DOCSLIB.ORG

The Role of Pumilio 2 in Axonal Outgrowth by Dani Sarkis a Thesis

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text

Load more

The Role of Pumilio 2 in Axonal Outgrowth by Dani Sarkis A thesis submitted in conformity with the requirements for the degree of Master of Science Graduate Department of Physiology University of Toronto Copyright c 2012 by Dani Sarkis Abstract The Role of Pumilio 2 in Axonal Outgrowth Dani Sarkis Master of Science Graduate Department of Physiology University of Toronto 2012 Pumilio 2 (PUM2) is a member of the Puf family of mRNA binding proteins and transla- tional regulators which are involved in various processes including embryonic patterning and memory formation. Nevertheless, its functions in the outgrowth of neuronal axons have not been studied. This study shows endogenous expression of PUM2 in neurites of dorsal root ganglia (DRG) neurons and transport of PUM2 along retinal ganglion cell (RGC) axons and their growth cones. Overexpression of PUM2 in DRG neurons resulted in shorter axons when compared to control neurons. Expression of either dominant nega- tive mutation (dnPUM2) or PUM2W349G displayed a reduction in axonal length. PUM2 downregulation with microRNA (miRNA) also caused a reduction in neurite length com- pared to control neurons. Finally, PUM2 silencing did not alter eye size at E4, which allows investigation of axonal outgrowth in RGC in vivo. These results suggest a novel role for PUM2 in axonal outgrowth. ii Dedication To Ahed Khouri and Farid Sarkis, my mother and father. For everything you have done for me. For your unconditional love and support. To Gabi Sarkis, my brother, my best friend, and idol. iii Acknowledgements First and foremost, I would like to thank my supervisor, Dr. Philippe Monnier, for giving me the chance to work on this project. I would also like to thank my advisory committee members, Dr. Zhong-Ping Feng and Dr. James Eubanks, for their insight and for ensuring I was on the right track towards obtaining my MSc. I would also like to acknowledge the help and guidance of our research associate, Dr. Nardos Tassew, PhD. Dr. Tassew taught me the skills that I needed to complete this project, and had very enlightening discussions with me to help me better understand my project, and I am grateful for her patience and help. Thank you Dr. Tassew and Dr. Paromita Banerjee for your help with editing this thesis. I must also thank my friends Gemma Higgs, MSc for help with editing this thesis, Dr. Anne Wheeler, PhD for her helpful feedback and getting me into research in the first place, and Elena Sidorova, MSc for her continuous support and encouragement. This project was funded by two scholarships from the Vision Science Research Pro- gram (VSRP) at the Toronto Western Research Institute. iv Contents 1 Introduction 1 1.1 mRNA Translation . 1 1.1.1 Overview of mRNA Translation in Eukaryotes . 1 1.1.2 Local Translation, RNA transport, and mRNA Binding Proteins . 2 1.2 Axonal Outgrowth . 4 1.3 The Embryonic Chicken Dorsal Root Ganglia as a Model for Axonal Out- growth . 6 1.4 Pumilio - a Founding Member of the Puf Family . 7 1.5 Pumilio is Involved in Embryonic and Germline Development and Cell Cycle Regulation . 9 1.6 Pumilio 1 and Pumilio 2: Vertebrate Homologues of the Drosophila Pumilio 10 1.7 Pumilio 2 and the Eukaryotic Initiation Factor 4E (eIF4E) . 12 1.8 Pumilio in the Nervous System . 13 1.9 Study Rationale . 14 1.10 Hypothesis and Aims . 17 2 Materials and Methods 18 2.1 Cloning . 18 2.1.1 Cloning of dominant negative PUM2 (dnPUM2) . 18 2.1.2 Cloning of PUM2W349G-EYFP in pEYFP-N1 . 19 v 2.1.3 Cloning of PUM2 microRNA (PUM2miRNA) . 20 2.1.4 Cloning of chicken PUM2 in pcDNA3.1 (-)/myc-His A . 21 2.1.5 Cloning of eIF4E-T2A-PUM2EYFP . 22 2.2 Cell Culture . 23 2.2.1 Transfection of Cell Lines . 24 2.3 Chicken Embryos . 24 2.4 Dorsal Root Ganglia (DRG) Neurons . 25 2.4.1 Dissection . 25 2.4.2 Nucleofection . 25 2.5 Retinal Flat Mounts . 26 2.6 Virus Preparation . 26 2.7 Fiber Tracing . 27 2.8 In Ovo Electroporation . 27 2.9 Immuno-cytochemistry . 29 2.10 Western Blots . 29 2.11 Microscopy . 30 2.12 Live Imaging . 31 2.13 Statistical Analysis . 31 3 Results 32 3.1 Construct Verification . 32 3.2 PUM2 is transported along the axons in DRG neurons and RGCs . 32 3.3 PUM2 overexpression results in shorter axons in dissociated DRG neurons 33 3.4 Expression of dnPUM2 or PUM2W349G results in shorter axons in disso- ciated DRG neurons . 33 3.5 PUM2 silencing . 34 3.5.1 PUM2 silencing hinders axonal outgrowth in dissociated DRG neu- rons . 34 vi 3.5.2 PUM2 silencing does not affect eye size at E4 . 35 3.6 eIF4E fails to rescue the short axon phenotype . 