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Brewis, Hilary Tyne

University of British Columbia

Chromatin is involved in all aspects of genome function and its dynamic structure is a key factor in many fundamental cellular processes such as transcription, DNA replication, and DNA repair. A variety of mechanisms regulate chromatin structure including the incorporation of histone variants. H2A.Z, encoded by the non-essential HTZ1 gene in budding yeast, is an evolutionarily conserved histone variant that replaces H2A in 5-10% of nucleosomes in a reaction catalyzed by the SWR1 complex, a conserved ATP-dependent chromatin remodeler. Specifically, H2A.Z is incorporated near centromeres, at the border of heterochromatic domains, and at the majority of transcription start sites. In this dissertation I utilized a combination of genome-wide approaches along with traditional molecular biology techniques to uncover how H2A.Z’s unique identity as a histone variant contributes to gene expression regulation in budding yeast. First, I identified three amino acid regions in the C-terminal half of H2A.Z that can confer specific H2A.Z-identity to H2A. Remarkably, the combination of only 9 amino acid changes, the H2A.Z M6 region, K79 and L81 (two amino acids in the α2-helix), were sufficient to fully recapitulate wild-type growth in genotoxic stress. Furthermore, combining H2A.Z K79 and L81, the M6 region, and the C-terminal tail was sufficient for expression of H2A.Z-dependent heterochromatin-proximal genes and GAL1 derepression. I then determined the impact of H2A.Z incorporation on gene expression during DNA replication stress by generating genome-wide mRNA transcript profiles for wild-type, htz1Δ, swr1Δ, and htz1Δswr1Δ mutants before and after exposure to hydroxyurea (HU). My findings revealed that removing H2A.Z incorporation did not result in mass dysregulation of gene expression, however H2A.Z was required for proper wild-type expression of several HU-induced genes. Collectively my dissertation uncovered the amino acids responsible for H2A.Z-specific identity and explored the importance of this unique identity in gene induction during cellular stress.

Text

2024-04-12

Attribution-NonCommercial-NoDerivatives 4.0 International

http://hdl.handle.net/2429/87779

Medical Genetics

University of British Columbia

UBCV

http://creativecommons.org/licenses/by-nc-nd/4.0/