A GENERAL PROTOCOL FOR EVALUATING THE SPECIFIC EFFECTS OF DNA-REPLICATION INHIBITORS

Inhibitors of DNA replication in mammalian cells are of great interest because of their potential use in chemotherapy and in cell synchroniz ing protocols in the laboratory. We have used a combination of isotopi c labelling protocols and a two-dimensional gel replicon mapping proce dure to determine the specific effects of five different replication i nhibitors in cultured cells. Utilizing this protocol, we show that hyd roxyurea, aphidicolin, and cytosine arabinoside, three known chain elo ngation inhibitors, are rather ineffective at preventing fork progress ion even at relatively high concentrations. In contrast, two related c ompounds that have been suggested to be G1/S inhibitors (mimosine and ciclopyrox olamine [CPX]) actually appear to inhibit initiation at ori gins. One of these agents (CPX) appears also to inhibit replication in yeast, opening the possibility that the gene encoding the target (ini tiator?) protein can first be identified in yeast by genetic approache s and can then be used to isolate the mammalian homologue.