studies (2001–2006) subsequently proved the existence of an analogous hierarchy that Brett became obsessed with how molecules were able in human breast and derived unique gene signatures for the to move around the cell – the process known as membrane different subpopulations. This work led to the discovery that trafficking. aberrant luminal progenitors, rather than stem cells, are the After moving to UQ’s IMB in Brisbane (2006), Brett’s focus transformation target in BRCA1-associated basal tumours. shifted towards understanding the molecular basis for Most recently, Jane’s extensive bank of human breast intracellular trafficking from organelles called endosomes. Page 20 AUSTRALIAN BIOCHEMIST Vol 47 No 1 April 2016 Medallist and Awardee Profiles

This continues to be a major interest of his lab, alongside biology, as well as an interest in the area of scholarship of studies of membrane remodelling in unusual membrane teaching and learning (SOTL), with the development and structures called caveolae, and probing the basis of vesicular implementation of innovative practices and curricula in the fusion in exocytosis. In recent years, Brett and his team have context of laboratory courses in the biological sciences. focused on the central role endosomal trafficking plays in Giovanna believes that engagement is a key element of neurodegenerative disorders, including Alzheimer’s and the learning process and ‘learning by doing’ is a teaching Parkinson’s disease. In particular, his lab is at the forefront of strategy she uses to achieve engagement. structural studies of retromer and sorting nexin proteins, as To achieve this, she has developed and implemented they work to understand how mutations and dysregulation innovative ‘hands on’ competency based stand alone of these proteins lead to the onset of neurological and laboratory courses, an example of which is Biotechniques neurodegenerative diseases. Laboratory. This course was developed as an introductory, Brett’s research spans the complementary disciplines of stand-alone, second year laboratory course with a focus structural biology, biomolecular interactions and cellular on the development of practical skills, competencies biology. His lab collaborates with cell biologists and and knowledge in experimental techniques commonly neurobiologists around the world in an attempt to perform used across the disciplines of biochemistry, molecular true structure-function analyses of protein complexes that biology and microbiology. The course facilitates the are essential for human cellular physiology. achievement of student laboratory competencies through: Brett has published over 65 papers in leading journals, pre-laboratory preparation that utilises innovative online including Nature, Cell, Nature Structural and Molecular resources; the teaching of explicit skills; a competency- Biology, Developmental Cell and PNAS. He has received based approach with the achievement of designated several prestigious fellowships, including an Australian competencies determining progression; opportunities to Research Council Future Fellowship and two NHMRC repeat to achieve competency; post-lab workshops linking Career Development Fellowships. In 2008, Brett received a theory with practice; and the consolidation of learned UQ Foundation Research Excellence Award, and in 2015, he skills in different and increasingly problem-based contexts. was awarded the Emerging Leader Award of the Australian While this course was initially developed to address an and New Zealand Society for Cell and Developmental identified deficiency in foundational practical laboratory Biology (ANZSCDB). skills amongst second and third year students, Giovanna’s meta goals as an educator have been to: engage students The Beckman Coulter Education Award rewards outstanding in order to enhance their learning; address the relevant achievement in education of biochemistry or molecular biology, science threshold learning outcomes; and align student especially innovation and creativity in education, with a view to fostering leadership in this important area of the Society’s learning with professional needs and standards to enhance objectives. The Award will enable the recipient to participate in employability. an international conference with a significant focus on education, Giovanna’s work has been recognised by teaching awards or to spend a period of time at another institution (in Australia including GU Faculty of Science Learning and Teaching or overseas) for the purposes of undertaking developments in education of biochemistry and molecular biology. The recipient will citations in 2007 and 2011, a GU Science Teaching award present a lecture within the Education Symposium at the ComBio in 2013, and recently an Office for Learning and Teaching meeting. Applicants must have been members of the Society for (OLT) Australian Award for University Teaching in 2014. at least two years before the year in which the Award application is More recently, Biotechniques Laboratory was included in to be considered. The contribution to travel expenses is provided through the generosity of Beckman Coulter. an articulation degree program with Nanjing University, China. Giovanna publishes in both areas of research and her scholarly work has been featured in two invited peer- The Beckman reviewed papers in international journals. She is also a team member of the Australian MathBench project, Coulter which was funded in 2013 through an OLT Innovation and Development grant to support the improvement of Education quantitative skills of undergraduate science students. Award: Giovanna Di Trapani

Dr Giovanna Di Trapani is a senior lecturer in the School of Natural Sciences at Griffith University (GU). She completed her undergraduate studies in Italy and her PhD in Biochemistry at GU. Her research covers scientific interests, in the field of redox and oxygen control in cancer

Vol 47 No 1 April 2016 AUSTRALIAN BIOCHEMIST Page 21 Medallist and Awardee Profiles

was less successful than we had hoped for, but the former The LabGear Australia Discovery Science Award is awarded to an has proven very popular. Australian biochemist or molecular biologist for distinguished More recently, we have been getting excited about the contributions to the field of biochemistry and molecular biology. The Award is intended as a Travelling Lectureship to enable the structural and mechanistic details of chromatin remodelling, awardee to present his/her work at a number of centres within as executed by the nucleosome remodelling and deacetylase Australia and New Zealand. Nominees must have been members of complex – a project that I suspect will occupy us for the next the Society for at least two years before the year in which the Award 20 years. We are also trying to understand the importance of nomination is to be considered. The award carries an honorarium to cover the travelling expenses, provided through the generosity post-translational modifications in transcription factors – in of LabGear Australia. what would be an expanded histone code. Finally, I have also enjoyed involvement for the last eight years in the CSIRO Scientists in Schools program. Through this and a range of other school activities, I’ve tried to get the The LabGear next generation excited – not necessarily about doing science themselves even, but in understanding how important it is Australia to ask sensible questions. Discovery The Eppendorf Edman Award is awarded to a biochemist or Science Award: molecular biologist with no more than seven years postdoctoral experience, in recognition of their outstanding research work. Joel Mackay The Award provides funds to assist the recipient to attend an overseas conference in a field associated with biochemistry or molecular biology or to briefly visit a research laboratory in Australia or elsewhere to access specialised equipment or to learn new research techniques. Applicants must have been members of the Society for at least 12 months before I have spent the major portion of my career working to the year in which the Award application is to be considered. understand aspects of how genes are switched on and The contribution to travel expenses is provided through the switched off. generosity of Eppendorf South Pacific. After BSc and MSc degrees at the University of Auckland, I ventured to Cambridge for a PhD with Dudley Williams in the Department of Chemistry. It was here that my interest in molecular recognition was born, with Dudley asking fundamental questions about the physical chemistry underlying biomolecular interactions and using this The Eppendorf information to draw conclusions and make predictions Edman Award: about systems such as antibiotic activity. It was also here that I became attached to NMR spectroscopy as a tool for Michelle probing protein structure and function. Dunstone Two years as an ARC Postdoctoral Fellow in Glenn King’s lab at the University of Sydney (1995–6) reinforced the lessons I’d learned with Dudley about trying to ask important questions and doing rigorous science – and also After completing an Honours degree at Monash brought the realisation that not all groups play cricket *in* University, Michelle Dunstone became fascinated with the lab. protein structure. This fascination led her to a PhD in Since 1997, I have stayed put and built a lab with the the laboratory of Professor Michael Parker (St Vincent’s assistance of fellowships and grants from the ARC Institute of Medical Research), a leading expert in the and NHMRC. It has all been built fundamentally on a structure of pore forming proteins including aerolysin, collaboration with Merlin Crossley, through an initial colicins and cholesterol dependent cytolysins (CDC). It was approach he made to Glenn – to probe the structural basis Michelle’s task to determine the first structure of another for an interaction between two zinc-finger domains. class of pore forming toxins, the MACPF family, which Early research highlights were (i) the realisation that zinc- are hole-punching proteins used by the human immune finger domains can act as protein recognition modules – system. Although Michelle’s PhD research did not lead an idea that still has a huge amount of potential to direct to a representative structure of a MACPF protein, her our understanding of transcriptional regulation and (ii) bioinformatics did identify the fact that MACPF proteins elucidation of the molecular mechanism of action of the were used in all kingdoms of life, including bacteria. protein chaperone of alpha hemoglobin (work driven Michelle went on to work with Professor Jamie Rossjohn by David Gell while he was at the University of Sydney). at Monash University (Department of Biochemistry and These projects have led to efforts to develop new research Molecular Biology), studying structural immunology, which tools, such as our DNA/RNA Pentaprobes, and attempts involved determining the structure of CD3 (published to engineer new classes of RNA-binding proteins. The latter in the journal PNAS). Success from this postdoc research Page 22 AUSTRALIAN BIOCHEMIST Vol 47 No 1 April 2016 Medallist and Awardee Profiles led to an NHMRC Peter Doherty Fellowship. Working in Yit-Heng (Heng) Chooi completed his PhD under the collaboration with Professor James Whisstock she was able guidance of Professor Ann Lawrie and Professor Dave to discover that bacterial MACPF proteins, and therefore the Stalker. Fascinated by how plants and microorganisms human MACPF proteins, shared the same fold as another produce chemically diverse bioactive molecules, he chose class of bacterial toxins, the CDCs (published in the journal to study the genes involved in the biosynthesis of a group Science). of secondary metabolites, called polyketides, in lichenized- The next challenge for Michelle was to determine exactly fungi, for his PhD. He had the opportunity to work with how MACPF and CDC pore forming proteins change eminent lichen chemist Professor Jack Elix (Australian shape to punch holes in membranes. During her NHMRC National University) and taxonomist Dr Simone Louwhoff Career Development Fellowship she combined X-ray (Royal Botanical Gardens, Melbourne), and visited crystallography, biophysics, computational biology and Professor Isao Fujii and Professor Yutaka Ebizuka’s lab at Single Particle cryo-Electron Microscopy (SP cryo-EM) to the University of Tokyo. While completing his PhD thesis at determine the mechanism and trajectory of pore formation the end of 2008, Heng took up a research assistant position by MACPF proteins (published in the journals PLoS Biology, briefly at the University of Minnesota with Professor Dirk Bioinformatics, PLoS Computational Biology). This was part of Hoffmeister (now at the Hans Knöll Institute). He then a great collaboration with Professor Helen Saibil at Birkbeck moved to the University of California, Los Angeles (UCLA) College, London. More recently Michelle, Helen and James in 2009 for his postdoctoral research at Professor Yi Tang’s determined the 8Å resolution SP cryo-EM structure of a lab; at the time, the lab was developing the technology polymerised version of the MACPF protein, C9 (published for recombinant production of large, intact, multidomain in Nature Communications). fungal megasynthases for biochemical characterisation. Michelle’s career path has not been straightforward. In Heng had an extremely productive time at UCLA, where he the last nine years, Michelle has been raising two daughters published 20 papers with Professor Tang, including many with a very supportive family, department and university. in top journals in the field such as Journal of the American Despite being based in Melbourne throughout her career, Chemical Society and Angewandte Chemie International Edition. Michelle has actively sought out opportunities to collaborate He was involved in the elucidation of the biosynthesis of internationally and perform experiments in cutting-edge several important fungal metabolites, including the anti- laboratories, such as Helen Saibil’s. methicillin-resistant Staphylococcus aureus viridicatumtoxin, What lies in the future? Michelle is now committed to antifungal drugs griseofulvin and echinocandin, actin determining how these proteins form in real time rather inhibitor cytochalasin, anti-cholesterol drug lovastatin, than the snapshots gleaned from crystallography and antiangiogenic agent fumagillin, protein transport inhibitor SP cryo-EM. To achieve this, she will collaborate with brefeldin. He also discovered a new immunosuppressive Associate Professor Mark Wallace, from Oxford Univeristy, compound from a human fungal pathogen. and Dr Bart Hoogenboom from the London Centre for In 2013, the award of an ARC Discovery Early Career Nanotechnology. Researcher Award (DECRA) fellowship allowed Heng to return to Australia. Hosted by Associate Professor Peter The Bioplatforms Australia Award is awarded to a biochemist Solomon at the Australian National University, Heng or molecular biologist with no more than 7 years postdoctoral employed his skills and knowledge to discover secondary experience working in the field of genomics, transcriptomics, metabolites from fungal plant pathogens. This is highly proteomics, metabolomics or relevant bioinformatics. The award is based on recognition of outstanding research and complementary to the research theme in Peter Solomon’s the potential to carry out independent research. Preference lab, which focuses on molecular plant pathology. Guided is given to those setting up an independent laboratory for by transcriptomics data from the lab, Heng has discovered the first time. The Award provides $10,000 worth of access the pathways for several metabolites in the wheat pathogen to the services provided by nodes of Bioplatforms Australia. Applicants must have been members of the Society for at least Parastagonospora nodorum. In 2015, Heng was to take up 12 months before the year in which the Award application is a position as a lecturer at the School of Chemistry and to be considered. Biochemistry, University of Western Australia, where he founded a new lab (www.chooilab.org). The lab focuses on employing synthetic biology and genomics to access the chemical diversity encoded in fungi and to uncover the biological functions of these molecules. The 2016 Bioplatforms Australia Award will be used to The Bioplaforms perform RNA-seq of two important fungal wheat pathogens P. nodorum and Zymoseptoria tritici. Combining RNA Australia sequencing and metabolite profiling, the project aims to Award: map out the global genotype-chemotype relationship of the two wheat pathogens and to advance our understanding of Yit-Heng Chooi the roles of small molecules in fungus-plant interactions.

