Preferential DNA-protein cross-linking by NiCl2 in magnesium-insoluble regions of fractionated Chinese hamster ovary cell chromatin.
Intracellular nickel ions (Ni2+) have been shown to cause single-strand breaks in DNA, that were rapidly repaired, and DNA-protein cross-links, that persisted for at least 24 h following removal of extracellular ionic nickel. In this study, we have used the techniques of alkaline elution, chromatin fractionation, and sodium dodecyl sulfate:polyacrylamide gel electrophoresis to examine the DNA-protein cross-linking induced by NiCl2 in Chinese hamster ovary cells. Continuous treatment of logarithmically growing Chinese hamster ovary cells with 2.5 mM NiCl2 in complete medium resulted in DNA single-strand breaks within 1 h, followed by a time-dependent increase in the induction of DNA-protein cross-links at 2, 3, and 6 h. Since the entry of nickel into cells was maximal within 2 h of exposure, the time delay for the formation of DNA-protein cross-links was not limited by metal uptake. The nickel-induced DNA-protein cross-linking appeared to require active cell cycling, since single-strand breaks but no cross-linking could be detected in confluent cells treated with 1, 2.5, or 5 mM NiCl2 for 3 h. DNA-protein cross-linking induced by nickel occurred in late S phase of the cell cycle. High-molecular-weight nonhistone chromatin proteins and possibly histone H1 migrating at the Mr 30,000 range became cross-linked to DNA after treatment of cells with NiCl2. All nickel-cross-linked proteins were concentrated in the magnesium-insoluble regions of fractionated chromatin and were stable to urea, 2-mercaptoethanol, and Nonidet P-40. Some proteins (Mr 48,000, 52,000, 55,000, 70,000, and 95,000), the association of which with DNA was also stable to Sarkosyl, salt, and EDTA, were detectable in DNA rigorously fractionated from untreated cells. Nickel therefore appeared to cause the cross-linking of proteins that normally reside in close association with DNA. Alterations of the normal association of these proteins with DNA by nickel may be an early event in the nickel transformation process.
Patierno, SR; Sugiyama, M; Basilion, JP; Costa, M
Volume / Issue
Start / End Page
Pubmed Central ID
International Standard Serial Number (ISSN)