Rfc5, in Cooperation with Rad24, Controls DNA Damage Checkpoints throughout the Cell Cycle in Saccharomyces cerevisiae

• Published in: Molecular and cellular biology 2000-08, Vol.20 (16), p.5888-5896
• Main Authors: Naiki, Takahiro, Shimomura, Toshiyasu, Kondo, Tae, Matsumoto, Kunihiro, Sugimoto, Katsunori
• Format: Article
• Language: English
• Subjects: Cell Cycle - genetics; Cell Cycle Proteins - genetics; DNA Dynamics and Chromosome Structure; DNA Replication - genetics; DNA-Binding Proteins - genetics; Gene Expression Regulation, Fungal; Homeodomain Proteins; Intracellular Signaling Peptides and Proteins; Minor Histocompatibility Antigens; Mutation; Proto-Oncogene Proteins c-bcl-2; RAD24 gene; Replication Protein C; Repressor Proteins; RFC5 gene; Saccharomyces cerevisiae; Saccharomyces cerevisiae - cytology; Saccharomyces cerevisiae - genetics; Saccharomyces cerevisiae Proteins
• ISSN: 0270-7306; 1098-5549; 1098-5549
• DOI: 10.1128/MCB.20.16.5888-5896.2000

RAD24 and RFC5 are required for DNA damage checkpoint control in the budding yeast Saccharomyces cerevisiae. Rad24 is structurally related to replication factor C (RFC) subunits and associates with RFC subunits Rfc2, Rfc3, Rfc4, and Rfc5. rad24Δ mutants are defective in all the G 1 -, S-, and G 2 /M-phase DNA damage checkpoints, whereas the rfc5-1 mutant is impaired only in the S-phase DNA damage checkpoint. Both the RFC subunits and Rad24 contain a consensus sequence for nucleoside triphosphate (NTP) binding. To determine whether the NTP-binding motif is important for Rad24 function, we mutated the conserved lysine 115 residue in this motif. The rad24-K115E mutation, which changes lysine to glutamate, confers a complete loss-of-function phenotype, while the rad24-K115R mutation, which changes lysine to arginine, shows no apparent phenotype. Although neitherrfc5-1 nor rad24-K115R single mutants are defective in the G 1 - and G 2 /M-phase DNA damage checkpoints, rfc5-1 rad24-K115R double mutants become defective in these checkpoints. Coimmunoprecipitation experiments revealed that Rad24 K115R fails to interact with the RFC proteins in rfc5-1 mutants. Together, these results indicate that RFC5, like RAD24, functions in all the G 1 -, S- and G 2 /M-phase DNA damage checkpoints and suggest that the interaction of Rad24 with the RFC proteins is essential for DNA damage checkpoint control.