Vol 47 No 1 April 2016 AUSTRALIAN BIOCHEMIST Page 23 ASBMB FELLOWSHIP PROFILES

The ASBMB Fellowships are awarded annually to biochemists or molecular biologists, in their early career and normally resident in Australia, in recognition of their outstanding work in an area of biochemistry and molecular biology. The Fellowships provide funds to assist the recipient to attend an overseas conference in a field associated with biochemistry or molecular biology or to briefly visit a research laboratory in Australia or elsewhere to access specialised equipment or to learn new research techniques. Applicants must be at least in the second year of PhD training and not more than 2 years subsequent to the award of the PhD degree. Applicants must have been members of the Society for least 1 year immediately prior to application.

STANLEY XIE - recipient of the Fred Collins Award for the most outstanding ASBMB Fellowship applicant Stanley Cheng Xie completed his Bachelor of Science in Biochemistry, Immunology and Microbiology at the University of Adelaide. In 2011, he undertook an Honours project in Structural Biology under the supervision of Associate Professor Charles Collyer at the University of Sydney. After Honours, Stanley received PhD scholarships (MIRS and MIFRS) from the University of Melbourne and joined Professor Leann Tilley’s team as a PhD student. Additionally, he was also awarded a top-up scholarship from the ARC Centre of Excellence for Coherent X-ray Science (CXS). Stanley’s PhD project was to investigate the mechanism of action of and resistance to artemisinin antimalarials. His PhD work led to three first/co-first author publications in PLoS Biology, Journal of Cell Science and International Journal for Parasitology. He also co- authored research articles in Nature and PNAS. In addition, Stanley is a co-author on a provisional patent application demonstrating a method for the treatment of artemisinin-resistant malaria. Stanley received travel awards that enabled him to present his work at the 2013 American Society of Tropical Medicine and Hygiene Annual Meeting in Washington, DC and the 8th International Conference on Heme Oxygenases, BioIron and Oxidative Stress in Sydney. He was awarded a best poster prize for the latter. Stanley has recently completed his PhD and is now continuing his research as a postdoctoral fellow in the Tilley laboratory. The ASBMB Fellowship will allow Stanley to visit Millennium Pharmaceuticals, Inc (Takeda Oncology Company) in Boston and perform a screen of their proteasome inhibitor library to identify Plasmodium-selective inhibitors for antimalarial treatment. STEPHEN FAIRWEATHER Stephen Fairweather completed a Bachelor of Philosophy (Science) degree with First Class Honours at the Australian National University in 2010. It was during his undergraduate degree that Stephen first became interested in protein structure–function relationships, particularly in membrane proteins. This led him to undertake first Honours, and then his doctorate in the laboratory of Dr Stefan Bröer at the Australian National University. Stephen’s research has focused on the molecular interactions between human epithelial secondary active amino acid transporters and their heteromeric protein partners and what this tells us about transporter function in healthy and diseased states. In addition, the last year of his doctorate has seen him branch out and begin the investigation of novel amino acid transporters from apicomplexan parasites. His research has led to numerous first and contributing author publications during his PhD, in journals such as Biochemical Journal, Journal of Biological Chemistry and Hypertension. In particular, Stephen is interested in investigating how basic biophysical processes at the molecular transport level can explain organism- or systems- level processes and disease states. His long-term goal is to study the biophysical processes underlying secondary transporter function and combine them with emerging techniques for in situ genome editing to bridge the gap between fundamental molecular understanding and human diseases. Stephen is currently finalising his thesis and some other doctoral research for publications. He will use the 2016 ASBMB Fellowship to attend and present at the Gordon Research Conference and ‘Membrane Transport Proteins’ Gordon Research Seminar in Barga, Italy, in June 2016.

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SWETA IYER Following completion of her Masters degree, Sweta undertook a PhD in Dr Ruth Kluck’s laboratory at the WEHI with the support of an International Research Scholarship from the University of Melbourne. Her thesis research focused on understanding the mitochondrial pathway of apoptosis regulated by the Bcl-2 protein family. In particular, she investigated how cells are killed when the pro-apoptotic proteins Bak and Bax form a pore in mitochondria. By utilising a combination of biochemical, cell biology and in silico modeling techniques, she identified how the Bak and Bax membrane anchors arrange in the apoptotic pore. During this time, she also identified an entirely new mechanism of triggering Bak- and Bax-mediated apoptosis using antibodies, providing new insights into developing treatments for cancer and degenerative disorders. Her studies so far have led to a first-author publication in Cell Death & Differentiation, a first-author manuscript that is currently under revision withNature Communications, co-author publications in PNAS and Cell Death & Differentiation as well as a patent. Sweta finished her doctoral studies in late 2015, and continues to pursue her research on Bak- and Bax-mediated apoptosis at the WEHI. The ASBMB Fellowship will allow Sweta to attend the Gordon Conference on Cell Death in Spain later this year to present her new findings to an international audience. She will also take this opportunity to visit labs and antibody- based companies in Europe to learn about emerging antibody technologies, and to investigate therapeutic opportunities to exploit her recent finding for clinical utility.

WEI HONG TOH Wei Hong Toh completed his Bachelor of Science at the University of Melbourne majoring in both Biochemistry and Molecular Biology and Pharmacology in 2009. He then pursued his interest in research under the supervision of Dr Catherine Butler and Dr Christine Seers at the Royal Dental Hospital of Melbourne, where he investigated the regulation of gene expression of a manganese-sensing metal transcriptional regulator in Porphyromonas gingivalis. He completed his Honours year with First Class Honours in 2010 and was also awarded the Dean’s Honour List, Oral Health CRC Top-up Scholarship and the Gordon Castles Scholarship. In 2011, he commenced on his PhD studies at the Department of Biochemistry and Molecular Biology, University of Melbourne, (under the supervision of Professor Paul Gleeson) after he was awarded the Melbourne International Fee Remission Scholarship and the Melbourne International Research Scholarship. His PhD project focused on the investigation of intracellular trafficking pathways of the amyloid precursor protein (APP) and BACE1 in Alzheimer’s disease as well as the signals and machinery that regulate their intracellular transport. His PhD work has resulted in a first and second author paper in Methods in Molecular Biology and Traffic as well as a review in Frontiers in Cell and Developmental Biology. He received several bursaries and travel awards to present his work at national and international conferences, including the American Society of Cell Biology meeting and also ComBio2011 where he received a best poster award. He completed his PhD in late 2015 and is working as a postdoc in the Gleeson lab, investigating the regulation of APP and BACE1 transport. The ASBMB Fellowship will allow him to present his work at the 2016 Lysosomes and Endocytosis Gordon Research Conference in Andover, New Hampshire.

Showcase on Research Great Expectations Technical Features SDS (Students) Page Coverage of all issues from 2000 to the present

Vol 47 No 1 April 2016 AUSTRALIAN BIOCHEMIST Page 25 Off the Beaten Track

Written by former researchers who have now established careers outside of research, Off the Beaten Track is intended to give the readers insights into the range of alternative careers available to them. Authors describe the paths they have taken to arrive at their present career and provide a detailed description of exactly what the job entails on a day-to-day basis. Old Dogs and New Tricks Michael Crouch, Director of Business Development, TGR BioSciences

As I am now in my late 50s, I consider myself an Old I was fortunate to receive. During this time, my lab was Dog (unfortunately). Looking back over my career, it is mostly doing cellular protein biology, often utilising scary how quickly the years have gone. The time from antibodies to target proteins we made in-house, running mid-career to late-career, in particular, has gone in the lots of gels and Westerns, and also doing a significant blink of an eye. I have been asked to reflect on these years amount of confocal microscopy. The combination of in this article, and offer some insights to young scientists these different techniques, particularly with our unique thinking about their future, and to those contemplating a antibodies, really differentiated us in the scientific area change in their career path. we were working on. The message I want to give to aspiring Australian How can I scientists is that you should expect your career path to summarise my change over time. Sometimes this is your choice, and scientific life up sometimes it is not. The key is to try to shrug off whatever to this point? I disappointment there may be when a change occurs, or felt I was pretty when your initial path is not your preferred option. You are good at what I was an intelligent person, who has a lot to offer the world. This doing, but was not contribution can take many forms, and it is not necessarily really competitive doing what you first started doing in your life. I would in total output say, almost invariably, that you will find whatever you with the big do to be more interesting than you anticipated, providing international you give it a good shot. labs. My lab at A snapshot of my scientific and business the ANU was on career is as follows: my PhD in the Department a block grant, or Michael Crouch. of Physiology at the University of Adelaide Wellcome funding, and for much of the time it was just me (1982–1985) was in cell signalling with Michael Roberts, and a very competent technician. The maximum size was specifically on the role of the inositol phospholipid four people, including one or two postgraduate students. signalling pathway in Ca-dependent responses. The I was at the bench every day, which I really enjoyed, but discovery of IP3 and diacylglyerol activating Ca release without access to other granting sources, there was no and PKC occurred near the end of my PhD, and was a way to increase my laboratory size at the institute. very interesting time, as there was an explosion of second Also, as I mentioned above, I had a desire to do research messenger research worldwide. I then went to the USA that was not just re-doing what someone else had done for three years, as a postdoctoral fellow at Burroughs in a different system, but broke new ground. This made Wellcome (a pharmaceutical company) in a department my research life very interesting and challenging, but of the company that carried out pure research. This work also made publication of data more difficult, as my data was in the study of the role of G-proteins and protein often challenged accepted norms. An example of this was kinases in platelet function, and I was working for a my work on G-proteins, which proposed that one such leader in the field at the time, Eduardo Lapetina. This was protein translocated into the nucleus and was important when I learned to really take chances experimentally, and in regulating cell division. At the time, these G-proteins explore things that were logical and important, but were a were largely considered to be only at the cell membrane bit out of left-field to mainstream thinking. It was a really and involved in receptor signal transduction. This work productive time, and we published a lot of interesting was pretty much considered as heresy at the time by scientific papers. I then returned to Australia, to the John some, but was shown some years after I left basic research Curtin School of Medical Research at the ANU. For most to be correct. of this 12 year period I continued studying G-proteins So, combined with the limited resources available, I can and kinases, but now focused on their role in regulating accept (sort of) that my style of research was, to a degree, cell proliferation and cell migration – which was really digging my own research grave. This is easier to see basic cancer research. Five of my 12 years there were looking back than it was then. I felt at the time (and still funded by a Wellcome Senior Research Fellowship that do) that if you are doing research, you might as well find

