Reduced dosage of a single fission yeast MCM protein causes genetic instability and S phase delay

MCM proteins are a conserved family of eukaryotic replication factors impli cated in the initiation of DNA replication and in the discrimination betwee n replicated and unreplicated chromatin, However, most mcm mutants in yeast arrest the cell cycle after bulk DNA synthesis has occurred. We investigat ed the basis for this late S phase arrest by analyzing the effects of a tem perature-sensitive mutation in fission yeast cdc19(+) (mcm2(+)). cdc19-P1 c ells show a dramatic loss of viability at the restrictive temperature, whic h is not typical of all S phase mutants. The cdc19-P1 cell cycle arrest req uires an intact damage-response checkpoint and is accompanied by increased rates of chromosome loss and mitotic recombination. Chromosomes from cdc19- P1 cells migrate aberrantly in pulsed-field gels, typical of strains arrest ed with unresolved replication intermediates. The cdc19-P1 mutation reduces the level of the Cdc19 protein at all temperatures. We compared the effect s of disruptions of cdc19(+) (mcm2(+)), cdc21(+) (mcm4(+)), nda4(+) (mcm5()) and mis5(+) (mcm6(+)); in all cases, the null mutants underwent delayed S phase but were unable to proceed through the cell cycle. Examination of p rotein levels suggests that this delayed S phase reflects limiting, but not absent, MCM proteins. Thus, reduced dosage of MCM proteins allows replicat ion initiation, but is insufficient for completion of S phase and cell cycl e progression.