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CREG1: Its Role As a Master Regulator of Liver Function Iffat Jahan Eastern Illinois University

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CREG1: Its Role As a Master Regulator of Liver Function Iffat Jahan Eastern Illinois University Eastern Illinois University The Keep Masters Theses Student Theses & Publications 2019 CREG1: Its Role as a Master Regulator of Liver Function Iffat Jahan Eastern Illinois University Recommended Citation Jahan, Iffat, "CREG1: Its Role as a Master Regulator of Liver Function" (2019). Masters Theses. 4477. https://thekeep.eiu.edu/theses/4477 This Dissertation/Thesis is brought to you for free and open access by the Student Theses & Publications at The Keep. It has been accepted for inclusion in Masters Theses by an authorized administrator of The Keep. For more information, please contact [email protected]. TheGraduate School� E>srf.RN111 1�11\USl\ 'F.R.S1n· Thesis Maintenance and Reproduction Certificate FOR: Graduate Candidates Completing Theses in Partial Fulfillment of the Degree Graduate Faculty Advisors Directing the Theses RE: Preservation, Reproduction, and Distribution of Thesis Research Preserving, reproducing, and distributing thesis research is an important part of Booth Library's responsibility to provide access to scholarship. In order to further this goal, Booth Library makes all graduate theses completed as part of a degree program at Eastern Illinois University available for personal study, research, and other not-for­ profit educational purposes. Under 17 U.S.C. § 108, the library may reproduce and distribute a copy without infringing on copyright; however, professional courtesy dictates that permission be requested from the author before doing so. Your signatures affirm the following: • The graduate candidate is the author of this thesis. •The graduate candidate retains the copyright and intellectual property rights associated with the original research, creative activity, and intellectual or artistic content of the thesis. •The graduate candidate certifies her/his compliance with federal copyright law (Title 17 of the U.S. Code) and her/his right to authorize reproduction and distribution of all copyrighted materials included in this thesis. • The graduate candidate in consultation with the faculty advisor grants Booth Library the nonexclusive, perpetual right to make copies of the thesis freely and publicly available without restriction, by means of any current or successive technology, including but not limited to photocopying, microfilm, digitization, or internet. •The graduate candidate acknowledges that by depositing her/his thesis with Booth Library, her/his work is available for viewing by the public and may be borrowed through the library's circulation and interlibrary loan departments, or accessed electronically. The graduate candidate acknowledges this policy by indicating in the foll7g manner: ___Yes, I wish to make accessible this thesis for viewing by the public ___No, I wish to quarantine the thesis temporarily and have included the Thesis Withholding Request Form • The graduate candidate waives the confidentiality provisions of the Family Educational Rights and Privacy Act (FERPA) (20 U. S. C. § 1232g; 34 CFR Part 99) with respect to the contents of the thesis and with respect to information concerning authorship of the thesis, including name and status as a student at Eastern Illinois University. I have conferred with my graduate faculty advisor. My signature below indicates that I have read and agree with the above statements, and hereby give my permission to allow Booth Library to reproduce and distribute my thesis. My adviser's signature indicates concurrence to reproduce and distribute the tl),e�s. uraouate 1..anu1udlt:.>1�11dLu1e IFFAT JAl·MN Printed Name �-lw, M Sc.Wn<.g. Graduate Degree Program Date Please submit in duplicate. CREG1: ITS ROLE AS MASTER REGULATOR OF LIVER FUNCTION (TITLE) BY IFFAT JAHAN THESIS SUBMITTED IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF Masters of Science IN THE GRADUATE SCHOOL, EASTERN ILLINOIS UNIVERSITY CHARLESTON, ILLINOIS 2019 YEAR I HEREBY RECOMMEND THAT THIS THESIS BE ACCEPTED AS FULFILLING THIS PART OF THE GRADUATE DEGREE CITED ABOVE f4L/? DATE (lµfDATE !f!