Page 26 AUSTRALIAN BIOCHEMIST Vol 47 No 1 April 2016 Off the Beaten Track really new things. Anyway, things were about to change. During these years, my activities within the company It was away from Canberra that I moved in order to have changed. For the first eigtht to ten years at TGR, I transition from academia to business. I did this mainly was both in the lab as well as doing administrative work. to ensure I had work and income to support my young I was also Chief Scientific Officer (CSO) for several years. family, as I was concerned about the certainty of my More recently, I have moved to the business development academic funding and employment. Additionally, I side, leaving the new technology development to our wasn’t averse to the commercial world, as I had always present CSO, Antony Sheehan. This means I am now very thought that the business world was the only conduit for rarely at the bench. Rather I spend my time talking to our scientific discoveries making it anywhere to help people collaborating companies, examining market opportunities – i.e. you ultimately had to get products to the market for new products, working on patent applications for if people were to benefit from basic discoveries. So, we our novel technologies, and looking for new business moved back to Adelaide, and I have been with a small opportunities. However, dealing directly with both biotech company called TGR BioSciences for the last 16 business and science people around the world is still very or so years. interesting, and I can get my ‘science-fix’ without carrying TGR is a company that detects cellular proteins involved a pipette anymore. in receptor activation and disease using immunoassay So, to summarise, I have gone from academia to business, technologies. This was pretty much the type of practical and had ups and downs in both. However, they have focus of my entire academic career, and was a good fit for both been challenging and interesting, but in different me. I joined TGR when the company was first formed. My ways. Your career is likely to have its ups and downs role was to lead the development of the company’s first as well, and you may have to consider employment assay technology and products. Similar to my academic options that were not part of your initial thinking. You approach, I didn’t want to use the old systems that were just need to keep your chin up, and remember you have really slow and laborious – I wanted new technologies a lot to offer the world. Keep your eyes open, try hard, that would speed up our own research, and form the basis stay positive, and make the most of the opportunities of products that would be totally new in the marketplace, presented to you! and really speed up therapeutic research around the world. What we were looking for was a system that could very rapidly measure and quantitate cells being activated or inhibited by a wide variety of stimuli, such Written by the Australian Society for Biochemistry as new therapeutic drugs (e.g. cancer inhibitors, anti- and Molecular Biology Past President inflammatories etc). At the time, the main methods were and Honorary Member William Elliott really slow (e.g. measuring cell growth), or were limited and long-standing member Daphne Elliott. to a small number of receptors or stimuli (e.g. measuring calcium signalling or cAMP). From my years of working with kinases, I thought protein phosphorylation would be a great measurement upon which to build a fast assay, as there are a lot of different kinases that can provide a readout of many different receptors and stimuli. However, no such rapid product existed at the time. So, we started from scratch to try to develop a system to rapidly measure endogenous cellular protein phosphorylation in up to thousands of cellular samples by incorporating a new technology that Perkin Elmer marketed, called Alpha. Without going into detail, this is a bead-based technology that is a homogeneous, mix-and-read system. It is, therefore, very fast and sensitive, and would give us the ability (if it worked for our application) to measure any cellular phosphoprotein we wanted. To cut a long story short, after about one to two years, we got it working, and the system is in our view the best in the world. We now manufacture about 100 products, which we call SureFire and SureFire Ultra, that we distribute worldwide in collaboration with Perkin Elmer. Order your copy today! This has been a really rewarding accomplishment, and the 1300 788 000 team at TGR has contributed all of their expertise to get [email protected] the products and company to where it is today. www.publish.csiro.au

Vol 47 No 1 April 2016 AUSTRALIAN BIOCHEMIST Page 27 Lateral Flow Immunoassays – How Fundamental Breakthroughs from the 1950s Led to the Inception of This Simple Yet Powerful Tool Jia Li 1 and Joanne Macdonald 1,2 1Inflammation and Healing Research Cluster, Genecology Research Centre, School of Science and Engineering, University of the Sunshine Coast, QLD email: [email protected] 2Division of Experimental Therapeutics, Columbia University, New York, NY, USA email: [email protected]

The lateral flow immunoassay (LFIA) has revolutionised innovation through the intersection of different fields. In diagnostics, enabling the development of rapid and this article, we explore the history of LFIA development, inexpensive assays that are very simple to perform. One and then introduce the basic LFIA detection principles of the most famous of these assays is the home pregnancy and formats. We conclude by considering some of the far- test, which has empowered women to monitor their reaching applications that have emerged from this novel reproductive health and future in complete privacy. LFIA combination of immunology and material science. technology continues to be developed and expanded today, and is expanding into various aspects of industry, such Lateral Flow Immunoassay – as the monitoring of food poisoning and quality control, agriculture, environmental monitoring, biodefence, Timeline drug abuse and the diagnoses of infectious diseases. In The LFIA is a paper-based test for the detection of particular, the rapid time to results (within minutes) is analytes, based on antigen and antibody interactions (4). a powerful tool in doctors’ surgeries and emergency The device emerged from a powerful combination of rooms. For example, flu test kits such as the QuickVue© individual technologies that arose during the 1950–1970s. Influenza AB test kit (1), BD Veritor SystemTM Flu A+B The idea of using paper-based assays was originally (2) or Clearview® Exact Influenza A&B (3) are lateral flow pioneered by Martin and Synge, who were awarded the immunoassays that provide clinicians with diagnoses of Nobel Prize in Chemistry in 1952 for their research in influenza virus type A or B within a few minutes. These partition chemistry. The technical basis of the LFIA can rapid results enable timely clinical decisions to be made, also be attributed to latex agglutination assays developed resulting in improvement of patient health outcomes. by Singer and Plotz in 1956 (5). However, the actual The operational simplicity of the LFIA, however, belies transformation of liquid phase immunoassays onto a a fascinating underlying complexity. In particular, the paper-based immunodetection system originated from inception of the technology is an interesting showcase on three fundamental demonstrations: 1. the immobilisation how incremental improvements can result in substantial of antibodies on solid phase by Catt et al. in 1967 (6);

Fig. 1. Principle of radioimmunoassay. A known amount of labelled antigen is mixed with a fixed quantity of antibody. Addition of an unknown amount of the same antigen (unlabelled) leads to competitive binding of the unlabelled antigen with the labelled antigen to the limited antibody. The bound antigen is then separated from the unbound one by immunoprecipitation. Radioactivity levels are measured from both the supernatant and the precipitates. The amount of antigen from the sample can be derived from the binding curve generated by the standard sample that was run in parallel with the unknown sample.

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Fig. 2. Principle of immunoradiometric assay. Antigen standard or unknown sample is reacted with excess radiolabelled antibody. Insulin is added to precipitate the unreacted antibody through centrifugation. The unknown sample can be directly quantified by measuring the radioactivity of the supernatant, and then derived from the binding curve generated by the standard sample that was run in parallel with the unknown sample.

2. the employment of sol particles (gold and silver) as Methods to avoid the use of radioactivity were developed tracers rather than the radiolabelled elements and enzymes by substituting radioactive labelling with enzymatic for immunoassay (7); and 3. the Dot-ELISA (enzyme- labelling, where the presence of enzyme results in a colour linked immunosorbent assay) by Pappas et al. in 1983 (8). change of a solution due to the reaction of an enzyme and its The combination of all of these technologies culminated substrate(s). In 1971, Perlmann and Engvall at Stockholm in the first commercial LFIA product – the now-famous University in Sweden, Schuurs and Van Weemen in the at-home pregnancy test, which was released in 1988 (9). Netherlands, together with Avrameas and Guilbert at the Since then, LFIA devices have gained huge popularity Pasteur Institute independently converted this idea into and are being applied in many diverse situations. a method (Fig. 3), ELISA or enzyme immunoassay (EIA) (12–14). ELISA/EIA was groundbreaking as it eliminated Plate-based Immunoassay from the requirement for radioactive elements and provided a much faster assay reaction. Forty years later, the ELISA/ 1960 to 1970 EIA is still a major procedure in diagnostics. This is because The immunoassay technologies that precede the LFIA ELISA/EIA allows rapid screening and quantitation of an (1960–1970) are particularly insightful incremental analyte or antibody in a large number of samples with innovations that fundamentally shaped LFIA high sensitivity (the typical detection range for an ELISA development. The earliest assay relevant to the LFIA is is 0.1 to 1 femtomole). the radioimmunoassay (RIA; Fig. 1), which dates back to 1960. Berson and Yalow used radiolabelled iodine (131I) to determine the concentration of insulin, a hormone which The LFIA was difficult to measure in the blood (10). The sample to LFIA technology emerged through the radical be assayed (an unknown concentration of insulin) was combination of the plate-based immunoassay with the run in parallel with a radiolabelled standard (known field of chromatography, where reagents move across a concentration of insulin), allowing the amount of antigen solid phase by capillary forces. As such, it is sometimes from the sample to be inferred. The invention of the RIA referred to as an immunochromatographic assay (ICA) was a fundamental advance in immunology, as it allowed (15). The first ICA was performed by Kohn for the the quantification of an antigen (low level of concentration detection of proteins. However, this assay involved in body) via antibody interactions in vitro. However, the several steps rather than the current one-step ICA (16). RIA faced two major disadvantages: the requirement to The all-steps-in-one ICA was first presented by Friesen separate the bound and the unbound antigens before the et al. (17) in a 1984 patent, and subsequently refined in a measurement, and the use of radioactivity and subsequent flurry of patents in 1987. Rosenstein and Bloomster (18) technical difficulties. described the idea of capillary flow (May, 1987), and In 1968, Miles and Hales developed an immuno- Eisinger et al. (19) coined the phrase ‘lateral flow’ (June, radiometric assay (IMRA) based on the RIA (11). Instead 1987). However, it was a patent by Unilever employees of labelling antigen, the antibody was radiolabelled (Fig. Clayton and Porter (20) that was filed first that year 2). This allowed the antigen to be directly quantified (January, 1987). This led to the Unipath Clearblue lateral and obviated the requirement to separate labelled and flow sensation, which was released commercially less unlabelled antigens. However, it did not tackle the issue than a year later in 1988, before publication in 1989 (21) of using radioactive substances, which require careful and subsequent refinements (22). Thus, while several handling due to their harmful effects on a user’s health. different agencies developed similar lateral flow concepts

Vol 47 No 1 April 2016 AUSTRALIAN BIOCHEMIST Page 29 Lateral Flow Immunoassays

Fig. 3. Principle of enzyme-linked immunosorbent assay. A detection antibody is pre-fixed onto a solid phase, and exposed to its corresponding antigen. A second antibody (conjugated to an enzyme) is subsequently added. This antibody binds to the antigen at a different epitope compared to the first antibody. After washing away unbound reagents, subsequent addition of enzyme substrate(s) triggers a colour change in the solution. The colour intensity correlates with the amount of antigen present and can be measured using a spectrophotometer.

Fig. 4. Principle of a lateral flow immunoassay conducted in sandwich assay format. On a reaction phase (e.g. nitrocellulose membrane): (A) The sample (green circle) is loaded onto the sample region. (B) Binding of the unknown analyte to a labelled antibody (the conjugate, red sun with blue arrows) occurs in the conjugate zone. (C) The antigen/conjugate complexes and the unbound antigen and conjugates, travel to both the test and control lines, respectively. If antigen or antigen/conjugate complexes are present they will bind to the test line. If conjugate is present it will bind to the control line. Binding of the labelled conjugate to either line results in the appearance of a coloured stripe along that line. (D) Appearance of a stripe in the test and control lines indicates that antigen was present, a positive result; appearance of a stripe in the control line only indicates a negative result; appearance of no coloured lines is an invalid result.

