Our hypothesis is that defects in the enzyme families we study result in aberrant base excision and homology-directed repair which is an engine driving human carcinogenesis. The majority of endogenous and radiation-induced DNA lesions are removed by BER and when this pathway fails, the resulting substrates are channeled into HDR. The overall goals of this Program Project are to understand at the atomic level how three families of DNA repair enzymes, the HhH and Fpg/Nei families of DNA glycosylases and the RecA-RAD51 family of recombinases recognize and process their substrates and how germ line and tumor associated variants of these proteins influence cancer susceptibility and carcinogenesis, respectively. Based on our strengths in biochemistry and structural biology, our program is now informed and driven by the identification and characterization of germ line and tumor-associated variants of human base excision repair and HDR enzymes. Core A will identify human germ line and somatic DNA sequence variants of the oxidative DNA glycosylases and RAD51 based on structure and phylogeny. Project 1 will demonstrate whether these repair variants induce cellular transformation, are mutagenic in mouse cells and whether they influence the cellular response to ionizing radiation and chemotherapeutic agents. Project 2 will examine the biochemical properties of the oxidative glycosylase variants and solve structures of wild type enzymes with substrates and where appropriate the glycosylase variants. Project 3 will examine the biochemical and where appropriate, structural characteristics of RAD51 variants as well as study the mechanisms of RAD51 filament formation. Project 4 will reconstitute the base excision repair pathway in the context of nucleosomes with wild type and variant glycosylases and examine the effect of histone primary sequence variants on chromatin accessibility during BER. Project 5 will define at the single molecule level the search mechanisms of the BER and HDR enzymes and their variants. Projects 1-5 will be serviced by Core B which will supply purified proteins and perform high throughput analysis of the proteins. Core A will perform bioinformatics analysis for all projects and kinetics analysis for Projects 2-4

Public Health Relevance

These studies will provide insight into how the BER and RAD51 variants in the normal population and in tumors may contribute to altered DNA repair capacity. Moreover, this aberrant DNA repair in tumor cells may be a key cause of genetic instability in cancer that leads to tumor progression. Our approach may also offer the possibility of devising therapeutic strategies to which tumors that express DNA glycosylase and RAD51 variants are susceptible.

National Institute of Health (NIH)
National Cancer Institute (NCI)
Research Program Projects (P01)
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Special Emphasis Panel (ZCA1-RPRB-0 (O1))
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Pelroy, Richard
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University of Vermont & St Agric College
Schools of Medicine
United States
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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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