DETERMINATION OF INITIATION OF DNA-REPLICATION BEFORE AND AFTER NUCLEAR FORMATION IN XENOPUS EGG CELL-FREE-EXTRACTS

Xenopus egg extracts prepared before and after egg activation retain M - and S-phase specific activity, respectively. Staurosporine, a potent inhibitor of protein kinase, converted M-phase extracts into interpha se-like extracts that were capable of forming nuclei upon the addition of sperm DNA. The nuclei formed in the staurosporine treated M-phase extract were incapable of replicating DNA, and they were unable to ini tiate replication upon the addition of S-phase extracts. Furthermore, replication was inhibited when the staurosporine-treated M-phase extra ct was added in excess to the staurosporine-treated S-phase extract be fore the addition of DNA. The membrane-depleted S-phase extract suppor ted neither nuclear formation nor replication; however, preincubation of sperm DNA with these extracts allowed them to form replication-comp etent nuclei upon the addition of excess staurosporine-treated M-phase extract. These results demonstrate that positive factors in the S-pha se extracts determined the initiation of DNA replication before nuclea r formation, although these factors were unable to initiate replicatio n after nuclear formation.