Appearance of rereplication and activation of DNA replication by calyculin A in a cell-free extract of activated Xenopus eggs.

When sperm nuclei were added to Xenopus S phase (activated) egg extracts pretreated with a serine/threonine protein phosphatase inhibitor, calyculin A, a density substitution experiment showed that newly synthesized sperm DNA migrated in two peaks, heavy-light DNA and heavy-heavy DNA, only in the presence of calyculin A and that the total DNA replication activity was activated. In contrast, the addition of calyculin A to S phase extracts about 30 min after addition of sperm nuclei had no effect on DNA replication activity. Calyculin A clearly prevented the decline of Replication Licensing Factor activity during S phase. These results imply that the occurrence of some rereplication and the activation of DNA replication by calyculin A in activated extracts may be due to the inhibition of inactivation of licensed sites of initiation or the increase in the number of the initiation sites of DNA replication because of lack of the fall in Licensing Factor activity during S phase.