35 4 Discussion 54 4.1 PUM2 localization in the growth cone and axons of RGCs and DRG neurons 55 4.2 Impaired axonal outgrowth in DRG neurons overexpressing PUM2 . 56 4.3 Impaired axonal outgrowth in DRG neurons expressing PUM2 mutants . 58 4.4 PUM2 silencing interferes with axonal outgrowth in DRG neurons . 60 4.5 eIF4E coexpression fails to rescue short axon phenotype . 61 4.6 Future Directions . 63 4.6.1 Are PUM2/Nos and PUM2/mRNA interactions necessary for nor- mal axonal outgrowth? . 63 4.6.2 Can the short axon phenotype be rescued? . 63 4.6.3 What is the cause of the short axon phenotype? . 65 4.6.4 Testing the role of PUM2 in axonal outgrowth and guidance in vivo 66 4.7 Conclusion . 67 References 67 vii List of Figures 1.1 In Situ hybridization showing the expression of Pum2 transcript in E9 eye 15 1.2 PUM2 Expression in Axons and Growth Cones of Retinal Ganglion Cells 16 2.1 Schematic Representation of eIF4E-T2A-PUM2EYFP in pT2K . 23 2.2 In Ovo Electroporation of E1.5 Embryos . 28 3.1 Western blot verifying the provided and cloned constructs . 37 3.2 Western blot verifying PUM2W349G-EYFP . 38 3.3 PUM2 is transported along the axon of RGCs . 38 3.4 PUM2 is endogenously expressed in DRG neurons . 39 3.5 PUM2 Overexpression Results in Short Axons . 40 3.6 Quantification of Axon Length in PUM2 Overexpression . 41 3.7 DRG neurons expressing dnPUM2 have shorter axons . 42 3.8 Quantification of Axonal Length in dnPUM2 Overexpression . 43 3.9 DRG neurons expressing PUM2W349G have short axons . 44 3.10 Quantification of Axonal Length in PUM2W349G Overexpression . 45 3.11 PUM2 Silencing with miRNA constructs . 46 3.12 PUM2miRNA transfected neurons had shorter axons . 47 3.13 Quantification of Axonal Length After PUM2 Silencing . 48 3.14 PUM2 Silencing and Eye Size . 49 3.15 Quantification of the effects of PUM2 silencing on eye size at E4 . 50 viii 3.16 Western Blot Confirming Coexpression of eIF4E and PUM2EYFP . 51 3.17 DRG neurons coexpressing eIF4E and PUM2 have short axons . 52 3.18 Quantification of Axonal Length in eIF4E and PUM2 Coexpression . 53 4.1 Known interactions of PUM2 and possible involvement in axonal outgrowth 64 4.2 Time Lapse Imaging of DRG Neurons . 68 ix Chapter 1 Introduction 1.1 mRNA Translation 1.1.1 Overview of mRNA Translation in Eukaryotes Messenger RNA (mRNA) translation is a stage of protein biosynthesis, where mRNA is used as a template for the assembly of amino acids to produce a polypeptide chain that undergoes modifications to form the protein. Translation is a complex and intricate process that involves many factors which are required to work in concert with one an- other. Briefly, eukaryotic initiation factors (eIFs) activate the mRNA in preparation for ribosomal binding. eIFs bind to the m7G cap and the poly-A tail at the 5' and 3' ends of the mRNA, respectively. Next, the small ribosomal subunit (40S), loaded with the methionyl tRNA specialized for initiation (Met-tRNAi) and the initiation factors 1, 1A, 2, 3, and 5 must come together to form a pre-initiation complex (PIC). PIC interacts with the m7G binding eIF4E as well as eIF4G. The PIC then scans the 5' untranslated region (UTR) of the mRNA for the start codon AUG (Sonenberg and Hinnebusch, 2009). Furthermore, poly(A) binding protein (PABP) binds the poly(A) tail at the 3' end of the mRNA and binds several factors, including the termination factor eRF-3. Direct interaction between PABP and eIF4G allows for the circularization of mRNA and the 1 Chapter 1. Introduction 2 formation of a closed loop complex (Sonenberg and Dever, 2003). The GTP (Guanosine Triphosphate) on eIF2 is hydrolyzed to GDP (Guanosine Diphosphate), enabling the 40S ribosomal subunit to bind with a larger (60S) ribosomal subunit, forming an 80S ribosome necessary for translation of the mRNA (de Moor et al., 2005). Other factors bind the 3' UTR and contribute to the translation of mRNA. One such factor is the cytoplasmic polyadenylation element binding protein (CPEB). CPEB recognizes the cytoplasmic polyadenylation element (CPE) in the 3' UTR of the mRNA and improves the recruitment of cleavage and polyadenylation specificity factor (CPSF), allowing for polyadenylation of the mRNA by poly(A) polymerase, thus increasing the stability of the mRNA (Radford et al., 2008). As one would expect, mRNA translation has the potential to be regulated in many ways and at different steps as there are many factors that are involved. For example, phosphorylation of CPEB by Aurora A kinase on serine 174 is required in order for CPEB to recruit CPSF to the poly(A) signal. Another example is the suppression of cyclin B1 translation by Maskin, which can bind CPEB and eIF4E.
Recommended publications
  • Program Nr: 1 from the 2004 ASHG Annual Meeting Mutations in A

