Browsing by Subject "nucleotide excision repair"
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Item Resistance of Bacillus Subtilis Spores Lacking Either Nucleotide Excision Repair or Spore Photoproduct Lyase to Ultraviolet (UV) Radiation from Artificial or Natural Sources(1996-06-01) Xue, Yaming; Tony Romeo; Ming-Chi Wu; Wayne L. NicholsonXue, Yaming, Resistance of Bacillus subtilis spores lacking either nucleotide excision repair or spore photoproduct lyase to ultraviolet (UV) radiation from artificial or natural sources. Master of Science (Biomedical Sciences), June 1996, pp., 4 tables, 12 illustrations, references, 38 titles. Exposure of bacterial spores to UV radiation causes the accumulation of a unique pyrimidine dimer in the DNA, “spore photoproduct” (SP). In Bacillus subtilis, two distinct DNA repair pathways are used for removal of SP: general nucleotide excision repair (the uvr parthway), or the SP-specific enzyme SP lyase (the spl pathway). Spores of four strains of Bacillus subtilis differing in their repair capabilities were irradiated under either artificial or solar UV. To determine the biologically-relevant cumulative UV dose under each irradiation condition, a sporocidal dosimeter was constructed. The results showed: (i) Both uvr and spl pathways contributed to the survival of spores under all tested conditions. The spl pathway was more efficient than uvr pathway in repairing the DNA damage caused by UV-C and solar UV-A, but no significant difference was noted in repairing DNA damage caused by UV-B or full-spectrum solar UV. (ii) Exposure of spores to solar UV can cause cellular lethal damage which is reparable by neither repair pathway.Item Studies of Checkpoint Responses Caused by Endogenous Oxidative DNA Damage in DNA Repair Deficient Saccharomyces Cerevisiae(2008-12-01) Pawar, Vaibhav; Alvarez, Rafael; Jiang, Yiwei; Krishnamoorthy, Raghu R.Pawar, Vaibhav., Studies of checkpoint responses caused by endogenous oxidative DNA damage in DNA repair deficient Saccharomyces cerevisiae. Doctor of Philosophy (Biomedical Sciences), December 2008, 163 pp., 21 illustrations, 189 references. In this dissertation project, I aimed to study checkpoint response of stationary phase yeast to DNA damage caused by basal oxidative stress. My study was focused on the regulation of Rad53 phosphorylation in different repair deficient strains to yeast. Rad53 plays decisive roles in cell cycle progression, cell death and transcriptional regulation of repair proteins to a plethora of DNA insults, including oxidative DNA damage. Rad53 activity is upregulated by phosphorylation, generating Rad53 species of various degrees of phosphorylation. I have measured steady state levels of Rad53 phosphorlyation by western blotting following SDS-polyacrylamide gel electrophoresis at different intervals in stationary phase, in various mutant backgrounds. To address the possible contribution of different repair pathways to endogenous DNA damage, I utilized two different sets of DNA repair deficient strains such as those deficient in Base excision repair (BER) and nucleotide excision repair (NER), and other set was deficient in Ku protein and NER. Interestingly, in both BERNER and Yku70rad4 strains, Rad53 phosphorylation was evident in stationary phase that is after 2 days, 4 days and 6 days but not in logarithmic phase. This covalent modification disappears after phosphatase treatment. This Rad53 modification was absent in their respective rho0 mutants, which lack mitochondrial DNA, indicating involvement of mitochondrial ROS in this checkpoint response. We analyzed mutants of different checkpoint proteins for Rad53 phosphorylation. Exclusive involvement of Rad17, Rad50 and Mec1 kinase in Rad53 phosphorylation strongly suggests processed DNA double strand breaks as critical lesions in BERNER cells. Analysis of Yku70 and NER deficient strain showed involvement of ssDNA, which is most likely at telomeres. This study consents with the model of unrepaired oxidative base damage, which can accelerate the appearance of single stranded DNA in the vicinity of double strand breaks (DSBs) or at telomeres.