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Fig. 5. Types of LFIA. A) Non-competitive immunoassay: the analyte (green circle) is sandwiched between the detection antibody and the capture antibody/ tracer (red sun with blue arrows) at the test zone to generate a coloured test line; the concentration of the analyte is proportional to the intensity of the test line. B) Competitive immunoassay with known concentration of analyte being pre- immobilised to the membrane. C) Competitive immunoassay with known concentration of analyte being pre- conjugated to the tracer. In both cases, addition of the analyte from the sample (unknown concentration; denoted by ‘U’) competes for binding with the known concentration of analyte to the capture or detection antibodies. When the concentration of analyte in the sample is lower than a ‘cut-off’ concentration, a coloured test line appears (top half of figures B and C); when the concentration of analyte in the sample is higher than a ‘cut-off’ concentration, the test line diminishes (bottom half of figures B and C). The decrease of test line intensity is proportional to the increase of the analyte in the sample. In both non-competitive and competitive immunoassays, the control line should always appear to validate the test.

Vol 47 No 1 April 2016 AUSTRALIAN BIOCHEMIST Page 31 Lateral Flow Immunoassays based on the prevailing ICA knowledge at the time, it was Competitive Immunoassay the unique commercialisation environment surrounding The limitations of the non-competitive assay can be Unipath that enabled immediate conversion to market compensated by using a competitive immunoassay. and ultimate dominance in the industry (23), with a test In the competitive format, the analyte (unknown that still prevails today under the Alere Inc. banner (24) concentration) competes with a known concentration of (Alere purchased Unipath in 2001). analyte for binding to a single antibody; the detection So how does a typical LFIA operate? The modern LFIA signal is inversely proportional to the analyte (unknown employs several reaction zones distributed across a solid concentration) (Fig. 5B and 5C). The RIA developed by phase, corresponding to where antigens or antibodies are Berson and Yalow (see above) is a classic competitive immobilised. An unknown analyte contained in liquid immunoassay. is applied to one side of the solid phase. The analyte is The competitive immunoassay has two formats, in subsequently wicked along the device and consecutive which the known concentration of analyte is either pre- reactions take place as it passes through each zone (Fig. immobilised on the membrane (Fig. 5B) or conjugated 4). The device employs three antibodies and a tracer: a to the tracer (Fig. 5C) (26–29). In both cases, a bright test capture antibody (antibody 1) is coupled with a tracer line appears when the analyte in the sample is present in (which acts as a transducer to generate the colour observed low concentrations, while a diminishing test line appears in the test/control line); a detection antibody (antibody 2) when the analyte in the sample is in high concentrations is embedded in a test line; and a third antibody (antibody (Fig. 5B and 5C). 3) that can directly bind to the capture antibody is immobilised at the control line (Fig. 4A–C). The capture and detection antibodies can bind an analyte (antigen) Applications of LFIA at different epitopes. The control line indicates that the A wave of LFIA applications originated from the first sample has migrated across the membrane as intended, launch of the Clearview home pregnancy test for the regardless of whether the analyte is present in the sample. detection of human chorionic gonadotropin (hCG) Observation of both a coloured test line and a coloured by Unipath in 1988 (9). Since then, LFIA has gained control line on the membrane is a positive result (Fig. 4D). wide acceptance due to its simplicity, speed and user- A single line in the control zone is a negative result (Fig. friendliness. LFIA devices are compact, portable in size 4D). If no lines appear at all (Fig. 4D), the result is invalid and offer reliable results. To date, the application of LFIA and must be repeated. devices is extremely broad, ranging from food safety, agriculture, aquaculture and environmental to forensic science, therapeutic monitoring, medical diagnostic LFIA Formats and military biodefense. LFIA devices have been used LFIA can be adapted to different formats based on for the detection of parasites (30), bacteria (31–34), cells the type of analyte, which can involve direct detection (35,36), viruses (37–40), presence of toxins in food (41), without competition (non-competitive immunoassay) environmental samples (42–44), usage of illicit drugs or competing with known concentrations of analyte (45), appearance of hormones (46) and biological markers (competitive immunoassay). Both of these formats are (47,48). The number of potential applications for LFIA useful for providing different information about analytes continues to grow, due to the major advantages of this when performed in a LFIA format. technique. In particular, by providing the capacity for at- home biomarker monitoring, this technology is improving Non-competitive Immunoassay our quality of life in a simple and low cost manner. The non-competitive immunoassay is usually called a ‘sandwich assay’, where the analyte is embedded References between the capture and detection antibodies (Fig. 4). In this format, the detection signal is proportional to the 1. QuickVue© Influenza AB diagnostic test CLIA increasing concentration of the analyte (25) (Fig. 5A). Waived, http://craigmedical.com/Influenza.htm, In this case, direct binding of the antigen to the excess accessed 24 February 2016 radiolabelled antibody correlated with an increase in 2. BD Veritor™ System – CLIA-waived for Rapid measurable radioactive signal, directly indicating the Detection of Flu A+B, http://www.bd.com/ds/ amount of antigen present in the sample. However, a productCenter/256045.asp, accessed 24 February 2016 potential problem of the sandwich immunoassay is the 3. Clearview® Exact Influenza A&B, http://www.alere. detection of small molecules that contain only a single com/en/home/product-details/clearview-exact- antigenic epitope. In addition, steric hindrance may influenza-a-and-b.html, accessed 24 February 2016 prevent simultaneous binding of the analyte to both of 4. Hage, D. S. (1995) Anal. Chem. 67, 455-462 the capture antibody/tracer complex and the detection 5. Plotz, C. M., and Singer, J. M. (1956) Am. J. Med. 21, 888- antibody. 892

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Vol 47 No 1 April 2016 AUSTRALIAN BIOCHEMIST Page 33 BOOMERANG REPORT Homeward (RNA-)bound: a Whirlwind Trip of Australian Science Mitchell O’Connell reports on his ASBMB Boomerang Award travels After two and a half years in Berkeley, California, I was her efforts to explore the epitranscriptome by charting excited to find out I’d won the ASBMB Boomerang Award novel m22G RNA modifications and possible links to to travel to Australia, where I’d organised a whirlwind intellectual disability, while Thomas Priess gave us an tour to present my work at ComBio2015 in Melbourne, exciting update on the role of 3’UTR shortening in cardiac the Australasian Genomic Technologies Association hypertrophy and its link with miRNA regulation, together Conference (AGTA) in the Hunter Valley, as well as a underscoring how far we have come (and how far we number of institutes in Sydney, Canberra and Perth. It have to go) in understanding the complex role of RNA in also allowed me to temporarily placate my undying need gene regulation. And to top it all off, we got to dine in the for decent coffee and a good-old Australian sausage roll. Melbourne Aquarium for the Conference Dinner in the After completing my PhD in Joel Mackay’s lab at the company of Pinjarra the 50 year old salt water croc! University of Sydney, I moved to the University of California, Berkeley, to take up a postdoc in the lab of Jennifer Doudna. The Doudna laboratory was the first to show the programmable DNA recognition and cleavage ability of a CRISPR-associated protein known as Cas9. This research led to an explosion in the use of CRISPR/ Cas9 as a genome-editing tool in multiple cell types and organisms. However, this protein had been thought to be incapable of targeting RNA. Recently, we were able to show that this is not the case and Cas9 can be used as a sequence-specific RNA-binding and cleavage enzyme, paving the way for tools to explore the myriad RNA species implicated in gene regulation and disease. Before ComBio, I had the opportunity to present my research at both my old department, the School of Molecular Biosciences, University of Sydney, and the Mitchell O’Connelll in King’s Park, Perth. Kinghorn Centre for Clinical Genomics at the Garvan After ComBio2015, I had the chance to speak at the John Institute. The Kinghorn Center houses the Illumina HiSeq Curtin School of Medical Research at ANU, Canberra, and X Ten sequencing system, which can sequence an entire the Harry Perkins Institute for Medical Research in Perth. human genome for essentially less than $1000 USD! I also got to spent time talking science to old friends, new Standing in the room with all these machines buzzing really friends and a number of PhD students, to whom I was able hit home the enormity of this resource; this technology to offer some advice about working overseas. After these has the capability of churning out 18,000 human genomes trips, I attended the AGTA Conference. As a molecular a year, which is a massive boon for Australian genomics biologist, these types of conferences have always acted as and brings us all a lot closer to routine whole genome great idea generators for my own work – I got to see talks sequencing as part of standard medical practice. on topics as vast as pseudo-time analysis of single-cell After being humbled by a room of sequencers, I travelled transcriptome data (Cole Trapnell), Neanderthal genome to Melbourne for ComBio2015. ComBio gathers great sequencing (Janet Kelso) and ‘wine’omics’ (Anthony researchers from Australia and covers a wide spectrum Borneman), which included a free wine-tasting during the of research across biochemistry, cell, developmental, session! molecular and plant biology. Pam Ronald (University of Rounding off the trip, I accepted a late offer to present California, Davis) gave probably my favourite talk of the my work at the newly-rebranded Western Sydney conference. Her talk was a tour-de-force of basic plant University, Campelltown. It was great meeting with genetics, chemical biology and novel mass spectrometry old friends and colleagues Liza Cubeddu and Roland methods to identify a tyrosine-sulfated bacterial protein, Gamsjaeger, and hearing about their research on DNA- RaxX, that betrays its owner to alert resistant rice plants of repair and chromatin-modifying complexes. It was also a imminent Xanthomonas attack. Other highlights included treat to revisit the very same campus I attended as a year the Sydney Protein Group Lecture by Alastair Stewart 9 high school student undertaking a ‘Summer Science exploring the evolution and stress adaptation of type II Experience’ program. All these years later, I’m still doing chaperonins, where he took us on an impressive structural science and loving it just as much. biology journey through the interesting symmetry and Mitchell O’Connell is an NHMRC CJ Martin Early subunit properties required for the assembly of Sulfolobus Career Research Fellow in the Department of Molecular chaperonins. While on the RNA side (my spiritual and Cell Biology at the University of California, Berkeley. home!), Nicole Schonrock gave a fantastic overview of

Page 34 AUSTRALIAN BIOCHEMIST Vol 47 No 1 April 2016 Science Teachers’ Association of Victoria Science Talent Search

Primary division: Experimental Research/Inventions/ Models/Science Photography. It was thrilling to see so many enthusiastic budding young scientists, who were excited to have their work acknowledged and awarded. A number of terrific projects received ASBMB awards, including: Sunlight and its Role in Our Life; Sustain the Food Chain; Light at the Bottom of the Ocean; Save Those Berries; Life Needs Life; and Looking at the Morphological Changes Leading to Colon Cancer. Like every other year, in the months after the STS I was greeted with many hand-written letters and emails thanking me and the ASBMB for our support. Just a few dollars seems to make a world of difference to these young aspiring scientists and I hope the ASBMB’s support of this initiative will continue for many years to come. The Victorian Branch of the ASBMB is proud to be a Diana Stojanovski, continued sponsor of the Science Talent Search, organised ASBMB Victorian State Representative by the Science Teachers’ Association of Victoria. The www.sciencevictoria.com.au/sts Science Talent Search was founded in 1952 and encourages school students to undertake scientific projects. Students are invited to submit work in eight categories: Computer Programs, Games, Science Photography, Posters and Scientific Wallcharts, Models and Inventions, Experimental Research, Creative Writing, and Videos. The competition is open to Victorian school students from Prep to Year 12. The theme for 2015 was ‘The Science of Light’. There were a total of 2052 entries from 2851 participants. These participants were from 158 different schools from around Victoria. The broad aims of the Science Talent Search are: 1. To stimulate ongoing interest in serious study of the sciences by encouraging independent, self-motivated project work amongst students of science, giving students the opportunity to communicate their achievements to a wider audience and recognising effort and achievement Above: in their scientific enterprise. Light box. 2. To promote the direct involvement of the students in the process of science and its communication. 3. To give the public an opportunity to see the quality of work being achieved in science, by both primary and secondary school students. The Victorian branch of ASBMB supported the 2015 Science Talent Search with a $1,000 donation, which was awarded as major and minor bursaries to 32 students from a number of Primary and High Schools from across the state, including: Fintona Girls School; Box Hill High School; Brighton Grammar Junior School; Essex Heights Primary; Santa Maria College and St Josephs School, Mildura. Entries from award-winning students were displayed at Right: La Trobe University on Monday 23 November 2015, prior Photosynthesis: to the award presentation ceremony. I was privileged to evergreens for life. be invited to the ceremony to present the awards to the

