DOCSLIB.ORG

The Role of Zyxin and LIMD1 in Mitosis and Cancer

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text

Load more

University of Nebraska Medical Center DigitalCommons@UNMC Theses & Dissertations Graduate Studies Spring 5-4-2019 The Role of Zyxin and LIMD1 in Mitosis and Cancer Jiuli Zhou University of Nebraska Medical Center Follow this and additional works at: https://digitalcommons.unmc.edu/etd Part of the Cancer Biology Commons, and the Cell Biology Commons Recommended Citation Zhou, Jiuli, "The Role of Zyxin and LIMD1 in Mitosis and Cancer" (2019). Theses & Dissertations. 353. https://digitalcommons.unmc.edu/etd/353 This Dissertation is brought to you for free and open access by the Graduate Studies at DigitalCommons@UNMC. It has been accepted for inclusion in Theses & Dissertations by an authorized administrator of DigitalCommons@UNMC. For more information, please contact [email protected]. THE ROLE OF ZYXIN AND LIMD1 IN MITOSIS AND CANCER by Jiuli Zhou A DISSERTATION Presented to the Faculty of the University of Nebraska Graduate College in Partial Fulfillment of the Requirements for the Degree of Doctor of Philosophy Pathology & Microbiology Graduate Program Under the Supervision of Professor Jixin Dong University of Nebraska Medical Center Omaha, Nebraska May, 2019 Supervisory Committee: Jennifer D. Black, Ph.D. Robert E. Lewis, Ph.D. Kaihong Su, Ph.D. i ACKNOWLEDGEMENTS Funding support: The China Scholarship Council; University of Nebraska Medical Center (UNMC) Graduate Studies Office Fellowship; National Cancer Institute/National Institutes of Health (NCI/NIH); Department of Defense; the COBRE Grant from the Nebraska Center for Cell Signaling/National Institute of General Medical Sciences (NIGMS/NIH). More important than the funding support, I would like to acknowledge the support and encouragement I received during my doctoral research. First and foremost, I would like to express my deepest appreciation to my advisor Dr. Jixin Dong. The completion of my dissertation would not have been possible without the support and nurturing of him. He taught me that it takes a long journey to dig deep into this field, which is not only a challenge of intelligence, but also a challenge of perseverance. Under his guidance, I am now equipped with a solid foundation for both molecular biology techniques and cancer research insights. Besides, I have strengthened my ability to design experiments and write papers and proposals independently. By pursuing various projects, I know how to deal with obstacles properly and turn disappointments into lessons, discoveries, alternatives and new plans set in motion. Thanks to his encouragement and patience for years, I have prepared myself to be a skillful and thoughtful researcher and will achieve my dream step by step. Additionally, Dr. Dong has led our lab to be a happy family, where I feel love and warmness. I would also like to extend my deepest gratitude to my supervisor committee, Dr. Jennifer Black, Dr. Kaihong Su, and Dr. Robert Lewis. They have set up themselves as excellent models for my research career. I really appreciate the time they have taken out of their busy schedules to help me out. Their insightful suggestion and critical comments ii have given me valuable clues for my projects and greatly improved my scientific knowledge. Besides, I am also grateful that they provided me with encouragement throughout the duration of my project. I am deeply indebted to my previous and present lab members: Shuping Yang, Lin Zhang, Xingcheng Chen, Seth Stauffer, Yu Ou, Zhan Wang, Yuanhong Chen, Yongji Zeng, Renya Zeng, and Ling Yin. I want to thank them for providing technique solutions as well as scientific suggestions on my research projects, and generous support and help when I was struggling. Special thanks to Yuanhong not only for helping us in research needs and laboratory management, but also for her countless supports and cares in my daily life like my family. Many thanks to my program director, Dr. Rakesh Singh and my program coordinator, Tuire Cechin for their help with my study and supports when I faced problems. I wish to thank UNMC core facilities, including Comparative Medicine, Advanced Microscope Core Facility, Tissue Sciences Facilities, Flow Cytometry Research Facility and DNA Sequencing Core for their great amount of assistance with my projects. My heartfelt appreciation also goes to my families and friends near and far. I especially thank my parents who nurtured me with endless love, and encouraged me to be a brave, honest, confident, and hard-working person. I also wish to express my thanks to my best friend, soul-mate, and husband Yijiu Ren. I married the best person, who never let me down and always stood by my side. Four years ago, I built a dream. Now, the dream builds me. I am so grateful for those who support me along the way of my Ph.D. training. They