    Program Nr: 1 from the 2004 ASHG Annual Meeting Mutations in A

    Program Nr: 1 from the 2004 ASHG Annual Meeting Mutations in a novel member of the chromodomain gene family cause CHARGE syndrome. L.E.L.M. Vissers1, C.M.A. van Ravenswaaij1, R. Admiraal2, J.A. Hurst3, B.B.A. de Vries1, I.M. Janssen1, W.A. van der Vliet1, E.H.L.P.G. Huys1, P.J. de Jong4, B.C.J. Hamel1, E.F.P.M. Schoenmakers1, H.G. Brunner1, A. Geurts van Kessel1, J.A. Veltman1. 1) Dept Human Genetics, UMC Nijmegen, Nijmegen, Netherlands; 2) Dept Otorhinolaryngology, UMC Nijmegen, Nijmegen, Netherlands; 3) Dept Clinical Genetics, The Churchill Hospital, Oxford, United Kingdom; 4) Children's Hospital Oakland Research Institute, BACPAC Resources, Oakland, CA. CHARGE association denotes the non-random occurrence of ocular coloboma, heart defects, choanal atresia, retarded growth and development, genital hypoplasia, ear anomalies and deafness (OMIM #214800). Almost all patients with CHARGE association are sporadic and its cause was unknown. We and others hypothesized that CHARGE association is due to a genomic microdeletion or to a mutation in a gene affecting early embryonic development. In this study array- based comparative genomic hybridization (array CGH) was used to screen patients with CHARGE association for submicroscopic DNA copy number alterations. De novo overlapping microdeletions in 8q12 were identified in two patients on a genome-wide 1 Mb resolution BAC array. A 2.3 Mb region of deletion overlap was defined using a tiling resolution chromosome 8 microarray. Sequence analysis of genes residing within this critical region revealed mutations in the CHD7 gene in 10 of the 17 CHARGE patients without microdeletions, including 7 heterozygous stop-codon mutations.
  • Evidence for Differential Alternative Splicing in Blood of Young Boys With

