Unphosphorylatable mutants of Cdc6 disrupt its nuclear export but still support DNA replication once per cell cycle

Cdc6 is essential for eukaryotic DNA replication. We have mutated highly conserved CDK phosphorylation sites in Cdc6. Contrary to their reported phenotypes in human cells, unphosphorylatable DeltaCDK mutants fully support DNA replication in Xenopus eggs. WtCdc6 is actively exported from the nucleus, which could explain why nuclear permeabilization is required for reinitiation within one cell cycle. However, DeltaCDK mutants are retained in the nucleus, yet surprisingly they still support only one round of replication. As these highly conserved CDK sites are unnecessary for replication once per cell cycle, an alternative checkpoint role for monitoring completion of the S phase is suggested.