are a reason to my greatness in life and I will never walk alone. iii ABSTRACT: THE ROLE OF ZYXIN AND LIMD1 IN MITOSIS AND CANCER Jiuli Zhou, Ph.D. University of Nebraska, 2019 Supervisor: Jixin Dong, Ph.D. The Hippo signaling pathway, originally discovered in Drosophila, consists of a core kinase cascade and has been subsequently demonstrated to control tissue growth and tumorigenesis. The core of this pathway contains MST1/2 (Mammalian sterile 20-like kinase 1/2), LATS1/2 (large tumor suppressor 1/2) and downstream effector named Yes- associated protein (YAP) and PDZ-binding motif (TAZ). MST1/2 transduce their kinase activity mainly through directly phosphorylating LATS1/2. Once phosphorylated and activated, LATS1/2 subsequently phosphorylate and inhibit YAP/TAZ from translocating to nucleus, thereby suppressing the expression of downstream pro-growth and survival genes. While recent studies provide important insight into the tumor suppressor properties of this pathway, the underlying molecular mechanisms through which the Hippo components exert their oncogenic/suppressing function are poorly understood. Zyxin and LIM domains containing 1 (LIMD1) are adaptor/scaffold proteins with three LIM domains, which are involved in protein-protein interactions during cellular signaling transduction. Several recent studies showed that Zyxin/LIMD1 affect Hippo-YAP signaling. Our studies found that Zyxin and LIMD1 are phosphorylated in mitosis via novel sites. We further characterized the phospho-regulation of Zyxin and LIMD1 in mitosis and examined the functional significance of the phosphorylation in colon cancer and lung cancer, respectively. Moreover, we demonstrated that Zyxin regulates YAP activity through colon cancer oncogene Cyclin-Dependent Kinase 8 (CDK8). Mechanistically, we identified that iv CDK8 directly phosphorylates YAP and promotes its activation. Together, our findings revealed novel layers of regulation for Zyxin/LIMD1 in mitosis and their roles in tumorigenesis. v TABLE OF CONTENT ACKNOWLEDGEMENTS ................................................................................................. i ABSTRACT: THE ROLE OF ZYXIN AND LIMD1 IN MITOSIS AND CANCER ............... iii LIST OF FIGURES ....................................................................................................... viii LIST OF ABBREVIATION ............................................................................................... x Chapter 1: Zyxin promotes colon cancer tumorigenesis in a mitotic phosphorylation- dependent manner and through CDK8-mediated YAP activation* ………………………..1 Abstract ................................................................................................................. 2 1.1. Introduction .................................................................................................. 3 1.2. Materials and Methods ................................................................................ 5 1.2.1. Expression constructs .................................................................................. 5 1.2.2. Cell culture and transfection ........................................................................ 5 1.2.3. Cell cycle synchronization ............................................................................ 6 1.2.4. Establishment of cell lines ............................................................................ 6 1.2.5. CRISPR-double nikase-mediated knockout ................................................. 6 1.2.6. Kinase inhibitors .......................................................................................... 7 1.2.7. Recombinant protein purification and in vitro kinase assay .......................... 7 1.2.8. Antibodies .................................................................................................... 8 1.2.9. Phos-tag and western blot assay ................................................................. 8 1.2.10. Immunofluorescence staining and cofocal mircoscopy ............................... 8 1.2.11. Cell proliferation, migration (wound healing) and colony formation assays . 9 1.2.12. Quantitative real time PCR ......................................................................... 9 1.2.13. Animal studies ............................................................................................ 9 1.2.14. Immunohistochemistry (IHC) staining and scoring ....................................10 vi 1.2.15. Statistical analysis .....................................................................................11 1.3. Results .......................................................................................................12 1.3.1. Zyxin is phosphorylated by CDK1 in vitro during mitosis .............................12
Recommended publications
  • Hidden Targets in RAF Signalling Pathways to Block Oncogenic RAS Signalling

