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Permanent link (DOI): https://doi.org/10.7939/R34B0J

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Studies on the control of tRNA transcription by the replication stress checkpoint Open Access

Descriptions

Other title
Subject/Keyword
transcription
tRNA
replication
Type of item
Thesis
Degree grantor
University of Alberta
Author or creator
Clelland, Brett William
Supervisor and department
Schultz, Michael (Biochemistry)
Examining committee member and department
Campbell, Shelagh (Biological Sciences)
Engelke, David (Biological Chemistry)
Simmonds, Andrew (Cell Biology)
MacMillan, Andrew (Biochemistry)
Department
Department of Biochemistry
Specialization

Date accepted
2011-09-21T23:38:55Z
Graduation date
2011-11
Degree
Doctor of Philosophy
Degree level
Doctoral
Abstract
RNA polymerase III (RNAPIII) pre-initiation complexes at tRNA genes naturally cause replication fork pausing in the yeast Saccharomyces cerevisiae, and interference with replication is known to have deleterious effects on genome stability. It follows that repression of tRNA gene transcription could be advantageous to minimize replication perturbation. Consistent with this idea, our lab has previously reported that the replication stress checkpoint inhibits tRNA gene transcription. Here, I describe how repression by checkpoint signalling, induced by treatment with the replication inhibitor hydroxyurea (HU), is associated with RNAPIII pre-initiation complex disassembly at tRNA genes. In addition, I show that active checkpoint signals likely impinge on Maf1, a key negative regulator of RNAPIII transcription, to signal to tRNA genes during HU exposure. Next, I report that checkpoint signalling affects the protein complex assemblage at tRNA genes during normal proliferation. Inactivation of the replication stress checkpoint, which is associated with an induction of tRNA gene transcription, results in greater RNAPIII occupancy at tRNA genes and a decrease in condensin association, condensin being an important tDNA localized complex that is vital for maintenance of genome integrity. Next, I extended these results by monitoring replication in cells with elevated tRNA gene transcription using cross-linking of replication proteins as proxy for replication fork movement. Despite the fact that tRNA gene transcription interferes with replication, by this method I detected no greater fork pausing at tRNA genes in strains with elevated transcription. These data are discussed in the context of current controversy in the literature about this type of replication perturbation. One possibility is that in cells unable to repress transcription, replication interference promotes greater genome instability in a way that does not include amplified fork pausing. Altogether, the results presented here are in harmony with the idea that the replication stress checkpoint functions to disassemble RNAPIII transcriptional machinery, likely to maintain genome stability. Lastly, I present preliminary data that identifies potential cell division cycle links to tRNA transcription. We propose a possible new pathway that restrains tRNA gene transcription involving Cdc28, the main cyclin-dependent kinase in yeast.
Language
English
DOI
doi:10.7939/R34B0J
Rights
License granted by Clelland Brett (brettc@ualberta.ca) on 2011-09-10T11:15:17Z (GMT): Permission is hereby granted to the University of Alberta Libraries to reproduce single copies of this thesis and to lend or sell such copies for private, scholarly or scientific research purposes only. Where the thesis is converted to, or otherwise made available in digital form, the University of Alberta will advise potential users of the thesis of the above terms. The author reserves all other publication and other rights in association with the copyright in the thesis, and except as herein provided, neither the thesis nor any substantial portion thereof may be printed or otherwise reproduced in any material form whatsoever without the author's prior written permission.
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