Publication Date

2010

Document Type

Dissertation

Committee Members

Gerald M. Alter (Committee Member), Steven J. Berberich (Advisor), Michael Leffak (Committee Chair), Robert W. Putnam (Committee Chair), Paula M. Termuhlen (Committee Member)

Degree Name

Doctor of Philosophy (PhD)

Abstract

A comprehensive understanding of the molecular signaling pathways that regulate cell growth and proliferation is essential in the realization of new therapeutic options to facilitate early detection and eradication of malignancy. Understanding the transcriptional regulation of the YPEL3 and FHIT genes forms the basis of this dissertation. YPEL3, or Yippee-like 3, is a newly identified p53 target gene that inhibits tumor cell growth and is thus itself, a novel tumor suppressor gene. FHIT, or Fragile histidine triad, is a well known tumor suppressor gene and is regulated at the transcriptional level by another growth inhibitory protein, FOXO3a, a Forkhead box transcription factor.

Our laboratory has determined that YPEL3 is a direct transcriptional target of the tumor suppressor gene p53. The first section of this dissertation provides significant experimental evidence to validate this observation. Briefly, YPEL3 was shown to be upregulated downstream of p53 protein stabilization in a microarray screen that explored global gene expression modulation after RNAi-mediated reduction of p53's negative regulators, Hdm2 and HdmX. Genotoxic stress induced by treatment with DNA-damaging agents resulted in stabilization of p53 protein along with elevation of YPEL3 transcript and protein levels. Moreover, there exists a cis-acting p53 response element within the YPEL3 promoter that is bound by p53 in response to this stress. YPEL3 also elicits growth inhibition and decreases in colony formation when expressed in tumor cells. It is apparent that cells which express exogenous YPEL3 are forced into a permanent cell cycle arrest, termed "premature senescence." Similar growth suppressive phenotypes are typical of many other known p53 target genes. However, most of these genes are associated with the regulation of transient inhibition of cell division or the induction of apoptosis. YPEL3 is unique in its ability to trigger premature cellular senescence. Indeed, YPEL3 stands out among many other p53 targets because it is among the first to play a role in this process.

To further understand the mechanism of YPEL3-induced senescence, the second portion of this dissertation focuses on the generation of a three-dimensional model of YPEL3's protein structure. It was hypothesized that predicting YPEL3's protein structure may also aid in understanding the molecular events involved in the induction and maintenance of premature senescence. Identifying structural homology between a predicted model of YPEL3 and other known structures may provide new insights into this process. This would especially be the case if homology existed between the predicted structure of YPEL3 and other proteins that have established functions in stress-responsive or senescence-associated molecular pathways. By using two independent structural prediction algorithms (Rosetta ab initio and I-TASSER), I have been able to estimate a three-dimensional model of the YPEL3 protein which has significant structural homology to a family of Methionine oxidoreductases. These enzymes catalyze redox-mediated antioxidant reactions which dissipate intracellular reactive oxygen species (ROS) through the repair of proteins damaged by ROX-mediated methionine to methionine sulfoxide oxidation. It is even plausible that YPEL3 could be involved in the mediation of cellular senescence in response to severe oxidative stress. If this is the case, the implications of this speculation, linking the oxidative stress response to YPEL3-dependent cellular senescence, are indeed profound. YPEL3 may not only play a role in the molecular biology of cancer but also may be involved in the cellular oxidative stress response as it relates to senescence and aging.

Page Count

219

Department or Program

Biomedical Sciences

Year Degree Awarded

2010

Download


Share

COinS