    Hidden Targets in RAF Signalling Pathways to Block Oncogenic RAS Signalling

    G C A T T A C G G C A T genes Review Hidden Targets in RAF Signalling Pathways to Block Oncogenic RAS Signalling Aoife A. Nolan 1, Nourhan K. Aboud 1, Walter Kolch 1,2,* and David Matallanas 1,* 1 Systems Biology Ireland, School of Medicine, University College Dublin, Belfield, Dublin 4, Ireland; [email protected] (A.A.N.); [email protected] (N.K.A.) 2 Conway Institute of Biomolecular & Biomedical Research, University College Dublin, Belfield, Dublin 4, Ireland * Correspondence: [email protected] (W.K.); [email protected] (D.M.) Abstract: Oncogenic RAS (Rat sarcoma) mutations drive more than half of human cancers, and RAS inhibition is the holy grail of oncology. Thirty years of relentless efforts and harsh disappointments have taught us about the intricacies of oncogenic RAS signalling that allow us to now get a pharma- cological grip on this elusive protein. The inhibition of effector pathways, such as the RAF-MEK-ERK pathway, has largely proven disappointing. Thus far, most of these efforts were aimed at blocking the activation of ERK. Here, we discuss RAF-dependent pathways that are regulated through RAF functions independent of catalytic activity and their potential role as targets to block oncogenic RAS signalling. We focus on the now well documented roles of RAF kinase-independent functions in apoptosis, cell cycle progression and cell migration. Keywords: RAF kinase-independent; RAS; MST2; ASK; PLK; RHO-α; apoptosis; cell cycle; cancer therapy Citation: Nolan, A.A.; Aboud, N.K.; Kolch, W.; Matallanas, D. Hidden Targets in RAF Signalling Pathways to Block Oncogenic RAS Signalling.
  • Bioinformatic Analysis of Structure and Function of LIM Domains of Human Zyxin Family Proteins

    Bioinformatic Analysis of Structure and Function of LIM Domains of Human Zyxin Family Proteins

    International Journal of Molecular Sciences Article Bioinformatic Analysis of Structure and Function of LIM Domains of Human Zyxin Family Proteins M. Quadir Siddiqui 1,† , Maulik D. Badmalia 1,† and Trushar R. Patel 1,2,3,* 1 Alberta RNA Research and Training Institute, Department of Chemistry and Biochemistry, University of Lethbridge, 4401 University Drive, Lethbridge, AB T1K 3M4, Canada; [email protected] (M.Q.S.); [email protected] (M.D.B.) 2 Department of Microbiology, Immunology and Infectious Disease, Cumming School of Medicine, University of Calgary, 3330 Hospital Drive, Calgary, AB T2N 4N1, Canada 3 Li Ka Shing Institute of Virology, University of Alberta, Edmonton, AB T6G 2E1, Canada * Correspondence: [email protected] † These authors contributed equally to the work. Abstract: Members of the human Zyxin family are LIM domain-containing proteins that perform critical cellular functions and are indispensable for cellular integrity. Despite their importance, not much is known about their structure, functions, interactions and dynamics. To provide insights into these, we used a set of in-silico tools and databases and analyzed their amino acid sequence, phylogeny, post-translational modifications, structure-dynamics, molecular interactions, and func- tions. Our analysis revealed that zyxin members are ohnologs. Presence of a conserved nuclear export signal composed of LxxLxL/LxxxLxL consensus sequence, as well as a possible nuclear localization signal, suggesting that Zyxin family members may have nuclear and cytoplasmic roles. The molecular modeling and structural analysis indicated that Zyxin family LIM domains share Citation: Siddiqui, M.Q.; Badmalia, similarities with transcriptional regulators and have positively charged electrostatic patches, which M.D.; Patel, T.R.
  • Establishment and Genomic Characterization of a Sporadic Malignant Peripheral Nerve Sheath Tumor Cell Line Jody Fromm Longo1, Stephanie N

