Enhanced release of nascent single strands from DNA synthesized in the presence of arabinosylcytosine

Abstract

Arabinosylcytosine at a 1 . 10(-4) molar concentration inhibited thymidine incorporation into DNA by more than 95%. In sucrose gradients the labelled dThd was predominantly found in short DNA chains. Labelled arabinosylcytosince (aC) was incorporated into DNA, as was labelled dThd, indicating that it causes a preferential inhibition of the chain polymerization rather than termination of nascent chains. This was confirmed by the observation of the conversion of short chains to normal size DNA molecules in chase experiments. In the absence of NaCl and at neutral pH a release of more than 50% of the nascent label as single strands was repeatedly observed upon sucrose gradient centrifugation. This release could be significantly reduced by 1 M NaCl, indicating that in the presence of aC the in vivo structure was preserved. This was further confirmed by the fact that aC did not cause detachment of DNA from a rapidly sedimenting nuclear structure. Based on these results, a replication model slightly different from the one suggested by Okazaki, is proposed, in which initiations of new nascent chains can occur at certain distances ahead of the replication fork.