    Evidence for Differential Alternative Splicing in Blood of Young Boys With

    Stamova et al. Molecular Autism 2013, 4:30 http://www.molecularautism.com/content/4/1/30 RESEARCH Open Access Evidence for differential alternative splicing in blood of young boys with autism spectrum disorders Boryana S Stamova1,2,5*, Yingfang Tian1,2,4, Christine W Nordahl1,3, Mark D Shen1,3, Sally Rogers1,3, David G Amaral1,3 and Frank R Sharp1,2 Abstract Background: Since RNA expression differences have been reported in autism spectrum disorder (ASD) for blood and brain, and differential alternative splicing (DAS) has been reported in ASD brains, we determined if there was DAS in blood mRNA of ASD subjects compared to typically developing (TD) controls, as well as in ASD subgroups related to cerebral volume. Methods: RNA from blood was processed on whole genome exon arrays for 2-4–year-old ASD and TD boys. An ANCOVA with age and batch as covariates was used to predict DAS for ALL ASD (n=30), ASD with normal total cerebral volumes (NTCV), and ASD with large total cerebral volumes (LTCV) compared to TD controls (n=20). Results: A total of 53 genes were predicted to have DAS for ALL ASD versus TD, 169 genes for ASD_NTCV versus TD, 1 gene for ASD_LTCV versus TD, and 27 genes for ASD_LTCV versus ASD_NTCV. These differences were significant at P <0.05 after false discovery rate corrections for multiple comparisons (FDR <5% false positives). A number of the genes predicted to have DAS in ASD are known to regulate DAS (SFPQ, SRPK1, SRSF11, SRSF2IP, FUS, LSM14A). In addition, a number of genes with predicted DAS are involved in pathways implicated in previous ASD studies, such as ROS monocyte/macrophage, Natural Killer Cell, mTOR, and NGF signaling.
  • A Role for Dazl in Commitment to Gametogenic Fate in Embryonic Germ Cells of C57BL/6 Mice

    A Role for Dazl in Commitment to Gametogenic Fate in Embryonic Germ Cells of C57BL/6 Mice

    A Role for Dazl in Commitment to Gametogenic Fate in Embryonic Germ Cells of C57BL/6 Mice by Yanfeng Lin (Yen-Hong Lim) B.S. Biochemistry and Molecular and Cell Biology University of Wisconsin-Madison, 1998 SUBMITTED TO THE DEPARTMENT OF BIOLOGY IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF DOCTOR OF PHILOSOPHY IN BIOLOGY AT THE MASSACHUSETTS INSTITUTE OF TECHNOLOGY SEPT 2005 C 2005 Yanfeng Lin. All rights reserved. The author hereby grants to MIT permission to reproduce and distribute publicly paper and electronic copies of this thesis document in whole or in part. Signature of Author __ _ __ Department of Biology August, 2005 V /-' ~J-2 Certified by David C. Page Professor of Biology Howard Hughes Medical Institute Thesis Supervisor Accepted b) Stephen P. Bell Co-chair, Biology Graduate Student Committee 'MACHUS ETT S NS1 I OF TECHNOLOGY ARCHIVES' OCT 0 2005 . ,. -~ I LIBRARIES .i. __ A Role for Dazl in Commitment to Gametogenic Fate in Embryonic Germ Cells of C57BL/6 Mice by Yanfeng Lin (Yen-Hong Lim) Submitted to the Department of Biology on June, 2005 in Partial Fulfillment of the Requirements for the Degree of Doctor of Philosophy in Biology Abstract Germ cells can be defined as the cells that undergo the terminal differentiating process of meiosis. In mice, as XX germ cells enter meiosis around Embryonic days 13.5-14.5 (E13.5-E14.5), they form meiotic figures and down-regulate pluripotency markers. XY germ cells enter proliferation arrest between E13.5 and E16.5, which is accompanied by a distinct morphological change as well.
  • Genetics of Azoospermia

