Abstract

The synthesis of purine nucleotides de novo in sarcoma 180 ascites cells propagated in mice was essentially completely inhibited for a prolonged period of time by azaserine (O-diazoacetyl-L-serine); this metabolic blockade resulted in a 60% decrease in the amount of adenine nucleotides in time cells. Concurrently, in the antibiotic-treated cells, the incorporation of thymidine-³H, formate-¹⁴C and deoxycytidine-³H into the thymine of DNA was markedly depressed, while the utilization of orotic acid-¹⁴C and leucine-¹⁴C for the formation of RNA and proteins, respectively, was unaffected. Azaserine did not decrease the rate of incorporation of adenine-¹⁴C into the purine nucleotides of DNA and RNA, suggesting that the polymerization processes per se were not altered by the drug. The inhibition of the incorporation of labeled precursors into the thymine of DNA in azaserine-treated cells was accounted for by a pronounced depression in the levels of thymidine kinase (ATP:thymidine 5'-phosphotransferase, EC 2.7.1.21) and thymidine monophosphate kinase (ATP:thymidine monophosphate phosphotransferase, EC 2.7.4.9). The availability to the cells of a supply of preformed adenine restored both the azaserinedepleted pool of adenine nucleotides and the activities of the thymine nucleotide-synthesizing enzymes; in this capacity, hypoxanthine was only slightly effective, while guanosine and azapurines were ineffective in reversing the azaserine-induced metabolic lesions. The results suggested a possible role for adenine nucleotides in regulating the activities of thymidine kinase and thymidine monophosphate kinase in intact cells.