    Establishment and Genomic Characterization of a Sporadic Malignant Peripheral Nerve Sheath Tumor Cell Line Jody Fromm Longo1, Stephanie N

    www.nature.com/scientificreports OPEN Establishment and genomic characterization of a sporadic malignant peripheral nerve sheath tumor cell line Jody Fromm Longo1, Stephanie N. Brosius3,5,7, Iya Znoyko1, Victoria A. Alers1, Dorea P. Jenkins1, Robert C. Wilson1,2, Andrew J. Carroll4, Daynna J. Wolf1, Kevin A. Roth6 & Steven L. Carroll1,2,3* Malignant peripheral nerve sheath tumors (MPNSTs) are aggressive Schwann cell-derived neoplasms that occur sporadically or in patients with neurofbromatosis type 1 (NF1). Preclinical research on sporadic MPNSTs has been limited as few cell lines exist. We generated and characterized a new sporadic MPNST cell line, 2XSB, which shares the molecular and genomic features of the parent tumor. These cells have a highly complex karyotype with extensive chromothripsis. 2XSB cells show robust invasive 3-dimensional and clonogenic culture capability and form solid tumors when xenografted into immunodefcient mice. High-density single nucleotide polymorphism array and whole exome sequencing analyses indicate that, unlike NF1-associated MPNSTs, 2XSB cells have intact, functional NF1 alleles with no evidence of mutations in genes encoding components of Polycomb Repressor Complex 2. However, mutations in other genes implicated in MPNST pathogenesis were identifed in 2XSB cells including homozygous deletion of CDKN2A and mutations in TP53 and PTEN. We also identifed mutations in genes not previously associated with MPNSTs but associated with the pathogenesis of other human cancers. These include DNMT1, NUMA1, NTRK1, PDE11A, CSMD3, LRP5 and ACTL9. This sporadic MPNST-derived cell line provides a useful tool for investigating the biology and potential treatment regimens for sporadic MPNSTs. Malignant peripheral nerve sheath tumors (MPNSTs) are aggressive neoplasms derived from the Schwann cell lineage1,2.
  • Inhibition of Polo-Like Kinase 1 During the DNA Damage Response Is Mediated Through Loss of Aurora a Recruitment by Bora

    Inhibition of Polo-Like Kinase 1 During the DNA Damage Response Is Mediated Through Loss of Aurora a Recruitment by Bora

    OPEN Oncogene (2017) 36, 1840–1848 www.nature.com/onc ORIGINAL ARTICLE Inhibition of Polo-like kinase 1 during the DNA damage response is mediated through loss of Aurora A recruitment by Bora W Bruinsma1,2,4,5, M Aprelia1,2,5, I García-Santisteban1,3, J Kool2,YJXu2 and RH Medema1,2 When cells in G2 phase are challenged with DNA damage, several key mitotic regulators such as Cdk1/Cyclin B, Aurora A and Plk1 are inhibited to prevent entry into mitosis. Here we have studied how inhibition of Plk1 is established after DNA damage. Using a Förster resonance energy transfer (FRET)-based biosensor for Plk1 activity, we show that inhibition of Plk1 after DNA damage occurs with relatively slow kinetics and is entirely dependent on loss of Plk1-T210 phosphorylation. As T210 is phosphorylated by the kinase Aurora A in conjunction with its co-factor Bora, we investigated how they are affected by DNA damage. Interestingly, we find that the interaction between Bora and Plk1 remains intact during the early phases of the DNA damage response (DDR), whereas Plk1 activity is already inhibited at this stage. Expression of an Aurora A mutant that is refractory to inhibition by the DDR failed to prevent inhibition of Plk1 and loss of T210 phosphorylation, suggesting that inhibition of Plk1 may be established by perturbing recruitment of Aurora A by Bora. Indeed, expression of a fusion in which Aurora A was directly coupled to Bora prevented DNA damage-induced inhibition of Plk1 activity, as well as inhibition of T210 phosphorylation. Taken together, these data demonstrate that DNA damage affects the function of Aurora A at multiple levels: both by direct inhibition of Aurora A activity, as well as by perturbing the interaction with its co-activator Bora.
  • Photoreceptor Proliferation and Dysregulation of Cell Cycle Genes in Early Onset Inherited Retinal Degenerations

    Photoreceptor Proliferation and Dysregulation of Cell Cycle Genes in Early Onset Inherited Retinal Degenerations

