University of Calgary PRISM: University of Calgary's Digital Repository Graduate Studies Legacy Theses 2008 A Role of ING1 Proteins in Cell Senescence Soliman, Mohamed A. Soliman, M. A. (2008). A Role of ING1 Proteins in Cell Senescence (Unpublished master's thesis). University of Calgary, Calgary, AB. doi:10.11575/PRISM/17019 http://hdl.handle.net/1880/46814 master thesis University of Calgary graduate students retain copyright ownership and moral rights for their thesis. You may use this material in any way that is permitted by the Copyright Act or through licensing that has been assigned to the document. For uses that are not allowable under copyright legislation or licensing, you are required to seek permission. Downloaded from PRISM: https://prism.ucalgary.ca UNIVERSITY OF CALGARY A Role of ING1 Proteins in Cell Senescence by Mohamed A. Soliman A THESIS SUBMITTED TO THE FACULTY OF GRADUATE STUDIES IN PARTIAL FULFILMENT OF THE REQUIREMENTS FOR THE DEGREE OF MASTER OF SCIENCE DEPARTMENT OF BIOCHEMISTRY AND MOLECULAR BIOLOGY CALGARY, ALBERTA SEPTEMBER 2008 © Mohamed A. Soliman 2008 The author of this thesis has granted the University of Calgary a non-exclusive license to reproduce and distribute copies of this thesis to users of the University of Calgary Archives. Copyright remains with the author. Theses and dissertations available in the University of Calgary Institutional Repository are solely for the purpose of private study and research. They may not be copied or reproduced, except as permitted by copyright laws, without written authority of the copyright owner. Any commercial use or publication is strictly prohibited. The original Partial Copyright License attesting to these terms and signed by the author of this thesis may be found in the original print version of the thesis, held by the University of Calgary Archives. The thesis approval page signed by the examining committee may also be found in the original print version of the thesis held in the University of Calgary Archives. Please contact the University of Calgary Archives for further information, E-mail: [email protected] Telephone: (403) 220-7271 Website: http://www.ucalgary.ca/archives/ Abstract The ING1 proteins affect cell growth and apoptosis by modulating chromatin structure. The major splicing isoforms of the ING1 locus are ING1a and INGlb. While INGlb mediates an apoptotic response, the function of ING1a is currently unknown. Here, we show that the INGla:INGlb ratio is altered by several-fold in senescent, compared to low passage primary fibroblasts. Furthermore, we show that INGla induces many features of senescence. To expand our understanding of ING1 functions, a cross-species in silico approach was used to identify potential human ING1 interacting proteins. Novel interactions, with p38MAPK and MEKK4, were identified and biochemically verified. None of the validated interactions were predicted by conventional tools tested. The Bioinformatics approach described can be used to predict novel interactions for other human proteins with yeast homolog(s). These data expand our knowledge of the roles that ING proteins play and demonstrate a novel role for ING1 proteins in differentially regulating senescence. ii Acknowledgements I would like to deeply thank my mentor Dr. Karl Riabowol for consistent support, guidance and encouragement. Thanks for his patience while my experiments were not working and his wisdom in dealing with both my frustrations and triumphs. Thanks for his welcoming character and very easy-going personality that encouraged me to overcome the cultural and language barriers that I encountered, in a relatively short time. He also let me discover significant questions to address in my research, and he respected my opinion and research findings. Thanks for his confidence in my scientific knowledge and for giving me the opportunity to improve myself in writing and in criticizing research work. Above all, I would like to thank him for the intellectual freedom he gave me, his trust in my capacity to think independently and accept my ideas even when we have different scientific opinions. Thanks for the great friendly journey! I would like to thank Drs. Randal Johnston and Jerome B. Rattner for the wonderful discussions we have had about science, politics, and global aspects of life, while serving on my supervisory committee. Their open-minded personalities helped me think out the box, celebrate diversity of ideas, and accept positive criticism. Thanks for their great help and advice during my Master’s thesis and their great support to continue my academic career the way I would like. I would also like to thank Dr. Greg Moorhead for serving on my thesis examination committee...much appreciated! Thanks also to all the Riabowol Lab members, past and current, for the collegial environment and their friendship. Special thanks to Dr. Michael Russell for helping with my first steps in the lab, Ms. Annie Fong for teaching me tissue culture techniques, and Drs. Xiaolan Feng and Keiko Suzuki for helping me appreciate science and teaching me how it is essential to be both constantly well-driven and a hard worker to succeed in academia. Thanks to Dr. Philip Berardi for establishing the first solid observations that led to my thesis project, Pinaki Bose and Sitar Shaw for revising my thesis writing. Thanks also to all the members of SACRI; I learned new techniques, shared ideas, used lab facilities, or borrowed reagents from almost every single lab in the cancer biology group, which was iii greatly appreciated. Thanks to Elizabeth Long and Alexander Klimowicz for the fantastic long discussions about the future of science in the country, and to Drs. Arcellana-Panlilio and Wang in the Sothern Alberta Microarray Facility for helping me with numerous microarray experiments. I also appreciate the help I received from the Bioinformatics group, especially Paul Gordon for the great collaboration we had with them. I would like to acknowledge the Alberta Heritage Foundation for Medical Research, the Alberta Cancer Board, and the Canadian Institute of Health Research for generous awards and studentships. Finally, thanks to my family and my friends here and back home for their constant support and love. We shared frustrations, happiness and success moments. Thanks for understanding the time- and effort-demanding nature of my work. Without you, my success would not have been possible. iv Table of Contents Approval Page ................................................................................................................... ii Abstract.............................................................................................................................. ii Acknowledgements .......................................................................................................... iii Table of Contents ...............................................................................................................v List of Tables....................................................................................................................viii List of Figures and Illustrations........................................................................................ix CHAPTER 1: INTRODUCTION....................................................................................1 Section I. The ING tumor suppressors ........................................................................2 ING tumor suppressors; interacting partners and biological functions .............3 Structural features of the ING family ....................................................................5 ING1 splicing isoforms.............................................................................................6 ING and p53: Do they need each other? ................................................................7 A role for ING stress signalling...............................................................................8 A role for ING in reading the histone code ..........................................................10 Section II. Cell senescence...........................................................................................22 Cell Senescence .......................................................................................................23 Causes of aging and senescence.............................................................................23 Tumor suppressors and senescence: a coherent link ..........................................25 Signalling pathways mediating senescence ..........................................................25 Oncogene-induced senescence (OIS).....................................................................28 Senescence markers................................................................................................29 Epigenetic contributions to senescence.................................................................32 Section III. Protein interaction networks ..................................................................34 Biochemical methods of determining large scale protein-protein interactions (PPI) ................................................................................................................35 Computational prediction of PPIs ........................................................................37 Genomic methods.............................................................................................38 Domain-based prediction of PPI ....................................................................39