Chromium-induced cross-linking of nuclear proteins and DNA

The in vivo cross-linking of proteins to DNA in intact Novikoff ascites hepatoma cells exposed to the chromium salt K2CrO4 was studied. DNA-protein complexes were assayed by high speed centrifugation of cells solubilized in buffered 4% sodium dodecyl sulfate and by electrophoretic identification of proteins associated with DNA-containing pellets. Further evidence of DNA-protein complexes, not dissociable in this buffer, was obtained by CsCl gradient centrifugation. Time dependence experiments showed that detectable cross-linking occurred after cells were exposed to chromium salt for at least 4 h, and the amount of DNA-protein complexes increased with longer incubation times. Complex formation occurred only with chromium salt concentrations of 200 microM or greater, and maximal cross-linking was effected at 5 mM. Immunotransfer methodology employing antibodies to nuclear matrix fraction and lamins was used to identify some of the polypeptides comprising the cross-linked complexes. These studies indicated specificity of chromium-induced complex formation within the nuclear protein fractions assayed. Our results document the ability of chromate to produce specific DNA-protein cross-links in living cells.