All ETDs from UAB

Advisory Committee Chair

Sunnie R Thompson

Advisory Committee Members

David Bedwell

Ching-Yi Chen

Elliot Lefkowitz

Casey Morrow

Document Type

Dissertation

Date of Award

2011

Degree Name by School

Doctor of Philosophy (PhD) Heersink School of Medicine

Abstract

Translation of the majority of eukaryotic mRNAs is initiated upon recognition of its 5' cap structure by translation initiation factors in so-called cap-dependent translation. Cap-dependent translation is repressed during cell stress and translation initiation by internal ribosome entry sites (IRESs) predominates. IRESs, located in the 5' untranslated region of specific mRNAs, recruit the ribosome independent of the 5' cap and are essential regulatory elements in many RNA viruses and cellular messages. The dysregulation of cellular IRESs has been linked to cancer because they regulate the expression of genes involved in growth, development and stress response. In comparison to cap-dependent translation, the mechanism of IRES-mediated translation is poorly understood. The intergenic region (IGR) IRES from the cricket paralysis virus serves as a model IRES because it is the most simplified one known, however little is known about other IGR IRESs. We compared the activity of nine IGR IRESs in yeast and HeLa cells. Our analysis demonstrated that all IGR IRES are functional and that the strength of the IRES is influenced by the ribosome binding domain and the sequence downstream of the IRES. We also discovered that many viral and cellular IRESs use a common strategy for translation initiation which requires ribosomal protein S25 (Rps25). Deletion of Rps25 in yeast, or depletion in human cells, modestly affects global translation but causes a specific defect in IRES-mediated translation which leads to inefficient replication of IRES-containing viruses and lower protein expression from IRES-containing cellular mRNAs. Rps25 is also vital for prostate cancer (PCa) survival in the bone. Rps25 depletion in the highly aggressive ARCaPM PCa cell line resulted in a failure of these cells to thrive in vivo, most likely because these cells were less invasive and more susceptible to apoptosis in vitro. Because Rps25 depletion specifically affects IRES-mediated translation, these data strongly suggest that translation by cellular IRESs is essential for establishing prostate cancer in the bone. Taken together, this work revealed the widespread, conserved and vital role of Rps25 in IRES-mediated translation and suggests that Rps25 would be a good target for drug development for anti-viral and anti-cancer therapeutics.

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