Abstract
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.