THE LARGE SUBUNIT OF REPLICATION FACTOR-C (RFC1P CDC44P) IS REQUIRED FOR DNA-REPLICATION AND DNA-REPAIR IN SACCHAROMYCES-CEREVISIAE/

We used genetic and biochemical techniques to characterize the phenoty pes associated with mutations affecting the large subunit of replicati on factor C (Cdc44p or Rfc1p) in Saccharomyces cerevisiae. We demonstr ate that Cdc44p is required for both DNA replication and DNA repair in vivo. Cold-sensitive cdc44 mutants experience a delay in traversing S phase at the restrictive temperature following alpha factor arrest; a lthough mutant cells eventually accumulate with a G2/M DNA content, th ey undergo a cell cycle arrest and initiate neither mitosis nor a new round of DNA synthesis. cdc44 mutants also exhibit an elevated level o f spontaneous mutation, and they are sensitive both to the DNA damagin g agent methylmethane sulfonate and to exposure to UV radiation. After exposure to UV radiation, cdc44 mutants at the restrictive temperatur e contain higher levels of single-stranded DNA breaks than do wild-typ e cells. This observation is consistent with the hypothesis that Cdc44 p is involved in repairing gaps in the DNA after the excision of damag ed bases. Thus, Cdc44p plays an important role in both DNA replication and DNA repair in vivo.