Vol 47 No 1 April 2016 AUSTRALIAN BIOCHEMIST Page 35 IP Lessons from Ground-breaking Developments in Basic Research

This feature is the first in the type of dispute where researchers from the same a series of regular articles laboratory claim to be the true inventors. Information on intellectual property, contained in laboratory notebooks is also critical for written by Sarah Hennebry, determining inventorship in inter-lab disputes but the Patent Attorney, Freehills issues around this are perhaps best considered in a future Patent Attorneys. Here, Sarah article! offers some basic advice Although most countries today are ‘first-to-file’ on patent applications. jurisdictions, in a fast-moving area of research, where there is an urgency to file patent applications, it is common for The Interplay Between Academic and patent applications to be filed before the full extent of the invention has been elucidated by the researchers. It is not Commercial Worlds enough to be the first filer: the content of that filing needs Breakthrough science conducted by academics to be sufficient to show the invention was sufficiently frequently opens doors for future developments in reduced to practice. Most patent offices now require biotechnology and medicine. For example, basic research specific data and examples in the patent application, into bacterial immunity led to the discovery of DNA demonstrating that the invention works (rather than mere restriction enzymes, which ultimately transformed speculation that it could work based on preliminary data). molecular biology research and paved the way for An example of this is where an application only contains recombinant DNA technologies. RNA interference in vitro data but the claims of the application extend to an technology was also borne out of research aimed at in vivo use of the invention. There is a risk that a patent understanding gene silencing machinery in plants and examiner would reject such an application on the basis nematodes and a similar story is now unfolding with the that the inventors have not shown that the invention advent of CRISPR/Cas9 technology. actually works. In addition, failure to adequately explain Being the group that leads the scientific breakthrough the invention in your application may allow someone else does not necessarily guarantee successful commercial to obtain a patent for a later-filed application. protection. What the above-mentioned stories all have in common is that they reveal certain challenges facing Carefully Manage Public Disclosures – Even academic researchers who develop commercially-relevant technologies, which include: After the Patent Application is Filed • balancing the urgency to file patent applications with Statements to the press or at conferences should also the need to ensure the content of the application is be carefully managed. If you say something about your sufficient; invention before you file your application, then you could • managing public disclosures about the invention seriously jeopardise your chances of patent protection. (before and after filing the patent application); and This includes casual statements made at the conference • awareness of intellectual property owned by others dinner while celebrating the success of your scintillating in addition to recognising when their own intellectual conference presentation. property is being generated. You should also be careful of what you say after you Below, we explore some of these concepts in greater have filed the application. For example, statements made detail. by the inventors, years after the first filing of their patent applications, can be used by third parties to argue that the Patent Application Filed – But is it Enough? inventors didn’t know that their invention worked at the time of filing. In some jurisdictions, this can be fatal to a Most countries apply ‘first-to-file’ rules for determining patent application. who is entitled to be granted a patent for an invention. While this is now also true of the US, up until a few years ago, the US applied ‘first-to-invent’ rules. This meant that Research Exemption to Patent Infringement if you could prove you were the first inventor, you could In Australia, there is an exemption to patent infringement be awarded a patent even if you filed your application after for research purposes (in certain circumstances). that of another party. This is decided in a procedure called Specifically, acts done for experimental purposes are an ‘interference proceeding’ which is used to determine exempted from patent infringement if the acts are for: who is legally entitled to an invention. • determining the properties of the invention; Interference proceedings can still be launched in the US, • determining the scope of a claim of the patent; depending on the filing date of the patent applications in • improving or modifying the invention; question. Commonly, excerpts from laboratory notebooks • determining the validity of the patent or of a claim; are used as evidence to show who was the first to get an and invention to work. • determining whether the patent would be, or has Inventorship disputes of this type are distinct from been, infringed by the doing of an act.

Page 36 AUSTRALIAN BIOCHEMIST Vol 47 No 1 April 2016 IP Lessons from Ground-breaking Developments in Basic Research

Take Home Messages application! However, you should also carefully consider There are some valuable lessons to be learned from making statements on potentially commercially important for researchers who are filing patent applications or are technology, even after you have filed the patent application. considering filing applications in the future. 3. Be mindful of others’ IP, not just your own 1. Be the first to file but make sure what you file is relevant When conducting research, many focus on the generation There is no value in being the first to file, unless you of their own IP, forgetting to consider that they may be can demonstrate that you have sufficiently developed using IP owned by someone else. In academic research, the invention and put it into practice. There is a balance this often does not amount to patent infringement. to be achieved between getting the first application in However, you should exercise caution and check with the early enough in a competitive environment (so you don’t IP manager at your university or organisation, who can get pipped at the post), but also, ensuring that you have best advise you on whether you have freedom to operate enough data to support your claims to the invention. in this area.

2. File before you publish and be careful what you say, not The best course of action in dealing with any of the above just when you say it situations is to speak with the intellectual property/ Those of you with some experience of the patenting business development managers at your university, process will have heard this ad nauseam from patent research institute or organisation. In consultation with attorneys: don’t ruin your chances of patenting by telling their patent attorneys, they will be able to provide you the world about your invention before you file your with practical advice relevant to your situation.

Nicholas Lim reports Meeting Legends - of on his ASBMB the Academic Kind Fellowship travels I am delighted to have received the table, and students/postdocs/PIs gathered around an ASBMB Fellowship in 2015 and asked anything they wanted – from latest research which enabled me to attend developments, to key outstanding gaps in the research the American Society for Cell field, to career advice. These were candid events that Biology (ASCB) meeting held provided opportunities for networking, and who knows, in San Diego, USA, from 12 to you might have been offered a job on the spot after 16 December 2015. The ASCB piquing someone’s interest with your interests/expertise/ meeting is one of the largest questions. For those that were interested, there were also international conferences numerous career advisors, industry representatives and covering multiple aspects of cell even immigration lawyers that could provide valuable biology such as trafficking, single-cell systems, epigenetics information if you were keen on pursuing a career in the and imaging, all of which intersect at different scales – United States. from the single cell to the macroscosmic level. Attending From the warm bright sunshine of San Diego, the ASCB the meeting was one of the most memorable moments Fellowship also enabled me to visit leading laboratories in of my young scientific career; highlights included the field of stem cell biology and epigenetics at Harvard attending key symposiums given by leading researchers Medical School. I remember walking into the Medical to interacting with the large numbers of like-minded and School and there was a screen showing upcoming passionate graduate students. It was a melting pot of seminars, and I was overawed at the presenters’ names – ideas and information, where you could have hundreds of it was a who’s who of stem cell biology. I had scheduled talks, exciting posters and microsymposias covering your my days there to meet with some of these researchers, favourite scientific question. The ASCB meeting truly was and had numerous candid and fruitful discussions. I look a showcase of the best work and best researchers in the forward to incorporating these ideas and discussions, and world, and it was exciting for me to listen and learn about importantly their mindsets, as I progress in my scientific the leading developments and partake in the discussions career and hopefully one day I can inspire others in my afterwards. field as well. Once again, thank you ASBMB for this Despite the scale of the event, the ASCB meeting was opportunity! very community-orientated. Throughout the event, Nicholas Lim is completing his PhD at the Department there were satellite sessions where leading experts in a of Biochemistry and Molecular Biology, University of field (or ‘legends’ as I call them) sit themselves down at Melbourne. Vol 47 No 1 April 2016 AUSTRALIAN BIOCHEMIST Page 37 The 41st Lorne Conference on Protein Structure and was a fabulous talk that was whole-heartedly enjoyed by Function was once again a superb showcase for leading everyone present. Lorne student poster prize winners. edge protein science, attracting over 350 registrants, including 62 from overseas. This year, perfect weather enabled attendees to enjoy the beautiful Lorne environs Bruce Kemp in between listening to the latest and greatest in protein (left) receives science, networking and socialising. As in previous years, the Leach the program (Program Chair, Jacqui Matthews) was Medal from diverse, covering topics ranging from chemical biology conference through to RNA regulation, synthetic biology, protein chair James design, signalling and quality control, amongst others. Whisstock. The emergence of electron microscopy as a technique that can rival traditional structural methods such as X-ray crystallography was a recurring theme throughout the meeting. Some stunning EM images of protein complexes Kinases were also featured in several other talks and the insights they produce into protein function were including from Titus Boggon (Yale University, USA), a highlight of a number of talks, including work from Irit Susan Taylor (UCSD) and Young Investigator Awardee, Sagi (Weizmann Institute) on collagenases, and Michelle Chris Langendorf (SVI). The other four YIA winners (Tim Dunstone (Monash University) and Doryen Bubeck Allison, Laura McCaughy, Thomas Ve and Takuya Shiota) (ICL) on complement membrane attack complexes. also delivered very high-calibre presentations. Another Grant Janson (Caltech) delivered an outstanding Special audience favorite was the Hot Topics session with seven Evening Lecture on the use of cryo-electron tomography rapid-fire, ten-minute talks on diverse topics. Karen techniques to understand the structure and function of Harris’ talk on the use of asparaginyl endopeptidase to large protein machines and their arrangement within cells. cyclise peptides won the ‘hottest topic’ crown by a narrow X-ray crystallography, of course, was still central to margin for generating a large amount of discussion. many talks including those from Cheryl Arrowsmith The speaking program was interspersed with afternoon (Structural Genomics Consortium, Toronto), Yvonne and evening poster sessions that generated plenty of Jones (University of Oxford) and local speakers Matthew interest and robust discussion. Hundreds of high-quality Call (Walter and Eliza Hall Institute) and Andrew Brooks posters were on display this year with 19 prizes being (University of Queensland). Protein Society Speaker awarded to Australian and some International students Dev Sidhu (Donnelly Centre for Cellular Biomolecular following the final session of the meeting. It was during Research) gave a wide-ranging talk covering the this final meeting wrap-up that James Whisstock, Chair of generation of synthetic antibodies using phage display the last three Lorne Protein Conferences, announced that techniques through to the production of D-peptides he would hand over the reins to a new Chair for 2017. A as potential therapeutics, with dozens of X-ray crystal number of top-quality speakers are already locked in for structures providing insights into the recognition modes next year’s meeting, which is shaping-up to being another of the great many novel ‘receptors’ his laboratory has highlight on the Australian scientific calendar. developed. Structure-based protein design was a feature Doug Fairlie (Olivia Newton-John of talks from International invited speakers Michael Cancer Research Institute, VIC) and Weiss (Case Western), Jeffrey Boyington (NIH) and Bill Charlie Bond (University of Western Australia) DeGrado (UCSF), whilst NMR got a look-in with in-cell Photo credit: Stephanie Pradier, Monash University, VIC studies from Mikael Oliveberg (Stockholm University) Call for Conference Reports showing how cellular components influence the folding landscape of proteins. As is often the case, a highlight was the lecture presented If you’ve recently organised or attended a by the Leach medallist, an award that recognises conference that you think might be of interest to ASBMB members, please consider submitting scientific excellence in the field of protein science. This Australian Biochemist year’s recipient was Professor Bruce Kemp (St Vincent’s a conference report to the Institute), one of Australia’s foremost protein scientists, for publication. Reports should be between who has made many critical discoveries in the area of 750–1000 words and written in a friendly, kinases and especially, AMP-activated protein kinase. easy to read style. 2–3 photos (with captions) Professor Kemp’s lecture provided an entertaining and to accompany the text would also be most informative overview of his stellar career spanning four welcome. Any photos should be in JPG format decades – from the very early days of kinase research at higher than 300 dpi resolution. (when there were fewer than an handful of known If you require further information, please kinases), through to the recent important structural work contact the Editor ([email protected]). he and his collaborators are currently doing at SVI. It

Page 38 AUSTRALIAN BIOCHEMIST Vol 47 No 1 April 2016 FAOBMB meets with the Society of Biological Chemists IN India

Leann Tilley, FAOBMB Representative, reports on the FAOBMB Congress and Council Meeting held in Hyderabad, India, during November 2015.

