Effect of nickel(II) on DNA-protein interactions.
Alterations in DNA-protein interactions (DPI) may play an important role in carcinogenesis. Although the mechanism of nickel carcinogenesis is unknown, nickel reportedly affects DPI. A microfiltration, nitrocellulose filter assay was utilized to study DPI in intact Chinese hamster ovary (CHO) cells and in isolated nuclei. Prior to exposure of CHO cells or isolated CHO cell nuclei, DNA and proteins were radiolabeled using 3H-thymidine and 35S-methionine, respectively. Nuclei were exposed to NiCl2 in 10 mM HEPES buffer (pH 6.8). CHO cells were exposed in either complete or a salts-glucose medium. Following exposure, nuclei or cells were incubated at 37 degrees C for 20 min in a high salt lysis solution; aliquots were loaded onto nitrocellulose filters and washed with a low salt solution. DNA (3H) retained on each filter was normalized to protein (35S) bound on the filter. Exposure of either whole cells or isolated nuclei to increasing, noncytotoxic concentrations of NiCl2 resulted in a dose dependent decrease in DPI. The effect of nickel on specific DNA-protein interactions was examined using a band shift assay and a cloned satellite DNA sequence. Nickel inhibited specific protein binding to the satellite DNA probe. The results of these two independent assays, which were conducted at physiological pH, indicate that NiCl2 inhibits specific DNA-protein interactions.
Coogan
T P
TP
Institute of Environmental Medicine, New York University Medical Center, Tuxedo 10987.
Latta
D M
DM
Imbra
R J
RJ
Costa
M
M
D000818:Animals; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D002478:Cells, Cultured; D003101:Collodion; D006224:Cricetinae; D004247:DNA / Q000187:drug effects*; D005260:Female; D006863:Hydrogen-Ion Concentration; D008715:Methionine / Q000378:metabolism; D051379:Mice; D009532:Nickel / Q000494:pharmacology*; D010053:Ovary / Q000166:cytology; D011485:Protein Binding / Q000187:drug effects; D011506:Proteins / Q000187:drug effects*; D013462:Sulfur Radioisotopes
D011506:Proteins; D013462:Sulfur Radioisotopes; D009532:Nickel; D003101:Collodion; D004247:DNA; D008715:Methionine