    University of Pennsylvania ScholarlyCommons Departmental Papers (Vet) School of Veterinary Medicine 3-11-2016 Photoreceptor Proliferation and Dysregulation of Cell Cycle Genes in Early Onset Inherited Retinal Degenerations Kristin L. Gardiner University of Pennsylvania Louise Downs University of Pennsylvania Agnes I. Berta-Antalics University of Pennsylvania Evelyn Santana University of Pennsylvania Gustavo D. Aguirre University of Pennsylvania, [email protected] See next page for additional authors Follow this and additional works at: https://repository.upenn.edu/vet_papers Part of the Veterinary Medicine Commons Recommended Citation Gardiner, K. L., Downs, L., Berta-Antalics, A. I., Santana, E., Aguirre, G. D., & Genini, S. (2016). Photoreceptor Proliferation and Dysregulation of Cell Cycle Genes in Early Onset Inherited Retinal Degenerations. BMC Genomics, 17 (221), http://dx.doi.org/10.1186/s12864-016-2477-9 This paper is posted at ScholarlyCommons. https://repository.upenn.edu/vet_papers/152 For more information, please contact [email protected]. Photoreceptor Proliferation and Dysregulation of Cell Cycle Genes in Early Onset Inherited Retinal Degenerations Abstract Background Mitotic terminally differentiated photoreceptors (PRs) are observed in early retinal degeneration (erd), an inherited canine retinal disease driven by mutations in the NDR kinase STK38L (NDR2). Results We demonstrate that a similar proliferative response, but of lower magnitude, occurs in two other early onset disease models, X-linked progressive retinal atrophy 2 (xlpra2) and rod cone dysplasia 1 (rcd1). Proliferating cells are rod PRs, and not microglia or Müller cells. Expression of the cell cycle related genes RB1 and E2F1 as well as CDK2,4,6 was up-regulated, but changes were mutation-specific.
  • Comprehensive Identification of Proteins in Hodgkin Lymphoma

    Comprehensive Identification of Proteins in Hodgkin Lymphoma

    Laboratory Investigation (2007) 87, 1113–1124 & 2007 USCAP, Inc All rights reserved 0023-6837/07 $30.00 Comprehensive identification of proteins in Hodgkin lymphoma-derived Reed–Sternberg cells by LC-MS/MS Jeremy C Wallentine1, Ki Kwon Kim1, Charles E Seiler III1, Cecily P Vaughn2, David K Crockett2, Sheryl R Tripp2, Kojo SJ Elenitoba-Johnson1,2 and Megan S Lim1,2 Mass spectrometry-based proteomics in conjunction with liquid chromatography and bioinformatics analysis provides a highly sensitive and high-throughput approach for the identification of proteins. Hodgkin lymphoma is a form of malignant lymphoma characterized by the proliferation of Reed–Sternberg cells and background reactive lymphocytes. Comprehensive analysis of proteins expressed and released by Reed–Sternberg cells would assist in the discovery of potential biomarkers and improve our understanding of its pathogenesis. The subcellular proteome of the three cellular compartments from L428 and KMH2 Hodgkin lymphoma-derived cell lines were fractionated, and analyzed by reverse- phase liquid chromatography coupled with electrospray ionization tandem mass spectrometry. Additionally, proteins released by Hodgkin lymphoma-derived L428 cells were extracted from serum-free culture media and analyzed. Peptide spectra were analyzed using TurboSEQUESTs against the UniProt protein database (5.26.05; 188 712 entries). A subset of the identified proteins was validated by Western blot analysis, immunofluorescence microscopy and im- munohistochemistry. A total of 1945 proteins were identified with 785 from the cytosolic fraction, 305 from the membrane fraction, 441 from the nuclear fraction and 414 released proteins using a minimum of two peptide identi- fications per protein and an error rate of o5.0%.
  • Molecular Signature Induced by RNASET2, a Tumor Antagonizing Gene, in Ovarian Cancer Cells