    Genetics of Azoospermia

    International Journal of Molecular Sciences Review Genetics of Azoospermia Francesca Cioppi , Viktoria Rosta and Csilla Krausz * Department of Biochemical, Experimental and Clinical Sciences “Mario Serio”, University of Florence, 50139 Florence, Italy; francesca.cioppi@unifi.it (F.C.); viktoria.rosta@unifi.it (V.R.) * Correspondence: csilla.krausz@unifi.it Abstract: Azoospermia affects 1% of men, and it can be due to: (i) hypothalamic-pituitary dysfunction, (ii) primary quantitative spermatogenic disturbances, (iii) urogenital duct obstruction. Known genetic factors contribute to all these categories, and genetic testing is part of the routine diagnostic workup of azoospermic men. The diagnostic yield of genetic tests in azoospermia is different in the different etiological categories, with the highest in Congenital Bilateral Absence of Vas Deferens (90%) and the lowest in Non-Obstructive Azoospermia (NOA) due to primary testicular failure (~30%). Whole- Exome Sequencing allowed the discovery of an increasing number of monogenic defects of NOA with a current list of 38 candidate genes. These genes are of potential clinical relevance for future gene panel-based screening. We classified these genes according to the associated-testicular histology underlying the NOA phenotype. The validation and the discovery of novel NOA genes will radically improve patient management. Interestingly, approximately 37% of candidate genes are shared in human male and female gonadal failure, implying that genetic counselling should be extended also to female family members of NOA patients. Keywords: azoospermia; infertility; genetics; exome; NGS; NOA; Klinefelter syndrome; Y chromosome microdeletions; CBAVD; congenital hypogonadotropic hypogonadism Citation: Cioppi, F.; Rosta, V.; Krausz, C. Genetics of Azoospermia. 1. Introduction Int. J. Mol. Sci.
  • Dynamic Changes in RNA–Protein Interactions and RNA Secondary Structure in Mammalian Erythropoiesis

    Dynamic Changes in RNA–Protein Interactions and RNA Secondary Structure in Mammalian Erythropoiesis

    Published Online: 27 July, 2021 | Supp Info: http://doi.org/10.26508/lsa.202000659 Downloaded from life-science-alliance.org on 30 September, 2021 Resource Dynamic changes in RNA–protein interactions and RNA secondary structure in mammalian erythropoiesis Mengge Shan1,2 , Xinjun Ji3, Kevin Janssen5 , Ian M Silverman3 , Jesse Humenik3, Ben A Garcia5, Stephen A Liebhaber3,4, Brian D Gregory1,2 Two features of eukaryotic RNA molecules that regulate their and RNA secondary structure. These techniques generally either post-transcriptional fates are RNA secondary structure and RNA- use chemical probing agents or structure-specific RNases (single- binding protein (RBP) interaction sites. However, a comprehen- stranded RNases (ssRNases) and double-stranded RNases [dsRNa- sive global overview of the dynamic nature of these sequence ses]) to provide site-specific evidence for a region being in single- or features during erythropoiesis has never been obtained. Here, we double-stranded configurations (4, 5). use our ribonuclease-mediated structure and RBP-binding site To date, the known repertoire of RBP–RNA interaction sites has mapping approach to reveal the global landscape of RNA sec- been built on a protein-by-protein basis, with studies identifying ondary structure and RBP–RNA interaction sites and the dynamics the targets of a single protein of interest, often through the use of of these features during this important developmental process. techniques such as crosslinking and immunoprecipitation se- We identify dynamic patterns of RNA secondary structure and RBP quencing (CLIP-seq) (6). In CLIP-seq, samples are irradiated with UV binding throughout the process and determine a set of corre- to induce the cross-linking of proteins to their RNA targets.
  • Genomic and Expression Profiling of Human Spermatocytic Seminomas: Primary Spermatocyte As Tumorigenic Precursor and DMRT1 As Candidate Chromosome 9 Gene

    Genomic and Expression Profiling of Human Spermatocytic Seminomas: Primary Spermatocyte As Tumorigenic Precursor and DMRT1 As Candidate Chromosome 9 Gene

    Research Article Genomic and Expression Profiling of Human Spermatocytic Seminomas: Primary Spermatocyte as Tumorigenic Precursor and DMRT1 as Candidate Chromosome 9 Gene Leendert H.J. Looijenga,1 Remko Hersmus,1 Ad J.M. Gillis,1 Rolph Pfundt,4 Hans J. Stoop,1 Ruud J.H.L.M. van Gurp,1 Joris Veltman,1 H. Berna Beverloo,2 Ellen van Drunen,2 Ad Geurts van Kessel,4 Renee Reijo Pera,5 Dominik T. Schneider,6 Brenda Summersgill,7 Janet Shipley,7 Alan McIntyre,7 Peter van der Spek,3 Eric Schoenmakers,4 and J. Wolter Oosterhuis1 1Department of Pathology, Josephine Nefkens Institute; Departments of 2Clinical Genetics and 3Bioinformatics, Erasmus Medical Center/ University Medical Center, Rotterdam, the Netherlands; 4Department of Human Genetics, Radboud University Medical Center, Nijmegen, the Netherlands; 5Howard Hughes Medical Institute, Whitehead Institute and Department of Biology, Massachusetts Institute of Technology, Cambridge, Massachusetts; 6Clinic of Paediatric Oncology, Haematology and Immunology, Heinrich-Heine University, Du¨sseldorf, Germany; 7Molecular Cytogenetics, Section of Molecular Carcinogenesis, The Institute of Cancer Research, Sutton, Surrey, United Kingdom Abstract histochemistry, DMRT1 (a male-specific transcriptional regulator) was identified as a likely candidate gene for Spermatocytic seminomas are solid tumors found solely in the involvement in the development of spermatocytic seminomas. testis of predominantly elderly individuals. We investigated these tumors using a genome-wide analysis for structural and (Cancer Res 2006; 66(1): 290-302) numerical chromosomal changes through conventional kar- yotyping, spectral karyotyping, and array comparative Introduction genomic hybridization using a 32 K genomic tiling-path Spermatocytic seminomas are benign testicular tumors that resolution BAC platform (confirmed by in situ hybridization).
  • Pumilio Proteins Regulate Translation in Embryonic Stem Cells and Are Essential for Early Embryonic Development Katherine Elizabeth Uyhazi Yale University

