Exposure of cells to ionizing radiation, UV light or chemicals can result in DNA damage, but usually repair mechanisms are available to mend the lesions induced. However, incomplete or faulty repair can occur and can have deleterious consequences, including cell lethality ,mutagenesis, teratogenesis or carcinogenesis. Therefore, an understanding of DNA repair mechanisms, a process fundamental to all living organisms, is important. To better understand the way cells respond to radiation or chemical exposure, this project focuses on molecular aspects of DNA repair primarily in the small eukaryote Schizosaccharomyces pombe. This yeast will serve as a model system because it affords several significant advantages over the use of mammalian cells in the laboratory, yet S. pombe shares many molecular, biological and biochemical features with mammalian systems. Recently, I isolated a S. pombe mutant exhibiting temperature-dependent sensitivity to UV light and ionizing radiation. Genetic analyses indicated that a previously unidentified, altered rad locus (i.e., called rad23-1) was responsible for the phenotype observed. In this proposal, I plan to characterize the mutant further, including the determination of chemical sensitivity, mutability and recombination ability. This mutant, as well as the S. pombe rad3-136 mutant, will be the focus of ability. This mutant, as well as the S. pombe rad3-136 mutant, will be the focus of molecular studies. No S. pombe DNA repair genes have been cloned to date. I propose to isolate the wild type rad23 and rad3 genes. The corresponding gene products encoded by these loci participate in repair of DNA damage induced by both UV light and ionizing radiation since alterations within these genes cause radiation sensitivity. Furthermore, it is known that the rad3 gene mediates UV light-induced mutagenesis. The structures of these cloned genes will be determined, and their inducibility by DNA damaging agents and/or preferential expression during a specific phase of the cell cycle will be tested to learn more about their regulation. The DNA sequence changes in the mutant rad genes, relative to wild type cognates, that make them responsible for radiation and chemical sensitivity will be identified. Limited studies will also be performed to determine whether S. pombe DNA repair gene sequences are conserved in other species. Together, these studies should provide important information relating to S. pombe DNA repair and possibly cognate systems in other organisms.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
First Independent Research Support & Transition (FIRST) Awards (R29)
Project #
5R29CA054044-05
Application #
2095654
Study Section
Radiation Study Section (RAD)
Project Start
1990-07-01
Project End
1995-04-30
Budget Start
1994-05-01
Budget End
1995-04-30
Support Year
5
Fiscal Year
1994
Total Cost
Indirect Cost
Name
Columbia University (N.Y.)
Department
Radiation-Diagnostic/Oncology
Type
Schools of Medicine
DUNS #
064931884
City
New York
State
NY
Country
United States
Zip Code
10027
Hang, H; Hager, D N; Goriparthi, L et al. (1996) Schizosaccharomyces pombe rad23 is allelic with swi10, a mating-type switching/radioresistance gene that shares sequence homology with human and mouse ERCC1. Gene 170:113-7
Zhao, Y; Lieberman, H B (1995) Schizosaccharomyces pombe: a model for molecular studies of eukaryotic genes. DNA Cell Biol 14:359-71
Lieberman, H B (1995) Extragenic suppressors of Schizosaccharomyces pombe rad9 mutations uncouple radioresistance and hydroxyurea sensitivity from cell cycle checkpoint control. Genetics 141:107-17
Lieberman, H B; Hopkins, K M (1994) Schizosaccharomyces malidevorans and Sz. octosporus homologues of Sz. pombe rad9, a gene that mediates radioresistance and cell-cycle progression. Gene 150:281-6
Zhao, Y; Goriparthi, L; Lieberman, H B (1994) A new shuttle vector system for the identification of spontaneous and radiation-induced mutations in the fission yeast Schizosaccharomyces pombe. Mutat Res 311:111-23
Zhao, Y; Hopkins, K M; Lieberman, H B (1993) A method for the preparation and storage of frozen, competent Schizosaccharomyces pombe spheroplasts. Biotechniques 15:238
Sidik, K; Lieberman, H B; Freyer, G A (1992) Repair of DNA damaged by UV light and ionizing radiation by cell-free extracts prepared from Schizosaccharomyces pombe. Proc Natl Acad Sci U S A 89:12112-6
Lieberman, H B; Hopkins, K M; Laverty, M et al. (1992) Molecular cloning and analysis of Schizosaccharomyces pombe rad9, a gene involved in DNA repair and mutagenesis. Mol Gen Genet 232:367-76