    Molecular Signature Induced by RNASET2, a Tumor Antagonizing Gene, in Ovarian Cancer Cells

    www.impactjournals.com/oncotarget/ Oncotarget, June, Vol.2, No 6 Molecular signature induced by RNASET2, a tumor antagonizing gene, in ovarian cancer cells Francesco Acquati1, Laura Monti1, Marta Lualdi1, Marco Fabbri2, Maria Grazia Sacco2, Laura Gribaldo2, and Roberto Taramelli1 1 Dipartimento di Biotecnologie e Scienze Molecolari, Università degli Studi dell’Insubria, via JH Dunant 3, 21100 Varese, Italy 2 European Commission - Joint Research Centre Institute for Health and Consumer Protection Molecular Biology and Genomics unit TP 464, Via E. Fermi, 2749 21027 Ispra (VA) - Italy Correspondence to: Roberto Taramelli, email: [email protected] Correspondence to: Francesco Acquati, email: [email protected] Keywords: RNases, cancer microenvironment, transcriptional profile Received: May 10, 2011, Accepted: June 2, 2011, Published: June 4, 2011 Copyright: © Acquati et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. ABSTRACT: Using the Hey3Met2 human ovarian cancer cell line, we previously found the RNASET2 gene to possess a remarkable in vivo tumor suppressor activity, although no in vitro features such as inhibition of cell proliferation, clonogenic potential, impaired growth in soft agar and increase in apoptotic rate could be detected. This is reminiscent of the behavior of genes belonging to the class of tumor antagonizing genes (TAG) which act mainly within the context of the microenvironment. Here we present transcriptional profiles analysis which indicates that investigations of the mechanisms of TAG biological functions require a comparison between the in vitro and in vivo expression patterns.
  • PDF Hosted at the Radboud Repository of the Radboud University Nijmegen

    PDF Hosted at the Radboud Repository of the Radboud University Nijmegen

    PDF hosted at the Radboud Repository of the Radboud University Nijmegen The following full text is a publisher's version. For additional information about this publication click this link. http://hdl.handle.net/2066/85871 Please be advised that this information was generated on 2021-09-29 and may be subject to change. ISOFORMS IN MUSCLE AND BRAIN CELLS localization and function • ralph j.a. oude ophuis • 2011 9 789088 912344 > ISBN 978-90-8891234,-4 DMPK ISOFORMS IN MUSCLE AND BRAIN CELLS LOCALIZATION AND FUNCTION Voor het bijwonen van de openbare verdediging van het proefschrift van RALPH J.A. OUDE OPHUIS DMPK ISOFORMS IN MUSCLE AND BRAIN CELLS LOCALIZATION AND FUNCTION op vrijdag 1 april 2011 om 13:00u precies in de Aula van de Radboud Universiteit Nijmegen aan de Comeniuslaan 2 te Nijmegen Na afloop van de verdediging is er een receptie ter plaatse PARANIMFEN Susan Mulders [email protected] Rinske van de Vorstenbosch r.vandevorstenbosch(§) ncmls.ru.nl DMPK ISOFORMS IN MUSCLE AND BRAIN CELLS LOCALIZATION AND FUNCTION ISBN-13 978-90-8891234-4 ISBN-10 90-8891-234-3 Printed by Proefsohriftmaken.nl || Printyourthesis.com Published by Uitgeverij BOXPress, Oisterwijk DMPK ISOFORMS IN MUSCLE AND BRAIN CELLS LOCALIZATION AND FUNCTION Een wetenschappelijke proeve op het gebied van de Medische Wetenschappen Proefschrift ter verkrijging van de graad van doctor aan de Radboud Universiteit Nijmegen op gezag van de rector magnificus prof. mr. S.C.J.J. Kortmann, volgens besluit van het college van decanen in het openbaar te verdedigen op vrijdag 1 april 2011 om 13:00 uur precies door Raphaël Johannes Antonius Oude Ophuis geboren op 24 oktober 1978 te Sint-Oedenrode Promotor Prof.
  • The Chromosome 3P21.3-Encoded Gene, LIMD1, Is a Critical Tumor Suppressor Involved in Human Lung Cancer Development

    The Chromosome 3P21.3-Encoded Gene, LIMD1, Is a Critical Tumor Suppressor Involved in Human Lung Cancer Development

