GINS, a novel multiprotein complex required for chromosomal DNA replication in budding yeast

Abstract

Eukaryotic chromosomal DNA replication requires a two-step assembly of replication proteins on origins; formation of the prereplicative complex (pre-RC) in late M and G1 phases of the cell cycle, and assembly of other replication proteins in S phase to load DNA polymerases to initiate DNA synthesis. In budding yeast, assembly of Dpb11 and the Sld3-Cdc45 complex on the pre-RC at origins is required for loading DNA polymerases. Here we describe a novel replication complex, GINS (Go, Ichi, Nii, and San; five, one, two, and three in Japanese), in budding yeast, consisting of Sld5, Psf1 (partner of Sld five 1), Psf2, and Psf3 proteins, all of which are highly conserved in eukaryotic cells. Since the conditional mutations of Sld5 and Psf1 confer defect of DNA replication under nonpermissive conditions, GINS is suggested to function for chromosomal DNA replication. Consistently, in S phase, GINS associates first with replication origins and then with neighboring sequences. Without GINS, neither Dpb11 nor Cdc45 associates properly with chromatin DNA. Conversely, without Dpb11 or Sld3, GINS does not associate with origins. Moreover, genetic and two-hybrid interactions suggest that GINS interacts with Sld3 and Dpb11. Therefore, Dpb11, Sld3, Cdc45, and GINS assemble in a mutually dependent manner on replication origins to initiate DNA synthesis.[Keywords: DNA replication; replication protein; Dpb11; Sld3; Sld5] Supplemental material is available at http://www.genesdev.org.

Mcm10 interacts with Rad4/Cut5TopBP1 and its association with origins of DNA replication is dependent on Rad4/Cut5TopBP1

“…Taken together with the data presented in Fig. 2 we favour the idea that Rad4 TopBP1 is not associated with 16 the moving replisome although we accept that the data remain inconclusive on this point due to the relatively low level of detection at the replication origins. A further intriguing possibility is that the low level of detection of Rad4 TopBP1 at replication origins, in contrast to increased accumulation at the stalled forks in HU, might reflect distinct activities of the protein and the dynamics of Rad4 TopBP1 turnover at the relevant sites within the genome.…”

“…Cdc45 loading 5 exhibits interdependence with Sld3, which in the light of the data discussed above, offers a compelling mechanism for combining the CDK and DDK activities in the initiation of DNA replication (3). Sld3 interacts with and is required for loading of the GINS complex in budding yeast (16). In turn the GINS complex has been reported to be necessary for Cdc45 S phase chromatin association suggestive of an intimate, conserved functional relationship between Sld3, Cdc45 and GINS (16,17).…”

“…Sld3 interacts with and is required for loading of the GINS complex in budding yeast (16). In turn the GINS complex has been reported to be necessary for Cdc45 S phase chromatin association suggestive of an intimate, conserved functional relationship between Sld3, Cdc45 and GINS (16,17). More recent evidence demonstrates that GINS is dispensable for Cdc45 origin loading but is required for its interaction with the active replisome whilst Sld3 is required for Cdc45 and GINS origin loading but does not move with the replisome (4).…”

Perpetuating the double helix: molecular machines at eukaryotic DNA replication origins

“…The protein complex between Sld5 and Psf1-3 has received the name of GINS (a Japanese acronym for five, one, two, three; Ref. 82). Mutant alleles sld5-12 and psf1-1 caused slow progression through S phase and loss of viability.…”

“…The association of GINS and Sld3-Cdc45 with the origins seems to be mutually dependent. (82) The Xenopus homologs of the GINS subunits also form a complex that associates with chromatin at the same time as Cdc45. (83) Depletion of XSld5 from the egg extracts impaired the replication of sperm chromatin.…”