MUTATIONAL EFFECT OF FISSION YEAST POL-ALPHA ON CELL-CYCLE EVENTS

Polar is the principal DNA polymerase for initiation of DNA replicatio n and also functions in postinitiation DNA synthesis. Ln this study, w e investigated the cell cycle responses induced by mutations in pol al pha(+). Germinating spores carrying either a deletion of pol alpha(+) (pol alpha Delta) or a structurally intact but catalytically dead pol alpha mutation proceed to inappropriate mitosis with no DNA synthesis. This suggests that the catalytic function, and not the physical prese nce of Pol alpha, is required to generate the signal that prevents the cells from entering mitosis prematurely. Cells with a pal alpha ts al lele arrest the cell cycle near the hydroxyurea arrest point, but, sur prisingly, pol alpha ts in cdc20 (pol epsilon mutant) background arres ted with a cdc phenoytpe, not a pol alpha ts-like phenotype. At 25 deg rees C, replication perturbation caused by pol alpha ts alleles induce s Cds1 kinase activity and requires the checkpoint Rads, Cds1, and Rqh 1, but not Chk1, to maintain cell viability. At 36 degrees C, replicat ion disruption caused by pol alpha ts alleles induces the phosphorylat ion of Chk1; however, mutant cells arrest with heterogeneous cell size s with a population of the cells entering aberrant mitosis. Together, our results indicate that the initiation DNA structure synthesized by Pol alpha is required to bring about the S phase to mitosis checkpoint , whereas replication defects of different severity caused by pol alph a ts mutations induce differential downstream kinase responses.