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.