The 14th Federation of Asian and Oceanian Biochemists drug targets and ‘off-targets’. Other highlight talks and Molecular Biologists (FAOBMB) Congress and 84th included the IUBMB Plenary Lecture by Ulrich Hartl, Annual Meeting of the Society for Biological Chemistry, Germany, describing the role of the nuclear compartment India (SBC-I) International (27–30 November 2015) was as a ‘safe-place’ for protein aggregates as part of the organized by the Centre for Cellular and Molecular homeostasis mechanisms; and the Plenary from Kanurty Biology (CCMB), Hyderabad, India. CCMB undertakes Rao (International Centre for Genetic Engineering and research activities in Biomedicine and Biotechnology Biotechnology, Delhi; ICGEB) who described the ways with a particular emphasis on providing centralised in which Mycobacterium tuberculosis manipulates the national facilities for new techniques in interdisciplinary central carbon metabolism of its host cell, inducing an research. The meeting was held at the Birla Institute of accumulation of neutral lipids, to counteract bactericidal Technology & Science (BITS Pilani, Hyderabad) on the pathways such as autophagy. Dr Manajit Hayer-Hartl, outskirts of Hyderabad. The meeting theme was: ‘Current Max Planck Institute of Biochemistry, described the Excitements in Biochemistry and Molecular Biology for complex chaperone machinery required for assembly Agriculture and Medicine’. Scientific sessions focused on of RuBisCO, the major protein in plants responsible for basic scientific discoveries, emerging research areas, and fixation of CO2. new synergies in translational sciences. The winner of the 2015 The Organizing Committee, chaired by Dr Ch. Mohan FAOBMB Award for Rao, Director of CCMB, and involving co-Chair, CCMB Research Excellence was group leader Dr K. Thangaraj, an international advisory David Craik, University committee and a number of enthusiastic volunteer helpers, of Queensland. David’s delivered a highly successful and enjoyable meeting, memorable talk on the both scientifically and socially. The food was particularly discovery and applications memorable. There were about 500 participants, including of cyclic peptides in 40 international speakers. biomedicine described the journey from basic biology to translation. His talk was Professor enlivened by images of Kiyoshi Fukui his heroic efforts to collect (FAOBMB cyclic peptides in different Professor David Craik, President) parts of the world, as well recipient of the 2015 FAOBMB on the left, as by some electrical failure Award for Research Excellence. congratulates issues. Professor Dr Martin Stone (Monash University, Australia) was Andy Wang a key speaker in the IUBMB-supported Education as IUBMB Symposium and talked about his efforts to prepare PhD President- students for careers beyond the standard research career Elect. path. The meeting was preceded by a Young Scientist Program (YSP) (24–26 November 2015), with 27 participants. The meeting commenced with the announcement of These included six Australians: two ASBMB fellowship President-Elect for the International Union of Biochemists recipients, Ash Hoque (University of Melbourne) and and Molecular Biologists (IUBMB); this was Professor Yogesh Khandokar (Charles Sturt University), two Andrew Wang, a Distinguished Research Fellow in the FAOBMB fellowship recipients, Justin Wong (Centenary Institute of Biological Chemistry at Academia Sinica, Institute, University of Sydney) and Rajesh Ghai (Victor Taiwan. Professor Wang is a well-known structural Chang Cardiac Research Institute), and two FAOBMB biologist, and Past-President of FAOBMB. Young Scientist Awardees, Urmi Dhagat (St Vincent’s Highlights of the scientific program included the twelve Medical Research Institute, Melbourne) and Victor Plenary or Award talks, including the Kunio Yagi Lecture Anggono (Queensland Brain Institute), all of whom spoke from Professor Shubha Tole, India, describing a single at the YSP Program. Urmi and Victor also gave excellent master transcription factor that controls the formation talks as part of the main program, describing their work on of the cerebral cortex, providing a blueprint for building interleukin-3 receptor signalling and glutamate receptor the brain. The FEBS Keynote was presented by Giulio trafficking, respectively. Superti-Furga, Director of CeMM, the Research Center The FAOBMB Council Meeting preceded the official for Molecular Medicine, Vienna, and described work Congress program (Thursday 26 November), with Leann on the molecular and systems biology of drug action, Tilley as the Australian delegate (ASBMB representative). including chemical proteomics approaches to determine The meeting was attended by delegates from 14 of the Vol 47 No 1 April 2016 AUSTRALIAN BIOCHEMIST Page 39 FAOBMB meets with the Society of Biological Chemists IN India

chaired by Leann Tilley, to develop a gender equality policy. The policy indicates that membership of FAOBMB- sponsored conference/ workshop organising committees, conference theme coordinators, symposium chairs and invited plenary speakers, should be at least 40% female and at least 40% male. In addition, opportunities should be provided for early-/mid-career researchers to be symposium chairs. Kiyoshi Fukui and IUBMB President- elect, Andy Wang, were very supportive and the Council approved the proposed FAOBMB Gender Equality Policy. The 25th FAOBMB Conference will be held in Manila, Australian winners of the FAOBMB Young Scientist Awards, Philippines, in December, 2016. In the year following Dr Urmi Dhagat and Dr Victor Anggono (second and third from the 26th FAOBMB Conference in Japan in 2017, the next left, respectively), with Professor Kiyoshi Fukui (President) and triennial IUBMB Congress will be held as a joint meeting Professor Sheila Nathan (Acting Secretary-General). with the FAOBMB Congress in Seoul, Korea, 4–9 June 21 constituent member countries and was chaired by the 2018. FAOBMB President, Professor Kiyoshi Fukui (Japan), and Acting Secretary-General Sheila Nathan (Malaysia). Kiyoshi Fukui announced Professor Zengyi Chang as President Elect for FAOBMB, commencing in January 2016. The President promoted the Exchange Fellowships, Visiting Lectureships and Travel Lectureship as a great way for students/ postdocs and more senior scientists to interact with colleagues in another FAOBMB country. Inspired by initiatives developed by the ASBMB, the Council had previously established a Working Group, FAOBMB Executive Committee and Council members, and Congress Convenors.

Australian Society for Biochemistry and Molecular Biology Inc PUBLICATION SCHEDULE FOR AUSTRALIAN BIOCHEMIST, volume 47, 2016

Issue ASBMB Content Copy Deadline Issue Date

April 2016 47 (1) Profiles of medal, award and fellowship winners Monday 8th February Monday 28th March Nominations for Executive/Council

August 2016 47 (2) Nominations for medals, awards and fellowships Monday 13th June Monday 25th July Notice of AGM/proposed constitutional changes

December 2016 47 (3) Annual Reports/finances Monday 10th October Monday 29th November ComBio2016 reports

The Program for ComBio2016 (Brisbane) will be placed on the ASBMB webpage (www.asbmb.org.au). The Proceedings of the Australian Society for Biochemistry and Molecular Biology is published in conjunction with the Annual Conference of the Society. The electronic version of the Proceedings (Volume 48) will be made available online.

Page 40 AUSTRALIAN BIOCHEMIST Vol 47 No 1 April 2016 Young Scientist Program in Hyderabad, the City of Pearls

Ash Hoque and Yogesh Khandokar, ASBMB–FAOBMB YSP Travel Fellowship awardees, report on the FAOBMB Young Scientist Program and Congress held in Hyderabad, India.

It was a huge honour for each of us to receive an ASBMB– FAOBMB YSP Travel Fellowship to attend the Young Scientist Program (YSP) (24–26 November 2015) and the 14th FAOBMB Congress (27–30 November 2015) in Hyderabad, India. The YSP was hosted by the Centre for Cellular and Molecular Biology (CCMB) in Hyderabad. The YSP event initiated collaborative networks among the 27 young scientists from the Asia and Oceania region with the two-day scientific program on 24 and 25 November. A half-day tour to the LaCONES (Laboratory for the Conservation of Endangered Species) laboratories and CCMB Foundation Day celebrations followed on 26 November. Ash Hoque outside BITS Pilani, Hyderabad. The organising committee led by Dr A.S. Sreedhar of CCMB put up a wonderful program and arranged warm hospitality for all the YSP participants. This commenced with the welcome at the airport (even late at night), continued with very good food and a hassle- free stay at the NGRI and CCMB guest houses, until we were safely dropped off at the airport for our homeward flights. We had two days of exciting oral presentations from the participants, with poster sessions during lunch breaks, where we got the chance to learn about each of the projects in detail. The oral sessions were chaired by Yogesh Khandokar receives the ASBMB-FAOBMB YSP Travel Professor Sheila Nathan (FAOBMB Fellowships Chair). Fellowship certificate from Professor Kiyoshi Fukui (FAOBMB We were very pleased to have exciting invited lectures President, right ), with Dr Ch. Mohan Rao (Congress Chair, centre). from Dr Jyotsna Dhawan of DBT-iStem, who talked about the quiescent state of adult stem cells, Professor Soumyadipta Pyne of the Indian Institute of Public Health who discussed modelling of high-resolution and high-throughput data even at single-cell level and Dr J. Gowrishankar of the Centre for DNA Fingerprinting and Diagnostics (CDFD, Hyderabad) who spoke on central dogma and Darwinian evolution. We appreciated CCMB as the optimal venue for hosting the YSP. Indeed, it was so exciting to see under one roof at the CCMB all sorts of cutting-edge facilities (high resolution microscopy, high YSP participants aiming high. throughput mass spectrometry, animal house facilities and excellent lab facilities). It was especially interesting to learn that all the CCMB students have unlimited access to all these resources. Despite the packed schedule we managed to go for sightseeing outings on the first evening. We watched the sound and light show at Golconda Fort in Hyderabad and we were thrilled to learn about its history. We also had a wonderful river cruise at Hussain Sagar Lake followed by a great dinner on the boat. Our visit to LaCONES on the third day was another important part of YSP. We were really excited to learn about the LaCONES laboratories where scientists are working on ‘Conservation Biotechnology’ that has provided insights on propagation of endemic endangered animals, storage of gametes and successful reintroduction of wild animals. The CCMB YSP participants on the rocks. Foundation Day celebrations followed in the afternoon, Vol 47 No 1 April 2016 AUSTRALIAN BIOCHEMIST Page 41 Young Scientist Program in Hyderabad, the City of Pearls with several speeches and lectures after which we had a Congress to be a big success. We feel very proud to have great roof-top dinner at CCMB Terrace with more than been a part of these exciting events. We thank ASBMB 1,000 guests. and FAOBMB, together with the YSP organisers, for The YSP was very interactive and somewhat the opportunity to travel to India for the YSP and the overwhelming after three days. Once we managed to Congress. learn the names of each of the other YSP participants, we Ashfaqul Hoque is a final year PhD student at the had great discussions about our scientific research, career Department of Biochemistry and Molecular Biology, goals and pathways, and possible future collaborations. University of Melbourne. He completed his BSc (Honours) On the morning of 27 November, we headed to BITS and MS in Microbiology at the University of Dhaka before Pilani, Hyderabad, to attend the 14th FAOBMB Congress. he joined the International Centre for Diarrhoeal Disease YSP participants stayed together at the Leonia Resorts Research, Bangladesh, in 2009. during the Congress. We enjoyed the Congress, where we Yogesh Khandokar is a final year PhD student in the presented our work in posters; two of the YSP participants Graham Centre for Agricultural Innovation, Charles Dr Urmi Dhagat and Dr Victor Anggono (recipients of Sturt University. He did BSc and MSc in Biotechnology the 2015 FAOBMB Young Scientist Awards), gave oral from at Sant Gadge Baba Amravati University, India. presentations at the Congress. We cannot forget the great food we had for the whole week. We had a great time overall and we considered the YSP and the 14th FAOBMB

Professor Sheila Nathan (left) chairing a YSP session with Assembled YSP participants and Faculty at CCMB, Hyderabad. Dr A.S. Sreedhar (centre) and Dr Ch. Mohan Rao (right).