    The chromosome 3p21.3-encoded gene, LIMD1, is a critical tumor suppressor involved in human lung cancer development Tyson V. Sharpa, Ahmad Al-Attarb,1, Daniel E. Foxlera,1, Li Dingc, Thomas Q. de A. Vallima, Yining Zhanga, Hala S. Nijmeha, Thomas M. Webba, Andrew G. Nicholsond, Qunyuan Zhange, Aldi Krajae, Ian Spendloveb, John Osbornec, Elaine Mardisc,e, and Gregory D. Longmoref,2 aSchool of Biomedical Sciences, Queen’s Medical Centre, University of Nottingham Medical School, Nottingham NG7 2UH, United Kingdom; bAcademic and Clinical Departments of Oncology, University of Nottingham, Nottingham NG5 1PB, United Kingdom; dDepartment of Histopathology, Royal Brompton Hospital, London SW3 6NP, United Kingdom; cDepartment of Genetics, Genome Sequencing Center, and eDivision of Statistical Genomics, Washington University School of Medicine, St. Louis, MO 63108; and fDepartments of Medicine and Cell Biology, Washington University, St Louis, MO 63110 Edited by George Klein, Karolinska Institutet, Stockholm, Sweden, and approved October 7, 2008 (received for review May 23, 2008) Loss of heterozygosity (LOH) and homozygous deletions at chro- toma protein (pRB), inhibits E2F-mediated transcription, and mosome 3p21.3 are common in both small and nonsmall cell lung suppresses expression of the majority of genes with E2F1- cancers, indicating the likely presence of tumor suppressor genes responsive elements (9). Moreover, forced expression of LIMD1 (TSGs). Although genetic and epigenetic changes within this region in A549 lung tumor cell line blocks tumor growth in vitro and in have been identified, the functional significance of these changes vivo (9). Here, we show that expression of the 3p21.3 gene, has not been explored.
  • Androgen Receptor

    Androgen Receptor

    RALTITREXED Dihydrofolate reductase BORTEZOMIB IsocitrateCannabinoid dehydrogenase CB1EPIRUBICIN receptor HYDROCHLORIDE [NADP] cytoplasmic VINCRISTINE SULFATE Hypoxia-inducible factor 1 alpha DOXORUBICINAtaxin-2 HYDROCHLORIDENIFENAZONEFOLIC ACID PYRIMETHAMINECellular tumor antigen p53 Muscleblind-likeThyroidVINBURNINEVINBLASTINETRIFLURIDINE protein stimulating 1 DEQUALINIUM SULFATEhormone receptor CHLORIDE Menin/Histone-lysine N-methyltransferasePHENELZINE MLLLANATOSIDE SULFATE C MELATONINDAUNORUBICINBETAMETHASONEGlucagon-like HYDROCHLORIDEEndonuclease peptide 4 1 receptor NICLOSAMIDEDIGITOXINIRINOTECAN HYDROCHLORIDE HYDRATE BISACODYL METHOTREXATEPaired boxAZITHROMYCIN protein Pax-8 ATPase family AAA domain-containing proteinLIPOIC 5 ACID, ALPHA Nuclear receptorCLADRIBINEDIGOXIN ROR-gammaTRIAMTERENE CARMUSTINEEndoplasmic reticulum-associatedFLUOROURACIL amyloid beta-peptide-binding protein OXYPHENBUTAZONEORLISTAT IDARUBICIN HYDROCHLORIDE 6-phospho-1-fructokinaseHeat shockSIMVASTATIN protein beta-1 TOPOTECAN HYDROCHLORIDE AZACITIDINEBloom syndromeNITAZOXANIDE protein Huntingtin Human immunodeficiency virus typeTIPRANAVIR 1 protease VitaminCOLCHICINE D receptorVITAMIN E FLOXURIDINE TAR DNA-binding protein 43 BROMOCRIPTINE MESYLATEPACLITAXEL CARFILZOMIBAnthrax lethalFlap factorendonucleasePrelamin-A/C 1 CYTARABINE Vasopressin V2 receptor AMITRIPTYLINEMicrotubule-associated HYDROCHLORIDERetinoidTRIMETHOPRIM proteinMothers X receptor tau against alpha decapentaplegic homolog 3 Histone-lysine N-methyltransferase-PODOFILOX H3 lysine-9OXYQUINOLINE
  • Activation of Diverse Signalling Pathways by Oncogenic PIK3CA Mutations

