title: GEMC1 is a TopBP1-interacting protein required for chromosomal DNA replication.
abstract: Many of the factors required for chromosomal DNA replication have been identified in unicellular eukaryotes. However, DNA replication is poorly understood in multicellular organisms. Here, we report the identification of GEMC1 (geminin coiled-coil containing protein 1), a novel vertebrate protein required for chromosomal DNA replication. GEMC1 is highly conserved in vertebrates and is preferentially expressed in proliferating cells. Using Xenopus laevis egg extract we show that Xenopus GEMC1 (xGEMC1) binds to the checkpoint and replication factor TopBP1, which promotes binding of xGEMC1 to chromatin during pre-replication complex (pre-RC) formation. We demonstrate that xGEMC1 interacts directly with replication factors such as Cdc45 and the kinase Cdk2-CyclinE, through which it is heavily phosphorylated. Phosphorylated xGEMC1 stimulates initiation of DNA replication, whereas depletion of xGEMC1 prevents the onset of DNA replication owing to the impairment of Cdc45 loading onto chromatin. Similarly, inhibition of GEMC1 expression with morpholino and siRNA oligos prevents DNA replication in embryonic and somatic vertebrate cells. These data suggest that GEMC1 promotes initiation of chromosomal DNA replication in multicellular organisms by mediating TopBP1- and Cdk2-dependent recruitment of Cdc45 onto replication origins.
authors.affiliation: Genome Stability, London Research Institute, Hertfordshire, UK.
authors.forename: Alessia
authors.initials: A
authors.lastname: Balestrini
authors.forename: Claudia
authors.initials: C
authors.lastname: Cosentino
authors.forename: Alessia
authors.initials: A
authors.lastname: Errico
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Garner
authors.forename: Vincenzo
authors.initials: V
authors.lastname: Costanzo
mesh_terms: D000818:Animals; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism* / Q000502:physiology*; D018797:Cell Cycle Proteins / Q000378:metabolism; D002875:Chromosomes; D051357:Cyclin-Dependent Kinase 2 / Q000378:metabolism; D004261:DNA Replication; D004268:DNA-Binding Proteins; D008969:Molecular Sequence Data; D010063:Ovum; D011485:Protein Binding; D018741:Replication Origin / Q000235:genetics; D029867:Xenopus Proteins / Q000378:metabolism; D014982:Xenopus laevis
chemical_list: D002352:Carrier Proteins; C502211:Cdc45 protein, Xenopus; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; C551026:GMNC protein, Xenopus; C509993:TopBP1 protein, Xenopus; D029867:Xenopus Proteins; C495891:Cdk2 protein, Xenopus; D051357:Cyclin-Dependent Kinase 2
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