The goal of this project is to explain how ionizing radiation causes cancer, mutation and cell death by elucidating the chemistry and enzymology of X-ray-induced DNA damage and repair in human cells. New approaches to the study of X-ray-induced single-strand breaks and base-modifications are described. We will use a repeated sequence (alpha sequence) in human DNA to 1) determine the location (sequence) of strand breaks after gamma-irradiation in vitro and in vivo; 2) determine the effect of oxygenation, pH, temperature, and free radical scavengers on the location and extent of strand breaks; 3) determine the location (sequence) of alkali-labile sites in gamma-irradiated DNA; 4) determine the level of production of a newly described X-ray lesion, 3' phosphoglycolate termini, in DNA irradiated in vitro and in intact cells. The approach to study of X-ray induced base modifications uses a sensitive """"""""post-modification labeling"""""""" method. This method will be used to determine 1) levels of formation of particular base-modifications after gamma-irradiation and 2) the rates and extent of repair of such base-modifications after gamma irradiation. The approach chosen to study human repair enzymes uses DNA substrates which contain single, defined and radiolabeled lesions of the types induced by X-rays. We will 1) determine the presence of human enzymes specific for DNA containing thymine glycols, HMU and 3' phosphoglycolate termini; 2) utilize such substrates for isolation and characterization of such enzymes; 3) determine whether a deficiency of such an enzyme can explain the susceptibility of ataxia telangiectasia cells to X-rays and radiomimetic drugs. Knowledge of the processes of X-ray DNA damage and repair will be useful in developing more rational approaches to cancer prevention and therapy and in predicting the effects of low level radiation exposure.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA035767-02
Application #
3173347
Study Section
Radiation Study Section (RAD)
Project Start
1984-07-01
Project End
1987-06-30
Budget Start
1985-07-01
Budget End
1986-06-30
Support Year
2
Fiscal Year
1985
Total Cost
Indirect Cost
Name
Dana-Farber Cancer Institute
Department
Type
DUNS #
149617367
City
Boston
State
MA
Country
United States
Zip Code
Zhao, B; Grandy, D K; Hagerup, J M et al. (1992) The human gene for apurinic/apyrimidinic endonuclease (HAP1): sequence and localization to chromosome 14 band q12. Nucleic Acids Res 20:4097-8
Ward, B E; Burkett, B; Petersen, C et al. (1990) Cytologic correlates of cervical papillomavirus infection. Int J Gynecol Pathol 9:297-305
Sanderson, B J; Chang, C N; Grollman, A P et al. (1989) Mechanism of DNA cleavage and substrate recognition by a bovine apurinic endonuclease. Biochemistry 28:3894-901
Jorgensen, T J; Furlong, E A; Henner, W D (1988) Gamma endonuclease of Micrococcus luteus: action on irradiated DNA. Radiat Res 114:556-66
Jorgensen, T J; Kow, Y W; Wallace, S S et al. (1987) Mechanism of action of Micrococcus luteus gamma-endonuclease. Biochemistry 26:6436-43
Doetsch, P W; Henner, W D; Cunningham, R P et al. (1987) A highly conserved endonuclease activity present in Escherichia coli, bovine, and human cells recognizes oxidative DNA damage at sites of pyrimidines. Mol Cell Biol 7:26-32
Henner, W D; Kiker, N P; Jorgensen, T J et al. (1987) Purification and amino-terminal amino acid sequence of an apurinic/apyrimidinic endonuclease from calf thymus. Nucleic Acids Res 15:5529-44
Furlong, E A; Jorgensen, T J; Henner, W D (1986) Production of dihydrothymidine stereoisomers in DNA by gamma-irradiation. Biochemistry 25:4344-9
Janicek, M F; Haseltine, W A; Henner, W D (1985) Malondialdehyde precursors in gamma-irradiated DNA, deoxynucleotides and deoxynucleosides. Nucleic Acids Res 13:9011-29