Relationship between Inhibition of Polyamine Biosynthesis and DNA Replication in Activated Lymphocytes¹

This paper examines the relationship between inhibition of DNA replication and reduced polyamine levels in bovine lymphocytes using several different inhibitors of polyamine biosynthesis and various administration regimens. Dose-response curves for inhibition of DNA synthesis by methylglyoxal bis(guanylhydrazone) failed to provide evidence for an intracellular inhibitory site of this drug aside from polyamine biosynthesis. Experiments with two analogs of methylglyoxal bis(guanylhydrazone) showed a correlation between inhibition of DNA replication and lowered levels of spermidine and spermine. A new inhibitor of polyamine biosynthesis, α-difluoromethylornithine, which irreversibly inhibits ornithine decarboxylase and is structurally unrelated to methylglyoxal bis(guanylhydrazone), specifically inhibited DNA replication without altering the number of cells in S phase or the rates of protein and RNA synthesis. This physiological response was identical to that observed previously when polyamine synthesis was blocked with methylglyoxal bis(guanylhydrazone). We conclude that the observed inhibition of [³H]thymidine incorporation was due to depressed intracellular polyamine levels and not to other pharmacological effects of the inhibitors that were unrelated to inhibition of polyamine biosynthesis.

In the polyamine-deficient cells, inhibition was also observed when [³H]thymidine incorporation was measured under conditions where de novo deoxythymidine 5′-phosphate synthesis was inhibited by amethopterin. It therefore appears that the observed inhibition was due to a depressed rate of DNA replication rather than to altered specific radioactivity of the intracellular DNA precursor, deoxythymidine 5′-triphosphate. Under the polyamine-deficient conditions, a lowered intracellular pool of deoxythymidine 5′-triphosphate was observed. However, it is concluded that this could not bring about the defects in replication observed and that polyamine deficiency probably induces defects in the nuclear replication machinery per se.