/11 !f DATE DA�f THESIS COMMITIEE MEMBER DATE THESIS COMMITIEE MEMBER DATE CREG1: Its Role as a Master Regulator of Liver Function Iffat Jahan Department of Biological Science Eastern Illinois University Charleston, IL-61920 2019 © 2019 by I ffat J ahan ii COMMITTEE IN CHARGE OF CANDIDACY Gary A. Bulla, Ph.D. Advisor Anthony Olouch, Ph.D. Britto Nathan, Ph.D. iii Abstract Mammalian development involves a complex system of regulatory signals and reactions resulting in highly differentiated cell types with specific structure and function. The liver is a major organ that has been studied extensively to understand underlying genetic processes responsible for specification, establishment and maintenance of tissue identity. Hepatoma and hepatoma variant cell lines have been used as models to understand genetic networks responsible for liver function. Whole genome microarray analysis of hepatocyte cell lines has revealed candidate genes that may serve as regulators or master regulators of liver specificity. In two previous studies in our lab, the role of candidate gene Cellular Repressor of EIA Stimulated Gene (CREG1) in regulation of liver-specific gene expression was determined using transfection studies combined with utilization of quantitative real-time polymerase chain reaction (qRTPCR). Both studies found strong activation (10-100-fold) of transcription factor Hn/6 and the downstream gene Serpinal (a marker gene used to identify liver function). These findings suggested that CREG 1 might act through Hnf6 to regulate Serpinal via a Locus Control Region (LCR). However, while one study showed that CREGI overexpression in a hepatoma variant cell line resulted in modest activation of liver-specific transcription factors Hnfl, Hn/4 and Hnf3, the second study suggested robust activation of these genes as measured by qRT-PCR. In the current study, a CREG1 expression vector was reintroduced into one of the variant cell lines (H 11) and gene activation profiles monitored to establish validity of the previous studies. However, we postulated that CREG-1 overexpression, validated by qRT-PCR, can activate expression of hepatocyte transcription factor Hn/6 and the downstream gene Serpinal along with other liver­ specific downstream genes. Results showed that CREGl can fully rescue expression of liver- iv specific genes including transactivation genes, suggesting its role as a master regulator of liver function. CREG l action appears to act at least partially through HNF6 gene activation, as well as through activation of HNF4/HNF1 pathway, both of which act to increase expression of the serpin locus through LCR activation through HNF6. v Dedicated to My Parents vi Acknowledgement I would like to express my sincere gratitude to all individuals who helped me in completing this project while working on my degree. It would not have been possible without their constant support, encouragement and help in facilitating this work. It is a pleasure to acknowledge them for their motivation, guidance and numerous suggestions. I express my deep sense of thanks and gratitude to my advisor and guide Dr. Gary A. Bulla, whose keen interest and, above all, his overwhelming attitude in helping his students. He has been largely responsible for my completing of this work. His timely advice, meticulous scrutiny, scholarly advice and scientific approach have helped me to accomplish this work in an interesting and expanding field. I am thankful to Dr. Britto P Nathan for providing indispensable advice, information and support on several aspects of my project while working in laboratory, and who make me look at research and my work in different ways. I owe a deep sense of gratitude to Dr. Anthony Oluoch for his keen interest at every stage of my research. His timely suggestions along with with kindness, enthusiasm and dynamism have enabled me to complete my thesis. Moreover, for providing an opportunity to serve as a teaching assistant in his class and helping me to learn the art of teaching and learning at the same time. The assistance, co-operation and experience of my fellow lab members was essential for completion of this project. Of course, family and friends who were there to give me the extra push necessary to succeed. Last, but not least, I am thankful to the Department of Biological Sciences and EasternIllinois University forthis opportunity to be trained and to be an alumnus. vii Table of Contents Abstract ........................................... ..................................................... ..................................................... iv Acknowledgement ................................................................................................................................... vi Table of Contents ...................................................................................................................................vii List of Figures .......................................................................................................................................... ix List of Tables ............................................................................................................................................. x Chapter 1 : Introduction ....... ............................................................................................. ................. 1 1.1. Liver Development .................................................................................................................... ! 1.2. Cell Culture Model Systems ................. ...................................
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