Phosphorylation of Mcm2 protein by Cdc7 kinase is not essential for the initiation of DNA replication in fission yeast

Previous studies in many species suggested that the minichromosome maintenance complex is a major physiological target of Cdc7. In this study, we examined the cellular role of Mcm2 phosphorylation by Cdc7 kinase in Schizosaccharomyces pombe. The in vitro phosphorylation of several truncated Mcm2 proteins by Cdc7 kinase showed that the major phosphorylation sites were located at N-terminus of Mcm2 protein. Alanine substitutions of several putative Cdc7 phosphorylation sites in this region resulted in the mutant Mcm2 proteins which were hardly phosphorylated by Cdc7 kinase in vitro. When these proteins were expressed in mcm2 mutant cells, all of alanine substituted mutant proteins still complemented the temperature sensitive phenotype of mutant cells. These results suggest that the phosphorylation of Mcm2 protein by Cdc7 kinase is not essential for the initiation of DNA replication. Cdc7 may play its essential roles by phosphorylating other Mcm subunits or proteins involved in the initiation of DNA replication.