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Jenel Nixon Cobb
Thesis Advisor: Marc. R. Gartenberg, PhD
Graduate Program of Cellular and Molecular Pharmacology
School of Public Health/Research Bldg.
Monday, November 9, 2009
The periphery of the nucleus constitutes a domain critical for transcriptional control. In yeast, many expressed genes associate with nuclear pore components for activation and mRNA export. Transcriptionally silenced genes also accumulate at the nuclear periphery and this contributes to gene repression. Esc1 is a perinuclear protein that appears to play a role in both processes. The protein and the DNA end binding protein Ku define redundant mechanisms that anchor telomeres and other silenced chromosomal domains in perinuclear positions. Esc1 also participates in mRNA surveillance by helping block transport of improperly spliced mRNAs out of the nucleus. To investigate how Esc1 exerts these pleitropic functions, I adapted the split-ubiquitin two-hybrid screen to identify Esc1 interacting proteins. This screen utilizes two complimentary fragments of ubiquitin, which when reconstituted generate an easily measured phenotypic signal. I have identified several putative Esc1 interacting partners whose functions range from transcriptional elongation to rRNA processing to the ER degradation pathway.
I have focused on two proteins recovered by my screen, Cue1 and Ubc6. Ubc6 is an E2 ubiquitin-conjugating enzyme that resides at the ER, yet executes nuclear functions because of the continuity of the ER and the inner nuclear membrane. Cue1 is a trans-membrane protein of the ER that anchors another E2 ubiquitin-conjugating enzyme, Ubc7. I have shown that Ubc6 and Cue1 influence the pathway by which Esc1 positions silenced chromatin. Specifically, I have shown that both proteins, like Esc1, are required for mitotic segregation of plasmids bound by the silent chromatin protein Sir4. I have also shown that Ubc6 and Cue1, like Esc1, are required for the nuclear retention of unspliced pre-mRNAs. These results indicate that the ER degradation contributes to the pathways controlled by Esc1.