FORTHCOMING MEETINGS

16th IUBMB Conference ComBio2016 • Genomes and Bioinformatics 17–21 July 2016 3–7 October 2016 • Cell Biology Vancouver, Canada • Cell Signalling Brisbane Convention and Exhibition The theme of the Conference is • Biochemistry and Metabolism Centre, Brisbane Signalling Pathways in Development, • Emerging Areas, Hot Topics Early Registration and Abstract Disease and Ageing. A Young and Enabling Technologies Deadline: Monday, 27 June 2016 Scientists Program (YSP) will be • Education and Career held; details to be advised. Development The program will feature 14 plenary Further information: presentations from some of the best www.iubmb2016.org There will be the three core ComBio international scientists (please see the Email: [email protected] societies participating (ASBMB, last page of this magazine for details), ASPS and ANZSCDB). plus a number of society speciality lectures. Several poster sessions are 25th FAOBMB also planned. The scientific program Further information: www.asbmb.org.au/combio2016 Conference of the conference will include the 5–7 December 2016 themes: Scientific Program: Manila, Philippines The theme of the Conference is • Plant Cell and Developmental Joe Rothnagel [email protected] Biochemistry and Molecular Biology Biology and Genetics in Health and Wellness. • Plant Physiology and Ecology Further information: • Developmental, Stem Cell and Registration/Exhibition www.psbmb.org Regenerative Biology Sally Jay Email: [email protected] • Proteins and Proteomics [email protected]

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field of cell biology. Some of these applications include: Disclaimer • Cell morphology The Australian Biochemist is • Intracellular processes published by the Australian Society • Cell–cell and cell–microorganism for Biochemistry and Molecular interactions Biology Inc. The opinions expressed • Nanoparticle uptake in this magazine do not necessarily • Tissue imaging represent the views of the Australian Society for Biochemistry and AXT Pty Ltd Molecular Biology Inc. Holographic Microscopy – See [email protected] Inside Living Cells Like Never www.axt.com.au Before Introducing a revolutionary way to instantly see inside living cells, in real time, in 3D and label-free! The disruptive technique of holographic microscopy works by passing light through the cell and recording the resulting light wave Integrated Solutions for Stem front. By collecting multiple packets of Cell Discovery information from multiple angles, we Regardless of the specific path your stem are able to reconstruct a 3D image of the LabGear Australia is the exclusive cell research takes, Enzo Life Sciences cell with incredible clarity. Australian distributor for the xxpress® provides innovative tools to help you Holographic microscopy enables real-time thermal cycler by BJS realise its full potential. us to measure cells in their native Technologies Ltd. The xxpress® qPCR Differentiate – Modulate Stem Cell Fate environment, with the advantages of instrument easily does 40 cycles of qPCR with Small Molecules and Proteins being: in under 10 minutes making it the fastest • Label-free qPCR instrument on the market. The Stem cell self-renewal and differentiation • Non-invasive instrument achieves this impressive involves complex events which lead to the • Interference-free feature using an ultra-low thermal mass generation of various phenotypes. Enzo • Low in photobleaching and high conductivity xxplates which offers an extensive collection of small • Fast come in 3 formats of 24, 54 and 96 wells molecules and proteins such as growth • 3D and 4D! easily covering workflows from low to factors and cytokines that can be used The Nanolive 3D Cell Explorer high throughput. to influence differentiation, biochemical is a self-adjusting, holographic and activity, and cell growth. The benefit of having exceptional tomographic microscope. Through temperature uniformity of better than Detect – Gene Expression Analysis, a combination of holography and 0.3°C during cycling is the acquisition Wnt Pathway Analysis, Epigenetics, rotational scanning the system precisely of qPCR results that you can trust and Live Cell Analysis and Worry-free measures the distribution of the physical will meet all MIQE guidelines, thereby, Antibodies refractive index (RI) within the cell. It minimising the bottleneck of repeating • Maximise genomic data with does this in 3D and in real-time with experiments. One of the instruments minimal sample input. impressive results. Being label-free, strengths is the active thermal • Discover Wnt pathway modulators cells are seen in their natural state. The management using highly accurate IR affecting stem cell development. innovative software package (STEVE) is sensors to measure and adjust sample • Robust epigenetic screening assays then able to digitally stain the images. temperature 100 times per second using for stem cell regulators. The TESCAN Q-Phase is a its patented algorithm. • Multiplex analysis of cell fate and multimodal, coherence-controlled differentiation by flow cytometry. holographic microscope for An easy to use intuitive user interface • Antibodies for identification of stem quantitative phase imaging (QPI). allows you to program runs the way cell populations. It uses incoherent light sources, you think via a colour touch screen. For further information please contact enabling imaging of samples in turbid The open platform allows users to United Bioresearch, Enzo’s Australian media and is purposefully designed continue running existing protocols partner. to observe living cells in vitro. Based with known and trusted qPCR reagents on a robust inverted transmission providing continuity now and flexibility United Bioresearch Products microscope platform, the whole system in the future. For more information, Kirrily Smith is situated in an incubator and includes please contact LabGear Australia on Phone (02) 4575 0309 full motorisation. 1800LABGEAR (1800 522 432) or email [email protected] [email protected] Holographic microscopy has wide www.enzolifesciences.com application, particularly within the

Vol 47 No 1 April 2016 AUSTRALIAN BIOCHEMIST Page 43 OUR SUSTAINING MEMBERS

iPSCs. Tested cell lines for spheroid can do this! Now Delong has turned its culture include HeLa, HepG2, LNCaP attention to 25 Kv Model – LVEM25; and HEK-293. Other tested cell lines a great fit without being hung up and include Vero, MDCKII, CaCo-2, CHO, concerned about getting systems that STO-DA. take up complete rooms. Does your High Throughput With non leachable properties, this Don’t forget about the RBC DNA Screening Lab Require the range is available as culture dishes (three extraction system, ISOGEN Imagers and Ultimate in Speed and sizes) and 6, 24, 48, 96 and 384 well plate DAICEL Chiral Columns. Talk to us or Sensitivity? formats. All items have a two year shelf website is at hand! BMG LABTECH introduces its new life, are non-cytotoxic, non-pyrogenic For further information contact:- product classified as the “gold standard” and are free of detectable DNase, RNase Scientex Pty Ltd ® microplate reader, the PHERAstar FSX. and human DNA. Tel 03 9899 6100 This latest evolution in microplate Please contact your local Interpath Fax 03 9899 6122 reader instrumentation continues the Services Sales Executive for further Email [email protected] success of the top selling PHERAstar information or a sample. www.scientex.com.au ® range with the new PHERAstar FSX. Interpath Services Providing real time dual colour readings Phone: (03) 9457 6277 in all fluorescence and luminescence Freecall: 1800 626 369 modes and the best sensitivity on the Email: [email protected] market. With the highest reading speed and unmatched flexibility in plate formats up to 3456 wells. Ebro EBI330 Transport Data The PHERAstar® FSX has improved Loggers features that make it superior to any other microplate reader currently on the VWR International is pleased to market. The reader performs all leading present the Ebro Single Use EBI 330 data non-isotopic detection technologies with loggers for monitoring temperature ease and speed. Unmatched sensitivity of food and other temperature-critical and speed in Fluorescence Intensity products during the transportation (FI) and Fluorescence Polarization process. ® Your Blue Science for R&D (FP) makes the PHERAstar FSX With the two single-use data loggers, the most sensitive reader today. The Hi, did you see the one about the EBI 330-T30 (-30°C to +60°C) and EBI reader offers an ultrahigh dynamic alternative to sample preservation from 330-T85 (-85°C to +50°C), Ebro extends range in Luminescence enabling a DENATOR-Stabilizor? If not it is well its EBI 300 family with two more data higher flexibility and more precise consideration for a Disrupter Technique loggers with USB plug and automatic measurement results. that allows great savings in freezer space. PDF generation. An outstanding feature No need to say more just look it up and The PHERAstar® FSX is the ultimate is the deep temperature measurement make up your own mind. microplate reader for HTS applications and down to -85°C with the EBI 330-T85. assay development. By the way, YOUR BLUE SCIENCE, is The single-use data loggers allow the our way of getting good stuff about Life Experience the “gold standard” and temperature control and documentation Science, because Life relates to our Blue book a demo today by emailing your in the food industry according to EN Planet! Nice way of looking at it and details to [email protected] 12830 as well as the transport monitoring moving away from the crowd using the of medicine, blood plasma and other term “LIFE this” and “LIFE that”! pharmaceutical products. These loggers APPLIED PHOTOPHYSICS continues are ideal for situations where return of its developments in Circular Dichroism multi-use loggers to the sender is either and recently introduced the Chirascan difficult or impractical because of efforts qCD for Quantitative CD with and costs. Cell-Repellent Surface from patented non-chemical, multi-point Greiner Bio-One Speak to your local VWR Equipment CD calibration. The Chirascan has been Specialist for further details. This surface technology inhibits cell the instrument of choice because of adhesion, making the range ideal for its excellent sensitivity and there are VWR International Pty Ltd culturing spheroid cultures and stem cell significant installs in Australia. Tel: 1300 727 696 aggregates, as well as suspension culture Fax: 1300 135 123 DELONG has long been involved in of semi-adherent and adherent cell lines. Email: [email protected] benchtop Low Voltage EM’s with TEM Web: au.vwr.com Tested cell lines for stem cell aggregate capabilities to complement SEM in the formation include mES-D3 and human model LVEM5. It is the only one that

