Sld2, Which Interacts with Dpb11 in Saccharomyces cerevisiae, Is Required for Chromosomal DNA Replication

TheDPB11 gene, which genetically interacts with DNA polymerase II (ɛ), one of three replicative DNA polymerases, is required for DNA replication and the S phase checkpoint inSaccharomyces cerevisiae . To identify factors interacting with Dbp11, we have isolatedsld (synthetically lethal withdpb11-1 ) mutations which fall into six complementation groups (sld1 to -6 ). In this study, we characterizedSLD2 , encoding an essential 52-kDa protein. High-copySLD2 suppressed the thermosensitive growth defect caused bydpb11-1 . Conversely, high-copyDPB11 suppressed the temperature-sensitive growth defect caused bysld2-6 . The interaction between Sld2 and Dpb11 was detected in a two-hybrid assay. This interaction was evident at 25°C but not at 34°C when Sld2-6 or Dpb11-1 replaced its wild-type protein. No interaction between Sld2-6 and Dpb11-1 could be detected even at 25°C. Immunoprecipitation experiments confirmed that Dpb11 physically interacts with Sld2.sld2-6 cells were defective in DNA replication at the restrictive temperature, as weredpb11-1 cells. Further, indpb11-1 andsld2-6 cells, the bubble-shaped replication intermediates formed in the region of the autonomously replicating sequence reduced quickly after a temperature shift. These results strongly suggest the involvement of the Dpb11-Sld2 complex in a step close to the initiation of DNA replication.