    Pumilio Proteins Regulate Translation in Embryonic Stem Cells and Are Essential for Early Embryonic Development Katherine Elizabeth Uyhazi Yale University

    Yale University EliScholar – A Digital Platform for Scholarly Publishing at Yale Yale Medicine Thesis Digital Library School of Medicine 12-2012 Pumilio Proteins Regulate Translation in Embryonic Stem Cells and are Essential for Early Embryonic Development Katherine Elizabeth Uyhazi Yale University. Follow this and additional works at: http://elischolar.library.yale.edu/ymtdl Part of the Medicine and Health Sciences Commons Recommended Citation Uyhazi, Katherine Elizabeth, "Pumilio Proteins Regulate Translation in Embryonic Stem Cells and are Essential for Early Embryonic Development" (2012). Yale Medicine Thesis Digital Library. 2189. http://elischolar.library.yale.edu/ymtdl/2189 This Open Access Dissertation is brought to you for free and open access by the School of Medicine at EliScholar – A Digital Platform for Scholarly Publishing at Yale. It has been accepted for inclusion in Yale Medicine Thesis Digital Library by an authorized administrator of EliScholar – A Digital Platform for Scholarly Publishing at Yale. For more information, please contact [email protected]. ABSTRACT Pumilio Proteins Regulate Translation in Embryonic Stem Cells and are Essential for Early Embryonic Development Katherine E. Uyhazi 2012 Embryonic stem (ES) cells are defined by their dual abilities to self-renew and to differentiate into any cell type in the body. This vast potential is precisely controlled by spatial and temporal gene regulation at transcriptional, post-transcriptional, and epigenetic levels. Recent studies have revealed several transcription factors that are essential for stem cell self-renewal and pluripotency, but the role of translational control in ES cells is poorly understood. Translational control is a fundamental mechanism of gene regulation during early development, and likely explains the discrepancies between the transcriptome and proteome profiles of stem cells and their differentiated progeny.
  • Intermolecular Interactions of Homologs of Germ Plasm Components in Mammalian Germ Cells Mark S

    Intermolecular Interactions of Homologs of Germ Plasm Components in Mammalian Germ Cells Mark S

    Dominican Scholar Collected Faculty and Staff Scholarship Faculty and Staff Scholarship 2007 Intermolecular Interactions of Homologs of Germ Plasm Components in Mammalian Germ Cells Mark S. Fox Center for Reproductive Sciences; Human Embryonic Stem Cell Research Center; Department of Obstetrics, Gynecology and Reproductive Sciences; Institute for Stem Cell and Tissue Biology; Departments of Physiology and Urology; University of California at San Francisco Amander T. Clark Center for Reproductive Sciences; Human Embryonic Stem Cell Research Center; Department of Obstetrics, Gynecology and Reproductive Sciences; Institute for Stem Cell and Tissue Biology; Departments of Physiology and Urology; University of California at San Francisco Mohammed El Majdoubi Center for Reproductive Sciences; Human Embryonic Stem Cell Research Center; Department of Obstetrics, Gynecology and Reproductive Sciences; Institute for Stem Cell and Tissue Biology; Departments of Physiology and Urology; University of California at San Francisco, [email protected] Recommended Citation Fox, Mark S.; Clark, Amander T.; El Majdoubi, Mohammed; Vigne, Jean-Louis; Urano, Jun; Hostetler, Chris E.; Griswold, Michael D.; Weiner, Richard I.; and Reijo Pera, Renee A., "Intermolecular Interactions of Homologs of Germ Plasm Components in Mammalian Germ Cells" (2007). Collected Faculty and Staff Scholarship. 308. https://scholar.dominican.edu/all-faculty/308 DOI http://dx.doi.org/https://doi.org/10.1016/j.ydbio.2006.08.047 This Article is brought to you for free and open access by the Faculty and Staff Scholarship at Dominican Scholar. It has been accepted for inclusion in Collected Faculty and Staff Scholarship by an authorized administrator of Dominican Scholar. For more information, please contact [email protected].
  • Protein-Protein Interaction Network Analysis and Identification of Key

