Rfc4 interacts with Rpa1 and is required for both DNA replication and DNA damage checkpoints in Saccharomyces cerevisiae.
The large subunit of replication protein A (Rpa1) consists of three single-stranded DNA binding domains and an N-terminal domain (Rpa1N) of unknown function. To determine the essential role of this domain we searched for mutations that require wild-type Rpa1N for viability in yeast. A mutation in RFC4, encoding a small subunit of replication factor C (RFC), was found to display allele-specific interactions with mutations in the gene encoding Rpa1 (RFA1). Mutations that map to Rpa1N and confer sensitivity to the DNA synthesis inhibitor hydroxyurea, such as rfa1-t11, are lethal in combination with rfc4-2. The rfc4-2 mutant itself is sensitive to hydroxyurea, and like rfc2 and rfc5 strains, it exhibits defects in the DNA replication block and intra-S checkpoints. RFC4 and the DNA damage checkpoint gene RAD24 were found to be epistatic with respect to DNA damage sensitivity. We show that the rfc4-2 mutant is defective in the G(1)/S DNA damage checkpoint response and that both the rfc4-2 and rfa1-t11 strains are defective in the G(2)/M DNA damage checkpoint. Thus, in addition to its essential role as part of the clamp loader in DNA replication, Rfc4 plays a role as a sensor in multiple DNA checkpoint pathways. Our results suggest that a physical interaction between Rfc4 and Rpa1N is required for both roles.
Kim
H S
HS
Department of Molecular Biology and Biochemistry, Center for Advanced Biotechnology and Medicine, Rutgers University, Piscataway, New Jersey 08854, USA.
Brill
S J
SJ
D018797:Cell Cycle Proteins / Q000378:metabolism; D004249:DNA Damage; D004261:DNA Replication; D004271:DNA, Fungal / Q000096:biosynthesis*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D018398:Homeodomain Proteins; D047908:Intracellular Signaling Peptides and Proteins; D015778:Minor Histocompatibility Antigens; D016296:Mutagenesis; D019253:Proto-Oncogene Proteins c-bcl-2; D051716:Replication Protein A; D051818:Replication Protein C; D012097:Repressor Proteins; D016196:S Phase; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D015398:Signal Transduction
C096816:BCL2-related protein A1; D018797:Cell Cycle Proteins; D004271:DNA, Fungal; D004268:DNA-Binding Proteins; D018398:Homeodomain Proteins; D047908:Intracellular Signaling Peptides and Proteins; C096063:MATA1 protein, S cerevisiae; D015778:Minor Histocompatibility Antigens; D019253:Proto-Oncogene Proteins c-bcl-2; C088385:RAD24 protein, S cerevisiae; D051716:Replication Protein A; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; D051818:Replication Protein C