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Role of SPDEF in Prostate Cancer

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Role of SPDEF in Prostate Cancer Role of SPDEF in Prostate Cancer A thesis submitted to the Graduate School of the University of Cincinnati in partial fulfillment of the requirements for the degree of Master of Science in the Department of Developmental Biology of the College of Medicine by Chen Gao B.S. Beijing Normal University June 2012 Committee Chair: Tanya V. Kalin, M.D., Ph.D. Abstract SAM Pointed Domain Containing ETS Transcription Factor (SPDEF) is a new member of ETS transcription factor family that is originally discovered in the epithelium of prostate. Like other ETS family members, SPDEF is involved in multiple biological processes including cell fate determination and specification, cell proliferation, epithelial-to-mesenchymal transformation and migration. Although much work has been done to elucidate the role of SPDEF in normal physiological conditions and in cancers, its expression level in cancer is still controversial. Similarly, there is no conclusive message on the role of SPDEF in cancer despite the fact that several in vitro studies have been done. We first overviewed the discovery and properties of SPDEF, followed by a comprehensive review of the role of SPDEF, with a special focus on its role in the development of cancer. In the second chapter, we utilized a xenograft model to manipulate SPDEF level in mouse prostate cancer. After orthotopic injection, we found that overexpression of SPDEF resulted in smaller tumors in mice. We also showed that several target genes related to cell proliferation and migration are down-regulated by SPDEF overexpression. We confirmed down-regulation of these genes in prostate cancer cell lines. In fact, we found that SPDEF overexpression inhibited cell migration, resulting in reduced aggressiveness of tumor cells. In addition, SPDEF overexpression was shown to suppress tumor growth via alteration of cell cycle profile. Collectively, these data indicate that SPDEF suppresses tumor growth via inhibition of cell proliferation and migration. ii iii Acknowledgements I would like to express my gratitude to the following people for their assistance and support to make this thesis possible. My deepest gratitude goes to my advisor, Dr. Tanya Kalin, for her guidance during my graduate study. Her patience, support, and advice helped me overcome many challenges during my research in the lab. I want to thank her for giving me the chance to learn how to conduct research. I would like to express my sincere gratitude to my thesis committee members for their guidance and helpful discussions through my thesis work. I also thank the Molecular and Developmental Biology Graduate Program for offering me the opportunity of graduate training. I’m also grateful to Dr. Vladimir Kalinichenko and my colleagues in Kalin and Kalinichenko labs. I deeply appreciate their scientific input and technical assistance to help me go through my research. I also thank the co-workers in the Division of Neonatology and Pulmonary Biology for their help. Finally, I want to give special thanks to my parents who always love, support and encourage me. iv Table of contents Table of Contents Abstract ............................................................................................................................................. ii Acknowledgements .......................................................................................................................... iv Table of contents ............................................................................................................................... v List of Symbols and Abbreviations .................................................................................................. vi Chapter 1: Literature Review of SPDEF and Its Implication in the Pathogenesis of Cancer ........... 1 Overview of ETS family ........................................................................................................... 1 General Information of SPDEF ................................................................................................. 3 Expression Pattern of SPDEF ................................................................................................... 5 1. SPDEF expression in normal tissues ............................................................................. 5 2. SPDEF expression in cancers ........................................................................................ 6 Regulation of SPDEF ................................................................................................................ 8 SPDEF Targets ........................................................................................................................ 11 1. Target genes in normal development ........................................................................... 11 2. Target genes regulating cell proliferation and apoptosis ............................................. 13 3. Target genes regulating cell migration and metastasis ................................................ 14 Biological Functions of SPDEF .............................................................................................. 16 1. SPDEF and cell differentiation ................................................................................... 16 2. SPDEF and cell proliferation ...................................................................................... 18 3. SPDEF and cell migration ........................................................................................... 19 Summary and Future Perspectives .......................................................................................... 21 References ............................................................................................................................... 24 Chapter 2: The role of SPDEF in prostate cancer ........................................................................... 29 Abstract ................................................................................................................................... 29 Introduction ............................................................................................................................. 29 Results ..................................................................................................................................... 32 Overexpression of SPDEF Reduced Tumor Weight In Mouse Prostate Cancer ............. 32 SPDEF Suppresses Tumor Cell Migration and Invasion ................................................ 33 SPDEF Inhibits Tumor Cell Growth Via Change Of Cell Cycle Profile ......................... 33 Discussion ............................................................................................................................... 34 Materials and Methods ............................................................................................................ 38 Figures..................................................................................................................................... 41 References ............................................................................................................................... 48 v List of Symbols and Abbreviations Agr2 anterior gradient 2 (Xenopus laevis) Akt also termed as protein kinase B AR androgen receptor ATOH1 atonal homolog 1 Cdc25b cell division cycle 25 homolog B CDK cyclin-dependent kinase CIP1 cyclin-dependent kinase inhibitor 1, also termed as p21 CK2 casein kinase 2 EMSA electrophoretic mobility shift assay ERG Ets related gene ETS E26 transformation-specific ETV-1 ETS translocation variant 1 FGF fibroblast growth factor FoxJ1 foxhead box protein J1 FoxM1 foxhead box protein M1 GFI1 growth factor independent protein 1 hPSE human prostate-specific ETS IAP inhibitor of apoptosis KLF4 Krueppel-like factor 4 LC-MS liquid chromatography - mass spectrometry MAPK MAPK mitogen activated protein kinase mammary serine protease inhibitor, also termed as Maspin SerpinB5 MMP-2 matrix metalloproteinase-2 MMP-7 matrix metalloproteinase-7 MMP-9 matrix metalloproteinase-9 mPSE mouse prostate-specific ETS MS mass spectrometry mTOR mammalian target of rapamycin Nkx3-1 NK3 homeobox 1 PDFGRα alpha-type platelet-derived growth factor receptor PEST domain proline, glutamic acid, serine, and threonine-rich domain PI Propidium iodide PI3K phosphoinositide 3-kinase PSA prostate-specific antigen RUNX1 Runt-related transcription factor 1 RUNX2 Runt-related transcription factor 2 Scgb1a1 secretoglobin, family 1A, member 1 (uteroglobin) serine (or cysteine) peptidase inhibitor, clade B Serpinb3a (ovalbumin), member 3a vi Sftpa surfactant protein A Sox17 SRY (sex determining region Y)-box 17 SPDEF SAM pointed domain containing ETS transcription factor Sqstm1 Sequestosome-1 Stmn stathmin TGFβ transforming growth factor beta TMPRSS2 transmembrane protease, serine 2 TRAMP transgenic adenocarcinoma of the mouse prostate thyroid transcription factor 1, also termed as Nkx2-1 TTF-1 (NK2 homeobox 1) uPA urokinase-type plasminogen activator VEGF vascular endothelial growth factor Zeb2 zinc finger E-box-binding homeobox 2 vii Chapter 1: Literature Review of SPDEF and Its Implication in the Pathogenesis of Cancer Overview of ETS family The E26 transformation-specific (ETS) family is one of the largest families of transcription factor proteins with currently 27 and 26 Ets genes found in humans and mice, respectively [1]. The evolutionarily conserved ETS domain is responsible for specific DNA binding and recognition of GGA(A/T) consensus sequence [2].
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