    Protein-Protein Interaction Network Analysis and Identification of Key

    bioRxiv preprint doi: https://doi.org/10.1101/281030; this version posted March 14, 2018. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. Protein-Protein Interaction Network Analysis and Identification of Key Players in nor-NOHA and NOHA Mediated Pathways for Treatment of Cancer through Arginase Inhibition: Insights from Systems Biology Ishtiaque Ahammad1* 1. Department of Biochemistry and Microbiology, North South University, Dhaka, Bangladesh. * Corresponding author. Email: [email protected] Abstract L-arginine is involved in a number of biological processes in our bodies. Metabolism of L- arginine by the enzyme arginase has been found to be associated with cancer cell proliferation. Arginase inhibition has been proposed as a potential therapeutic means to inhibit this process. N- hydroxy-nor-L-Arg (nor-NOHA) and N (omega)-hydroxy-L-arginine (NOHA) has shown promise in inhibiting cancer progression through arginase inhibition. In this study, nor-NOHA and NOHA-associated genes and proteins were analyzed with several Bioinformatics and Systems Biology tools to identify the associated pathways and the key players involved so that a more comprehensive view of the molecular mechanisms including the regulatory mechanisms can be achieved and more potential targets for treatment of cancer can be discovered. Based on the analyses carried out, 3 significant modules have been identified from the PPI network. Five pathways/processes have been found to be significantly associated with nor-NOHA and NOHA associated genes. Out of the 1996 proteins in the PPI network, 4 have been identified as hub proteins- SOD, SOD1, AMD1, and NOS2.
  • (12) Patent Application Publication (10) Pub. No.: US 2011/0044954 A1 Stice Et Al

    (12) Patent Application Publication (10) Pub. No.: US 2011/0044954 A1 Stice Et Al

    US 20110044954A1 (19) United States (12) Patent Application Publication (10) Pub. No.: US 2011/0044954 A1 Stice et al. (43) Pub. Date: Feb. 24, 2011 (54) METHODS OF PRODUCING GERM-LIKE Publication Classification CELLS AND RELATED THERAPES (51) Int. Cl. A6II 35/12 (2006.01) CI2N 5/071 (2010.01) (76) Inventors: Steven Stice, Athens, GA (US); CI2N 5/07 (2010.01) Franklin West, Athens, GA (US) CI2N 5/00 (2006.01) A6IP 5/00 (2006.01) (52) U.S. Cl. ........ 424/93.7:435/377; 435/366; 435/354; Correspondence Address: 435/325 Henry D. Coleman (57) ABSTRACT 714 Colorado Avenue The present invention relates to methods of producing germ Bridgeport, CT 06605-1601 (US) like cells (GLCs) from embryonic stem cells and induced pluripotent stem cells, GLCs produced by such methods, gametes derived from Such GLCs, pharmaceutical composi (21) Appl. No.: 12/583,402 tions and kits containing Such GLCs, screens that use GLCs to identify agents useful in enhancing mammalian reproductive health, and methods of treatment that use GLCs to enhance (22) Filed: Aug. 20, 2009 mammalian reproductive health. Patent Application Publication Feb. 24, 2011 Sheet 1 of 15 US 2011/0044954 A1 Patent Application Publication Feb. 24, 2011 Sheet 2 of 15 US 2011/0044954 A1 A 1000 to a post. : : Patent Application Publication Feb. 24, 2011 Sheet 3 of 15 US 2011/0044954 A1 xxx x missism &r-o-o-o-o-o-o-o-o-o: aaraaaaaaaaaaaaaaaaaaaaaaaaaaaaaa to reexier : & Patent Application Publication Feb. 24, 2011 Sheet 4 of 15 US 2011/0044954 A1 xix.
  • Genome Provides Insights Into Vertebrate Evolution

