Pruneski, Justin
(2012)
IDENTIFICATION AND CHARACTERIZATION OF FACTORS REQUIRED FOR REPRESSION OF THE YEAST SER3 GENE BY SRG1 INTERGENIC TRANSCRIPTION.
Doctoral Dissertation, University of Pittsburgh.
(Unpublished)
Abstract
Recent studies have shown that transcription is widespread in eukaryotic genomes occurring over noncoding regions (ncDNA) as well as protein-coding genes. This pervasive transcription can have profound effects on DNA-mediated processes such as the regulation of gene expression. In one well characterized example, repression of the Saccharomyces cerevisiae SER3 gene was shown to be dependent on transcription of SRG1 from ncDNA initiating within intergenic DNA 5’ of SER3 and extending across the SER3 promoter region, preventing transcription factors from binding to the SER3 promoter.
To understand the details of this transcription interference mechanism, I performed a genetic screen in yeast to identify factors required for SER3 repression. I identified 21 candidates including known regulators of SER3, factors involved in histone gene expression, and transcription elongation factors. The regulators of histone gene expression led us to discover a role for chromatin in SER3 repression. The combined activities of the Swi/Snf chromatin remodeling factor, the HMG-like factor Spt2, and the Spt6 and Spt16 histone chaperones allow SRG1 transcription to deposit and maintain nucleosomes over the SER3 promoter to prevent transcription factors from binding and activating SER3.
Through investigations of the transcription elongation factors identified in the screen, I uncovered a role for the Paf1 transcription elongation complex in SER3 repression. I found that SER3 repression is primarily dependent on the Paf1 and Ctr9 subunits of this complex, with minor contributions by the Rtf1, Cdc73, and Leo1 subunits. Importantly, the defect in SER3 repression in strains lacking Paf1 subunits is not a result of reduced SRG1 transcription or reduced levels of known Paf1 complex-dependent histone modifications. Rather, we find that strains lacking subunits of the Paf1 complex exhibit reduced nucleosome occupancy and reduced recruitment of Spt16 and, to a lesser extent, Spt6 at the SER3 promoter, suggesting a novel role for the complex. Taken together, my work demonstrates that SER3 repression is mediated by nucleosome occupancy of the SER3 promoter, which is facilitated by the disassembly and assembly of nucleosomes by Spt6 and Spt16, which requires Spt2 and the Paf1 complex during SRG1 transcription.
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Details
| Item Type: |
University of Pittsburgh ETD
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| Status: |
Unpublished |
| Creators/Authors: |
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| ETD Committee: |
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| Date: |
1 February 2012 |
| Date Type: |
Publication |
| Defense Date: |
1 December 2011 |
| Approval Date: |
1 February 2012 |
| Submission Date: |
8 December 2011 |
| Access Restriction: |
No restriction; Release the ETD for access worldwide immediately. |
| Number of Pages: |
217 |
| Institution: |
University of Pittsburgh |
| Schools and Programs: |
Dietrich School of Arts and Sciences > Biological Sciences |
| Degree: |
PhD - Doctor of Philosophy |
| Thesis Type: |
Doctoral Dissertation |
| Refereed: |
Yes |
| Uncontrolled Keywords: |
Yeast, transcription, chromatin, nucleosome occupancy, non-coding DNA, gene expression |
| Date Deposited: |
01 Feb 2012 14:29 |
| Last Modified: |
15 Nov 2016 13:55 |
| URI: |
http://d-scholarship.pitt.edu/id/eprint/10740 |
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