Early effects of protein kinase modulators on DNA synthesis in rat cerebral cortex

By using tissue miniunits, protein kinase modulators, and topoisomerase inh ibitors in short-term incubation (0-90 min) we studied (1) the role of prot ein phosphorylation in the immediate control of DNA replication in the deve loping rat cerebral cortex and (2) the mechanism of action for genistein-me diated DNA synthesis inhibition. Genistein decreased the DNA synthesis with in less than 30 min. None of the other protein kinase inhibitors examined ( herbimycin A, staurosporine, calphostin-C) or the protein phosphatase inhib itor sodium orthovanadate inhibited DNA synthesis and they did not affect t he genistein-mediated inhibition. The selective topoisomerase inhibitors ca mptothecin and etoposide decreased the DNA synthesis to an extent similar t o that of genistein and within less than 30 min. In addition, the effects o f these substances on topoisomerase I and II were studied. Etoposide and ge nistein but not herbimycin A, staurosporine, or calphostin-C strongly inhib ited the activity of topoisomerase II. Our results (1) strongly suggest tha t the net rate of DNA replication during the S phase of the cell cycle is i ndependent of protein phosphorylation and (2) indicate that the early inhib itory effect of genistein on DNA synthesis is mediated by topoisomerase II inhibition rather than protein tyrosine kinase inhibition, (C) 1999 Academi c Press.