Every day, normal cellular metabolism produces 30,000 or more oxidative lesions In the DNA of each human cell. Most of these lesions are removed by the base excision repair (BER) machinery. Certain defective BER enzymes have been shown to be oncogenic, Nucleosomes that package DNA into chromafin provide little if any protection from oxidative damage. Project 4 was added to the original POI in September, 2007, to identify rate-limiting steps in the BER of lesions in nucleosomes, and to investigate mechanisms that cells use to circumvent these limits. Studies to date indicate that the human bifunctional DNA glycosylase hNTHI can excise oxidative lesions from nucleosomal DNA without irreversibly disrupting the nucleosome, and that spontaneous, transient partial unwrapping of DNA from the histone octamer facilitates access to sterically occluded lesions. With these results as a foundation, the goals for this new proposal are to test HYPOTHESIS 1, that during BER of lesions in nucleosomes, there is a stepwise hand-off of substrates from one enzyme to the next, facilitated by BER accessory factors. In the course of testing this hypothesis, we will investigate changes in nucleosome structure that accompany individual steps in BER, These structural analyses will help guide future studies of histone chaperones and chromatin remodeling agents that facilitate BER in vivo. We also will test HYPOTHESIS 2, that histone primary sequence variants associated with either transcriptionally active or inactive chromatin influence the capacity of selected human DNA glycosylases to repair oxidative damages in nucleosomes. Information from Project 2 will help elucidate the molecular basis for interactions between hNTHI and lesion-containing nucleosomes. Information and reagents provided by Core A and Projects 1 and 3 will enable us to test HYPOTHESIS 3, that DNA polymerase p (Pol P) variants with demonstrated oncogenic potential differ from wild type Pol p in BER of lesions in nucleosomes. We also will determine If, in the context of nucleosomal templates, Pol p (or oncogenic variants of Pol P) can bind or recruit hRadSI. Core B will provide critical reagents for all of the studies in Project 4.
The information generated by the proposed studies will advance our understanding of the carcinogic consequences of damages arising from endogenous metabolism and radiafion exposure. Additionally, radiation threatment is widely used in cancer therapy. The proposed studies will advance our understanding of factors that affect radiafion sensitivity of cells and modulation of such factors could help maximize therapeutic gain during radiotherapy.
|Maher, R L; Marsden, C G; Averill, A M et al. (2017) Human cells contain a factor that facilitates the DNA glycosylase-mediated excision of oxidized bases from occluded sites in nucleosomes. DNA Repair (Amst) 57:91-97|
|Marsden, Carolyn G; Dragon, Julie A; Wallace, Susan S et al. (2017) Base Excision Repair Variants in Cancer. Methods Enzymol 591:119-157|
|Galick, Heather A; Marsden, Carolyn G; Kathe, Scott et al. (2017) The NEIL1 G83D germline DNA glycosylase variant induces genomic instability and cellular transformation. Oncotarget 8:85883-85895|
|Robey-Bond, Susan M; Benson, Meredith A; Barrantes-Reynolds, Ramiro et al. (2017) Probing the activity of NTHL1 orthologs by targeting conserved amino acid residues. DNA Repair (Amst) 53:43-51|
|Cannan, Wendy J; Rashid, Ishtiaque; Tomkinson, Alan E et al. (2017) The Human Ligase III?-XRCC1 Protein Complex Performs DNA Nick Repair after Transient Unwrapping of Nucleosomal DNA. J Biol Chem 292:5227-5238|
|Silva, Michelle C; Bryan, Katie E; Morrical, Milagros D et al. (2017) Defects in recombination activity caused by somatic and germline mutations in the multimerization/BRCA2 binding region of human RAD51 protein. DNA Repair (Amst) 60:64-76|
|Zhou, Jia; Chan, Jany; Lambelé, Marie et al. (2017) NEIL3 Repairs Telomere Damage during S Phase to Secure Chromosome Segregation at Mitosis. Cell Rep 20:2044-2056|
|Prakash, Aishwarya; Moharana, Kedar; Wallace, Susan S et al. (2017) Destabilization of the PCNA trimer mediated by its interaction with the NEIL1 DNA glycosylase. Nucleic Acids Res 45:2897-2909|
|Lee, Andrea J; Wallace, Susan S (2017) Hide and seek: How do DNA glycosylases locate oxidatively damaged DNA bases amidst a sea of undamaged bases? Free Radic Biol Med 107:170-178|
|Lee, Andrea J; Wallace, Susan S (2016) Visualizing the Search for Radiation-damaged DNA Bases in Real Time. Radiat Phys Chem Oxf Engl 1993 128:126-133|
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