    Activation of Diverse Signalling Pathways by Oncogenic PIK3CA Mutations

    ARTICLE Received 14 Feb 2014 | Accepted 12 Aug 2014 | Published 23 Sep 2014 DOI: 10.1038/ncomms5961 Activation of diverse signalling pathways by oncogenic PIK3CA mutations Xinyan Wu1, Santosh Renuse2,3, Nandini A. Sahasrabuddhe2,4, Muhammad Saddiq Zahari1, Raghothama Chaerkady1, Min-Sik Kim1, Raja S. Nirujogi2, Morassa Mohseni1, Praveen Kumar2,4, Rajesh Raju2, Jun Zhong1, Jian Yang5, Johnathan Neiswinger6, Jun-Seop Jeong6, Robert Newman6, Maureen A. Powers7, Babu Lal Somani2, Edward Gabrielson8, Saraswati Sukumar9, Vered Stearns9, Jiang Qian10, Heng Zhu6, Bert Vogelstein5, Ben Ho Park9 & Akhilesh Pandey1,8,9 The PIK3CA gene is frequently mutated in human cancers. Here we carry out a SILAC-based quantitative phosphoproteomic analysis using isogenic knockin cell lines containing ‘driver’ oncogenic mutations of PIK3CA to dissect the signalling mechanisms responsible for oncogenic phenotypes induced by mutant PIK3CA. From 8,075 unique phosphopeptides identified, we observe that aberrant activation of PI3K pathway leads to increased phosphorylation of a surprisingly wide variety of kinases and downstream signalling networks. Here, by integrating phosphoproteomic data with human protein microarray-based AKT1 kinase assays, we discover and validate six novel AKT1 substrates, including cortactin. Through mutagenesis studies, we demonstrate that phosphorylation of cortactin by AKT1 is important for mutant PI3K-enhanced cell migration and invasion. Our study describes a quantitative and global approach for identifying mutation-specific signalling events and for discovering novel signalling molecules as readouts of pathway activation or potential therapeutic targets. 1 McKusick-Nathans Institute of Genetic Medicine and Department of Biological Chemistry, Johns Hopkins University School of Medicine, 733 North Broadway, BRB 527, Baltimore, Maryland 21205, USA.
  • SS18-SSX–Dependent YAP/TAZ Signaling in Synovial Sarcoma

    SS18-SSX–Dependent YAP/TAZ Signaling in Synovial Sarcoma

    Published OnlineFirst February 27, 2019; DOI: 10.1158/1078-0432.CCR-17-3553 Translational Cancer Mechanisms and Therapy Clinical Cancer Research SS18-SSX–Dependent YAP/TAZ Signaling in Synovial Sarcoma Ilka Isfort1,2, Magdalene Cyra1,2, Sandra Elges2, Sareetha Kailayangiri3, Bianca Altvater3, Claudia Rossig3,4, Konrad Steinestel2,5, Inga Grunewald€ 1,2, Sebastian Huss2, Eva Eßeling6, Jan-Henrik Mikesch6, Susanne Hafner7, Thomas Simmet7, Agnieszka Wozniak8,9, Patrick Schoffski€ 8,9, Olle Larsson10, Eva Wardelmann2, Marcel Trautmann1,2, and Wolfgang Hartmann1,2 Abstract Purpose: Synovial sarcoma is a soft tissue malignancy Results: Asignificant subset of synovial sarcoma characterized by a reciprocal t(X;18) translocation. The chi- showed nuclear positivity for YAP/TAZ and their tran- meric SS18-SSX fusion protein acts as a transcriptional dysre- scriptional targets FOXM1 and PLK1. In synovial sarco- gulator representing the major driver of the disease; however, ma cells, RNAi-mediated knockdown of SS18-SSX led to the signaling pathways activated by SS18-SSX remain to be significant reduction of YAP/TAZ-TEAD transcriptional elucidated to define innovative therapeutic strategies. activity. Conversely, SS18-SSX overexpression in SCP-1 Experimental Design: Immunohistochemical evaluation cells induced aberrant YAP/TAZ-dependent signals, mech- of the Hippo signaling pathway effectors YAP/TAZ was per- anistically mediated by an IGF-II/IGF-IR signaling loop formed in a large cohort of synovial sarcoma tissue specimens. leading to dysregulation of the Hippo effectors LATS1 SS18-SSX dependency and biological function of the YAP/TAZ and MOB1. Modulation of YAP/TAZ-TEAD–mediated Hippo signaling cascade were analyzed in five synovial sarco- transcriptional activity by RNAi or verteporfintreatment ma cell lines and a mesenchymal stem cell model in vitro.