The ultimate goal of this program is to construct an in vitro system composed of defined proteins to study the mechanisms employed by higher eukaryotes to contend with DNA damage. Drosophila will be employed in these studies, because a parallel genetic analysis will be performed to also define the in vivo function of the enzymes under investigation. Since the available mutants and those we expect to isolate are analogues of human repair-related disorders, these studies will provide evidence relevant to human DNA repair and its involvement in mutagenesis, carcinogenesis, and recombination. The isolation and characterization of several key enzymes of DNA metabolism will be undertaken. These enzymes include DNA polymerase Beta, DNA polymerase Gamma, two ATP-dependent deoxyribonucleases, a rec A-like protein, and a damage-specific endonuclease. Following their enzymological characterization, defects in these enzymes will be sought among the available repair-deficient mutants. Selected genes will be cloned either by screening a cDNA expression library or by screening genomic libraries with mixed-sequence oligonucleotide probes. The clones will be employed to study the normal regulation of these genes and to alter that regulation to define their role in DNA repair, recombination and synthesis. The clones will also be employed to guide a genetic study which will further define the in vivo function of the proteins. The recovered clones will provide probes which can potentially be employed to recover analogous genes from the human genome. Mitochondrial DNA metabolism will also be investigated in an effort to develop a model in vitro system.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
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Genetics Study Section (GEN)
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University of California Davis
Schools of Earth Sciences/Natur
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Banga, S S; Yamamoto, A H; Mason, J M et al. (1995) Molecular cloning of mei-41, a gene that influences both somatic and germline chromosome metabolism of Drosophila melanogaster. Mol Gen Genet 246:148-55
Harris, P V; Boyd, J B (1993) Re-evaluation of excision repair in the mus304, mus306 and mus308 mutants of Drosophila. Mutat Res 301:51-5
Leonhardt, E A; Henderson, D S; Rinehart, J E et al. (1993) Characterization of the mus308 gene in Drosophila melanogaster. Genetics 133:87-96
Leonhardt, E A; Boyd, J B (1993) Identification of a new locus, mus115, in Drosophila melanogaster. Mutat Res 301:121-4
Sakaguchi, K; Zdzienicka, M Z; Harris, P V et al. (1992) Nuclease modification in Chinese hamster cells hypersensitive to DNA cross-linking agents--a model for Fanconi anemia. Mutat Res 274:11-8
Harosh, I; Mezzina, M; Harris, P V et al. (1992) Purification and characterization of a mitochondrial endonuclease from Drosophila melanogaster embryos. Eur J Biochem 210:455-60
Harosh, I; Binninger, D M; Harris, P V et al. (1991) Mechanism of action of deoxyribonuclease II from human lymphoblasts. Eur J Biochem 202:479-84
Sakaguchi, K; Harris, P V; Ryan, C et al. (1991) Alteration of a nuclease in Fanconi anemia. Mutat Res 255:31-8
Banga, S S; Velazquez, A; Boyd, J B (1991) P transposition in Drosophila provides a new tool for analyzing postreplication repair and double-strand break repair. Mutat Res 255:79-88
Oliveri, D R; Harris, P V; Boyd, J B (1990) X-ray sensitivity and single-strand DNA break repair in mutagen-sensitive mutants of Drosophila melanogaster. Mutat Res 235:25-31

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