Aminoimidazole carboxamide ribonucleoside (AIC-R), a purine precursor, has biphasic effects on the growth of Chinese hamster fibroblasts. At 200 microM AIC-R cell growth is almost completely arrested, while at 50 and 700 microM AIC-R cell growth is comparable to that observed in the absence of nucleoside. The growth inhibition produced by AIC-R is the consequence of inhibition of the orotate phosphoribosyltransferase-orotidylic decarboxylase (OPRT-ODC) reactions, as evidenced by a 87% reduction in the intracellular concentrations of UTP and CTP, accumulation of orotate in the medium, and restoration of normal growth by inclusion of 100 microM uridine in the medium. Inhibition of pyrimidine nucleotide synthesis at 200 microM AIC-R is associated with an 82% reduction in the intracellular concentration of PP-ribose-P and a 150% increase in the concentration of purine nucleotides. Restoration of cell growth to a normal rate at 700 microM AIC-R--a condition under which PP-ribose-P remains depressed and purine nucleotide concentrations are also depressed (40% of control)--and absence of toxicity at 50 microM AIC-R--a condition under which purine nucleotide concentrations are increased by 150% and PP-ribose-P concentration is normal--suggest that the inhibition of OPRT-ODC observed at 200 microM AIC-R is caused by the combination of the reduction in PP-ribose-P and increase in purine nucleotides. These studies provide a better understanding of the control of the OPRT-ODC reactions in the cell and provide additional insight into the basis of pyrimidine starvation induced by purine nucleosides.

Proteins cross-linked to DNA after nitrogen mustard (HN2) treatment of cells or isolated nuclei were purified in CsCl gradients. The protein-DNA cross-links could be cleaved by incubation in dilute acid and could be stabilized by alkali pretreatment. These results indicate that proteins cross-linked to DNA by HN2 are bound to alkylated purines. Analysis of the DNA-bound proteins on NaDodSO4-polyacrylamide gels showed that primarily large nonhistone proteins are cross-linked to DNA in cells treated with HN2. Very little if any histone is cross-linked to the DNA. Comparison of DNA bound proteins from HN2-treated cells and HN2-treated nuclei showed that in general the same proteins are linked to DNA in both cases, but some qualitative and quantitative differences exist.

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