    Genome Provides Insights Into Vertebrate Evolution

    ARTICLES OPEN Sequencing of the sea lamprey (Petromyzon marinus) genome provides insights into vertebrate evolution Jeramiah J Smith1,2, Shigehiro Kuraku3,4, Carson Holt5,37, Tatjana Sauka-Spengler6,37, Ning Jiang7, Michael S Campbell5, Mark D Yandell5, Tereza Manousaki4, Axel Meyer4, Ona E Bloom8,9, Jennifer R Morgan10, Joseph D Buxbaum11–14, Ravi Sachidanandam11, Carrie Sims15, Alexander S Garruss15, Malcolm Cook15, Robb Krumlauf15,16, Leanne M Wiedemann15,17, Stacia A Sower18, Wayne A Decatur18, Jeffrey A Hall18, Chris T Amemiya2,19, Nil R Saha2, Katherine M Buckley20,21, Jonathan P Rast20,21, Sabyasachi Das22,23, Masayuki Hirano22,23, Nathanael McCurley22,23, Peng Guo22,23, Nicolas Rohner24, Clifford J Tabin24, Paul Piccinelli25, Greg Elgar25, Magali Ruffier26, Bronwen L Aken26, Stephen M J Searle26, Matthieu Muffato27, Miguel Pignatelli27, Javier Herrero27, Matthew Jones6, C Titus Brown28,29, Yu-Wen Chung-Davidson30, Kaben G Nanlohy30, Scot V Libants30, Chu-Yin Yeh30, David W McCauley31, James A Langeland32, Zeev Pancer33, Bernd Fritzsch34, Pieter J de Jong35, Baoli Zhu35,37, Lucinda L Fulton36, Brenda Theising36, Paul Flicek27, Marianne E Bronner6, All rights reserved. Wesley C Warren36, Sandra W Clifton36,37, Richard K Wilson36 & Weiming Li30 Lampreys are representatives of an ancient vertebrate lineage that diverged from our own ~500 million years ago. By virtue of this deeply shared ancestry, the sea lamprey (P. marinus) genome is uniquely poised to provide insight into the ancestry of vertebrate genomes and the underlying principles of vertebrate biology. Here, we present the first lamprey whole-genome sequence and America, Inc. assembly. We note challenges faced owing to its high content of repetitive elements and GC bases, as well as the absence of broad-scale sequence information from closely related species.
  • Multiscale Genomic Analysis of The

    Multiscale Genomic Analysis of The

    University of Tennessee Health Science Center UTHSC Digital Commons Theses and Dissertations (ETD) College of Graduate Health Sciences 5-2009 Multiscale Genomic Analysis of the Corticolimbic System: Uncovering the Molecular and Anatomic Substrates of Anxiety-Related Behavior Khyobeni Mozhui University of Tennessee Health Science Center Follow this and additional works at: https://dc.uthsc.edu/dissertations Part of the Mental and Social Health Commons, Nervous System Commons, and the Neurosciences Commons Recommended Citation Mozhui, Khyobeni , "Multiscale Genomic Analysis of the Corticolimbic System: Uncovering the Molecular and Anatomic Substrates of Anxiety-Related Behavior" (2009). Theses and Dissertations (ETD). Paper 180. http://dx.doi.org/10.21007/etd.cghs.2009.0219. This Dissertation is brought to you for free and open access by the College of Graduate Health Sciences at UTHSC Digital Commons. It has been accepted for inclusion in Theses and Dissertations (ETD) by an authorized administrator of UTHSC Digital Commons. For more information, please contact [email protected]. Multiscale Genomic Analysis of the Corticolimbic System: Uncovering the Molecular and Anatomic Substrates of Anxiety-Related Behavior Document Type Dissertation Degree Name Doctor of Philosophy (PhD) Program Anatomy and Neurobiology Research Advisor Robert W. Williams, Ph.D. Committee John D. Boughter, Ph.D. Eldon E. Geisert, Ph.D. Kristin M. Hamre, Ph.D. Jeffery D. Steketee, Ph.D. DOI 10.21007/etd.cghs.2009.0219 This dissertation is available at UTHSC Digital