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Global genome nucleotide excision repair factors and the ubiquitin-proteasome system regulate the DNA damage response in Saccharomyces cerevisiae

Humphryes, Neil 2010. Global genome nucleotide excision repair factors and the ubiquitin-proteasome system regulate the DNA damage response in Saccharomyces cerevisiae. PhD Thesis, Cardiff University.

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The DNA damage response triggers a complex regulatory network to protect genomic integrity and promote cell survival by inducing effects such as cell cycle arrest, upregulation of dNTP synthesis and DNA repair. The Nucleotide Excision Repair (NER) pathway removes a broad array of DNA lesions by excising the damage with surrounding DNA and resynthesising the DNA using the undamaged strand as the template. NER consists of two sub-pathways that remove lesions from transcribed DNA (TC-NER), or from the rest of the genome (GG-NER). It has been proposed more recently that NER factors have additional regulatory roles in the DNA damage response. The initial aim of the study was to identify a possible regulatory interaction between the Abf1 protein and the 19S proteasome. abfl and sug (proteasome) mutant alleles were employed to identify a functional interaction between the factors but a significantly altered mutant phenotype could not be identified. The study proceeded to investigate a regulatory role of the Rad4-Rad23 NER complex in the DNA damage response via Rad7-dependent ubiquitination of Rad4. Previous evidence suggested that post-UV Rad4 ubiquitination has a regulatory role in DNA damage-responsive transcription. Global analysis of gene expression revealed that genes involved in dNTP synthesis, including the Ribonucleotide Reductase (RNR) pathway, were misregulated in a strain unable to ubiquitinate Rad4. The study progressed to investigate the function of the Rad4-Rad23 complex in regulation of the RNR pathway. It was discovered that constitutive activation or expression of the RNR pathway could suppress the UV sensitivity of the E3 ligase-defective strain, thus suggesting that the Rad7-E3 ligase has a role in regulating cellular dNTP levels in response to DNA damage. The Rad4-Rad23 complex binds at the DUN1 promoter, a positive regulator of the RNR pathway. E3 ligase-defective mutant strains were also found to exhibit defective cell cycle progression in the presence and absence of DNA damage. This defective cell cycle progression is suppressed by upregulating the RNR pathway. This further supports a role of the Rad7 E3 ligase in regulation of cellular dNTP levels. These results indicate a role of the Rad7 E3 ubiquitin ligase in regulation of cellular dNTP levels in the DNA damage response.

Item Type: Thesis (PhD)
Status: Unpublished
Schools: Medicine
Subjects: Q Science > QH Natural history > QH426 Genetics
R Medicine > R Medicine (General)
Date of First Compliant Deposit: 30 March 2016
Last Modified: 10 Jan 2018 03:54

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