Wu, Chun Kit Andrew; (2020) Control of divergent noncoding transcription in Saccharomyces cerevisiae. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

The regulation of gene expression underlies all cellular processes and fundamentally enables complexity of eukaryotic organisms. Aberrant expression of noncoding RNAs can compromise normal gene expression. Gene promoters are inherently bidirectional and generate divergent noncoding RNAs along with protein-coding messenger RNAs. Chromatin and RNA turnover pathways limit expression of noncoding RNAs, but how sequence-specific transcription factors regulate divergent noncoding transcription and promoter directionality is not well understood. Here, I investigate how divergent transcription is repressed at highly expressed genes in Saccharomyces cerevisiae. I find that the sequence-specific transcription factor Rap1 limits divergent noncoding transcription at a large fraction of its target genes. Rap1 safeguards normal gene expression by limiting aberrant transcription that overlaps with neighbouring loci. Divergent RNAs initiate at or extremely close to Rap1 binding sites, indicating that Rap1 limits initiation of transcription from divergent core promoters. Stable binding of Rap1 near cryptic promoters is required and sufficient to suppress divergent transcription. Silencing cofactors or transcriptional coactivators associated with Rap1 are not required for repression of noncoding RNAs at promoters. In contrast, a small region within the Rap1 carboxy-terminal domain is required for repression of divergent transcription and affects interaction between Rap1 and the RSC chromatin remodeller. RSC and Rap1 regulate divergent transcription at Rap1-regulated gene promoters in distinct ways. Promoter output shifts from unidirectional to bidirectional transcription in the absence of Rap1, which is partially suppressed after co-depletion of Rap1 and RSC. RSC activity is not strictly required for divergent transcription suggesting that additional regulators also play important roles. I propose that certain sequence-specific transcription factors limit the access of transcription machinery and coactivators to divergent core promoters by steric hindrance, thereby providing directionality towards productive transcription of coding genes.

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