Page 44 AUSTRALIAN BIOCHEMIST Vol 47 No 1 April 2016 OUR SUSTAINING MEMBERS DAINTREE scientific AUSTRALIA New England Biolabs and Daintree welcomes two new molecular Genesearch Bring You biology products to the range this month Monarch™ Nucleic Acid – we are excited to introduce two highly Purification Kits effective and affordable methods of sample preparation, the It’s time to transform your DNA Tissue Grinder GRYPHAX® NAOS 20 MP and the . purification experience. NEB’s Monarch Cellcrusher™ Microscopy Camera Nucleic Acid Purification Kits are The new handheld Portable Tissue The PROGRES GRYPHAX® NAOS optimized for maximum performance Grinder features either 12V DC power 20 MP microscopy camera is the latest and minimal environmental impact. or a Lithium battery capable of 10 hours addition to the portfolio of Jenoptik Providing fast and reliable purification of continuous use making the grinder PROGRES cameras made for routine of high quality DNA from bacterial perfect for field use or in laboratories microscopic applications. Using a back cultures, agarose gels, and enzymatic with limited power points. illuminated, sensitive, scientific grade reactions using best-in-class technology. Three models are available; G10 high 1” CMOS sensor, USB 3.0 interface and Our unique column design offers speed (12000rpm), G20 low speed included software, this camera provides elution in lower volumes than standard (1200rpm) and adjustable speed G50 fast live images and is ideal for use in purification kits, providing concentrated, from 3000–8000rpm. The replaceable science, research and industry. high quality DNA suitable for use plastic grinder pestles are autoclavable, The PROGRES GRYPHAX® NAOS in downstream applications such as helping to avoid cross-contamination. camera provides fast live images DNA sequencing, PCR, restriction The grinder is one of the most powerful at short exposure times, brilliant enzyme digests and other enzymatic motor driven grinders on the market, and colour reproduction and excellent manipulations. Our column design also a perfect solution for DNA extraction. image quality. Position and focus the eliminates buffer retention and the risk The Cellcrusher™ features an preparation being examined quickly of carryover contamination, providing innovative tissue cryopulveriser design and easily. Even moving objects can be fast, worry-free DNA purification. with curved inner surfaces which allow analyzed with minimal effort. They can Designed with sustainability in mind, for effective sample recovery. The curved be used on any microscope and are also Monarch kits use significantly less plastic surfaces also facilitate easy cleaning. compatible with Windows, MacOS and and responsibly-sourced packaging. Physically tough samples such as Linux operating systems. cartilage and venous tissue are routinely For more information visit: https:// The user-friendly software programs disrupted. Particularly tough tissues www.neb.com/monarch/monarch- are easy to install and intuitive to like seeds or skin can utilise the Drill- nucleic-acid-purification-kits use. A host of additional features Bit accessory for secondary grinding. and parameter settings allow you to Available kits include: Customers report the Cellcrusher™ is so optimally capture, process, analyze and • Monarch Plasmid Miniprep Kit much faster than a pestle and mortar and document images. The camera features • Monarch DNA Gel Extraction Kit uses far less muscle work for extraction a modern USB 3.0 SuperSpeed interface • Monarch PCR & DNA Cleanup Kit on smaller tissues for DNA and RNA. that makes the camera easy to connect (5 μg) For information please contact to your PC or laptop. For a limited time, try one of our Murdoch Macaskill Benefits include high image quality, Monarch Nucleic Acid Purification Kits. Daintree Scientific Australia ease of use, compatible with Win/MAC/ Request your sample at https://www.neb. Phone (03) 6376 3335 LinUX and German manufactured and com/monarch/try-a-monarch-sample Email [email protected] includes a two year warranty. www.daintreescientific.com.au For ordering information please visit SciTech Pty Ltd www.genesearch.com.au or contact us (03) 9480 4999 on 1800 074 278. Also available via your [email protected] local Genesearch e•Freezer! www.scitech.com.au

Vol 47 No 1 April 2016 AUSTRALIAN BIOCHEMIST Page 45 OUR SUSTAINING MEMBERS immunofluorescence (IF) and histological Sapphire Bioscience staining (IHC). Nano-Boosters are the www.sapphirebioscience.com Nano-Trap GFP or RFP antibodies [email protected] coupled to a range of the ATTO-TEC Ph: +61 2 9896 2022 fluorescent dyes. Monoclonal Antibodies for BioNovus Life Sciences GFP, RFP and Tag Proteins, Ph: (02) 9484-0931 Epigenetic Markers and RNA Email: [email protected] Polymerase II CTD Web: www.bionovuslifesciences.com.au ChromoTek’s products are centred Scanning Electron Microscope around the proprietary Chromobody® (SEM) with EDX for Elemental technology. These innovative single Analysis domain camelid antibody fragments Hitachi have released the latest are either coupled to a matrix (Nano- Sapphire Bioscience partners model of their tabletop SEM range, the Traps); to a fluorescent dye (Nano- with Avanti Polar Lipids, TM3030Plus. This SEM features an Ultra Booster); fused to a fluorescent Antibodies Inc and Diaclone Variable-pressure SE Detector (UVD) protein (Chromobodies®) or are part for low-vacuum SE imaging and a high of an assay system to study protein– Sapphire Bioscience is proud to sensitivity 4-segment BSE detector for protein interactions (F2H-Assay). announce distribution partnerships high quality compositional imaging. Their extremely small size and high with Avanti Polar Lipids, Antibodies Low-vacuum minimises moisture stability make them ideal reagents for Incorporated and Diaclone in Australia evaporation and shrinkage of samples, biomolecular research and live cell and New Zealand. These partnerships reducing the need for chemical assays. are a part of Sapphire’s continued processing and Critical Point Drying commitment to source high quality and In addition to their range of Alpaca (CPD). 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ESGRO® – 2500 Publications and Counting Knockout (KO) Validated Merck provides the largest and most comprehensive range of products for Antibodies: Specificity NEW! Animal Origin Free Types your mouse ES cell culture needs. Confirmed AFA, AFB and AFC, STZ1 and STZ2 collagenases are derived from cultures ® Highlights include the unique ESGRO The Need grown in medium completely devoid Antibodies are amongst the most supplement, a special formulation of of animal based components and commonly used tools in life science mouse LIF protein. Unlike regular LIF, designed for bioprocessing applications to study proteins and their functions which is sold by weight, each lot of where introduction of potential animal within various biological pathways ESGRO supplement is sold based on derived pathogens must be prevented. and diseases. However, an increasing its biological activity for reproducible Levels of secondary proteases are number of studies have shown that not results. The benefits of using ESGRO similar to Types 1 and 2 collagenase. mLIF medium supplement include: all of these tools are specific, leading • Consistent inhibition of ES cell to a growing issue of experimental • CLSAFA is the original AFA grade differentiation irreproducibility, which is leading designed to have collagenase and • No batch-to-batch variation to wasted time, resources and, secondary proteases similar to • Feeder-free cell culture unfortunately, the retraction of papers Types 1 and 2 collagenase. within the scientific community. • CLSAFB contains higher For certain cell lines, the use of ESGRO collagenase and caseinase activities supplement allows feeder-free cell The Solution: Knockout Validation than CLSAFA. One of the most accepted and trusted culture, saving time and giving you • CLSAFC has especially low validation processes for antibody more control over your cells. tryptic activity similar to Type 4 specificity is through knockout ® collagenase. Further complementing the ESGRO validation. Antibody specificity is • STZ1 and STZ2, 0.22μ filtered supplements are: confirmed by testing the antibody of STEMzyme™ AOF collagenase/ • ESGRO Complete PLUS, a serum- interest on a knockout sample or cell neutral protease (Dispase®) blends free media for the maintenance and line, which doesn’t express the target for primary and stem cell isolation. derivation of mouse ES cell lines in protein. Knockout validation offers a the absence of FBS and feeder cells true negative control to confirm the ScimaR • EsGRO-2i medium is suitable for ES antibody specificity to the protein Email [email protected] and iPS cell derivation, passaging of interest. We have incorporated Freecall 1800 639 364 and maintenance. This media can knockout based validation as a standard Phone 03 9842 3386 be used on feeder dependent and level of validation for our new antibody Fax 03 9842 3407 independent pluirpotent stem cell products. When you see the knockout lines. validated seal, you can trust that the • RESGRO™, a culture medium for antibody has not only been validated the rescue of partially differentiated in the recommended applications ES cell lines and improved ES cell and species, but its specificity has derivation. been confirmed through our in-house For more information contact knockout validation approach. Merck on 1800 335 571 or visit Discover more at Abcam www.merckmillipore.com www.abcam.com/ko Email: [email protected]

Vol 47 No 1 April 2016 AUSTRALIAN BIOCHEMIST Page 47 DIRECTORY COUNCIL FOR 2016 STATE SPECIAL INTEREST PRESIDENT REPRESENTATIVES GROUPS Professor Michael Ryan Department of Biochemistry and AUSTRALIAN CAPITAL TERRITORY ADELAIDE PROTEIN GROUP Molecular Biology Dr Peter Mabbitt Contact: Dr Christopher McDevitt Monash University ANU College of Physical and Mathematical Research Centre for Infectious Diseases CLAYTON VIC 3800 Sciences University of Adelaide Ph (03) 9902 4909 Australian National University ADELAIDE SA 5005 Email: [email protected] CANBERRA ACT 0200 Ph (08) 8313 0413 Email: [email protected] Email: [email protected] PRESIDENT ELECT Professor Leann Tilley NEW SOUTH WALES AUSTRALIAN YEAST GROUP Department of Biochemistry and Dr Katharine Michie Chair: Dr Alan Munn Molecular Biology School of Physics Griffith University Gold Coast University of Melbourne University of New South Wales PMB 50, Gold Coast Mail Centre PARKVILLE VIC 3010 SYDNEY NSW 2052 SOUTHPORT QLD 9726 Ph (03) 8344 2227 Ph (02) 9385 4587 Ph (07) 07 5552 9307 Email: [email protected] Email: [email protected] Email: [email protected] TREASURER QUEENSLAND BIOCHEMICAL EDUCATION Associate Professor Terrence Piva Dr Dominic Chi Hiung Ng Chair: Associate Professor Susan Rowland School of Medical Sciences School of Biomedical Sciences School of Chemistry and Molecular Biosciences RMIT University, PO Box 71 University of Queensland University of Queensland BUNDOORA VIC 3083 ST LUCIA QUEENSLAND 4072 ST LUCIA QLD 4072 Ph (03) 9925 6503 Ph (07) 3365 3077 Ph: (07) 3365 4615 Email: [email protected] Email: [email protected] Email: [email protected] SECRETARY SOUTH AUSTRALIA MELBOURNE PROTEIN GROUP Associate Professor Briony Forbes Dr Steven Polyak Chair: Dr Marc Kvansakul Medicinal Biochemistry School of Molecular and Biomedical Science Department of Biochemistry Flinders University University of Adelaide La Trobe University BEDFORD PARK SA 5042 ADELAIDE SA 5005 BUNDOORA VIC 3086 Ph (08) 8204 4221 Ph (08) 8313 6042 Ph (03) 9479 2263 Email: [email protected] Email: [email protected] Email: [email protected] EDITOR TASMANIA METABOLISM AND MOLECULAR Dr Chu Kong Liew Dr Adele Holloway MEDICINE GROUP Email: [email protected] School of Medicine, University of Tasmania Chair: Dr Nigel Turner HOBART TAS 7008 School of Medical Science EDUCATION REPRESENTATIVE Ph (03) 6226 2670 University of New South Wales Associate Professor Janet Macaulay Email: [email protected] Department of Biochemistry and KENSINGTON NSW 2052 Ph (02) 9385 2548 Molecular Biology, Monash University VICTORIA Email: [email protected] CLAYTON VIC 3800 Dr Diana Stojanovski Ph (03) 9905 3730 Biochemistry and Molecular Biology QUEENSLAND PROTEIN GROUP Email: [email protected] Bio21 Institute Chair: Dr Brett Collins University of Melbourne Institute for Molecular Bioscience FAOBMB REPRESENTATIVE PARKVILLE VIC 3010 University of QLD, ST LUCIA QLD 4072 Professor Paul Gleeson Ph (03) 9035 3197 Ph (07) 3346 2043 Department of Biochemistry and Email: [email protected] Molecular Biology Email: [email protected] University of Melbourne WESTERN AUSTRALIA RNA NETWORK AUSTRALASIA PARKVILLE VIC 3010 Associate Professor Nicolas Taylor Chair: Dr Archa Fox Ph (03) 8344 2354 ARC Centre of Excellence in Plant Energy Harry Perkins Institute of Medical Research Email: [email protected] Biology 6 Verdun Street University of Western Australia NEDLANDS WA 6009 SECRETARY FOR CRAWLEY WA 6009 Ph (08) 6151 0762 SUSTAINING MEMBERS Ph (08) 6488 7005 Email: [email protected] Sally Jay Email: [email protected] c/- ASBMB National Office SYDNEY PROTEIN GROUP PO Box 2331 Chair: Dr Liza Cubeddu KENT TOWN SA 5071 School of Science and Health, University of Ph (08) 8362 0009 Western Sydney, PENRITH NSW 2751 Fax (08) 8362 0009 Ph (02) 4620 3343 Email: [email protected] Email: [email protected] ASBMB NATIONAL OFFICE PO Box 2331 KENT TOWN SA 5071 FOLLOW US ON: Ph (08) 8362 0009 Fax (08) 8362 0009 Email: [email protected] http://www.asbmb.org.au COPY DEADLINE FOR NEXT ISSUE: Monday, 13 June 2016

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