The DNA-protein crosslink lesion caused by carcinogenic nickel and chromium compounds will be studied. The formation and repair of this lesion will be examined by the alkaline elution technique. Individual proteins crosslinked to the DNA by nickel and chromium will be studied in intact cells and in vitro. To study the proteins crosslinked in intact cultured mammalian cells, DNA will be isolated from cells treated with metal compounds, and proteins intrinsically labeled with 35S methionine that cannot be dissociated from the DNA with high salt and 1% SDS will be analyzed by SDS polyacrylamide gel electrophoresis. The crosslinking reaction occurring in the intact cell will be modeled in vitro by reacting purified or crude nuclear protein fractions with metal and DNA. Order of addition and metal binding studies conducted with this in vitro system will facilitate an understanding of the reaction sequence occurring in the intact cell. The nature of the crosslink reaction will be examined in some detail. The amino acid and the DNA base involved in the metal-bridged crosslinks will be studied. The significance of the nickel or chromium induced DNA-protein lesion will also be examined. DNA containing proteins crosslinked in vitro or in the intact cell will be transfected into NIH 3T3 cells to assess the contribution of this lesion towards the development of transformation. Proteins crosslinked to DNA by nickel or chromium may protect certain DNA sequences from degradation with nucleases. These protected sequences will be examined for enrichment in DNA homologous to specific genomic probes of interest (i.e. oncogenes). The effect of DNA protein crosslinks on RNA and DNA synthesis will also be examined to provide a more complete understanding of the early effects of this lesion on these important cellular processes.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA043070-02
Application #
3184968
Study Section
Chemical Pathology Study Section (CPA)
Project Start
1986-01-01
Project End
1989-04-30
Budget Start
1986-05-01
Budget End
1987-04-30
Support Year
2
Fiscal Year
1986
Total Cost
Indirect Cost
Name
New York University
Department
Type
Schools of Medicine
DUNS #
004514360
City
New York
State
NY
Country
United States
Zip Code
10012
Miller 3rd, C A; Costa, M (1990) Immunodetection of DNA-protein crosslinks by slot blotting. Mutat Res 234:97-106
Costa, M (1989) Perspectives on the mechanism of nickel carcinogenesis gained from models of in vitro carcinogenesis. Environ Health Perspect 81:73-6
Conway, K; Costa, M (1989) The involvement of heterochromatic damage in nickel-induced transformation. Biol Trace Elem Res 21:437-44
Conway, K; Costa, M (1989) Nonrandom chromosomal alterations in nickel-transformed Chinese hamster embryo cells. Cancer Res 49:6032-8
Miller 3rd, C A; Costa, M (1989) Immunological detection of DNA-protein complexes induced by chromate. Carcinogenesis 10:667-72
Miller 3rd, C A; Costa, M (1989) Analysis of proteins cross-linked to DNA after treatment of cells with formaldehyde, chromate, and cis-diamminedichloroplatinum(II). Mol Toxicol 2:11-26
Wang, X W; Imbra, R J; Costa, M (1988) Characterization of mouse cell lines resistant to nickel(II) ions. Cancer Res 48:6850-4
Zelikoff, J T; Li, J H; Hartwig, A et al. (1988) Genetic toxicology of lead compounds. Carcinogenesis 9:1727-32
Miller 3rd, C A; Costa, M (1988) Characterization of DNA-protein complexes induced in intact cells by the carcinogen chromate. Mol Carcinog 1:125-33
Sugiyama, M; Costa, M; Nakagawa, T et al. (1988) Stimulation of polyadenosine diphosphoribose synthesis by DNA lesions induced by sodium chromate in Chinese hamster V-79 cells. Cancer Res 48:1100-4

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