Evidence for the covalent interaction of carbon tetrachloride with mouse liver chromatin DNA in vitro

Whole chromatin was isolated from livers of 8-12 week-old male B6C3Fl hybrid mice and incubated with 14C-labelled carbon tetrachloride (14CCL4) in the presence of hepatic microsomes from the same animals and an NADPH-regenerating system. The experiments were carried out at various concentrations of 14CCl4 and incubation times. Under the conditions used, 14CCl4 metabolite(s) bind quantifiably to chromatin DNA (ch.DNA) in direct response to increases in 14CCl4 concentration and incubation time. Maximal binding (0.34 nmoles/mg DNA) occurs at 2 hrs when 10 umoles 14CCl4 was used for the incubation. Binding of 14CCl4 metabolite(s) to ch.DNA was/were significantly inhibited (84%) in the presence of the strong nucleophile, L-cysteine (5 mM). These results provide evidence for the covalent interaction of CCl4 metabolite(s) with DNA when organized as chromatin under aerobic incubation condition, which suggest the importance of the higher-order organization of DNA in chromatin and its reactivity with CCl4 metabolite(s).