Effect of 1-beta-D-arabinofuranosylcytosine triphosphate on DNA synthesis in isolated HeLa cell nuclei

Abstract

1-beta-D-Arabinofuranosylcytosine 5'-triphosphate (ara-CTP) was found to be a competitive inhibitor of DNA synthesis in nuclei isolated from synchronized S-phase HeLa S3 cells. The inhibition was reversed by dCTP. The main inhibitory effect was to reduce the rate of elongation of the primary DNA pieces from ara-CTP-treated nuclei after a 60-s pulse was about 115-120 nucleotides vs. 215-220 in control nuclei. A reduction in the frequency of initiation of primary DNA pieces probably also occurred. A slight inhibition of the transfer of labeled material from the primary pieces into high-molecular weight DNA was noted and probably arose from the fact that the reduced rate of growth of the primary pieces caused the critical size for ligation to be reached later. Synchronized HeLa S3 cells in suspension were treated with 1-beta-D-arabinofuranosylcytosine for 2 h in S-phase to give a 99% inhibition of DNA synthesis prior to isolation of nuclei. Neither the rate nor the extent of [3H]TTP incorporation into DNA decreased when such nuclei were incubated in the absence of ara-CTP, but the rate of ligation of primary pieces was reduced.