title: CDC28 phosphorylates Cac1p and regulates the association of chromatin assembly factor I with chromatin.
abstract: Chromatin Assembly Factor I (CAF-I) plays a key role in the replication-coupled assembly of nucleosomes. It is expected that its function is linked to the regulation of the cell cycle, but little detail is available. Current models suggest that CAF-I is recruited to replication forks and to chromatin via an interaction between its Cac1p subunit and the replication sliding clamp, PCNA, and that this interaction is stimulated by the kinase CDC7. Here we show that another kinase, CDC28, phosphorylates Cac1p on serines 94 and 515 in early S phase and regulates its association with chromatin, but not its association with PCNA. Mutations in the Cac1p-phosphorylation sites of CDC28 but not of CDC7 substantially reduce the in vivo phosphorylation of Cac1p. However, mutations in the putative CDC7 target sites on Cac1p reduce its stability. The association of CAF-I with chromatin is impaired in a cdc28-1 mutant and to a lesser extent in a cdc7-1 mutant. In addition, mutations in the Cac1p-phosphorylation sites by both CDC28 and CDC7 reduce gene silencing at the telomeres. We propose that this phosphorylation represents a regulatory step in the recruitment of CAF-I to chromatin in early S phase that is distinct from the association of CAF-I with PCNA. Hence, we implicate CDC28 in the regulation of chromatin reassembly during DNA replication. These findings provide novel mechanistic insights on the links between cell-cycle regulation, DNA replication and chromatin reassembly.
authors.affiliation: a Department of Molecular and Cellular Biology ; University of Guelph ; Guelph , Ontario , Canada.
authors.forename: Daniel C B
authors.initials: DC
authors.lastname: Jeffery
authors.forename: Naoko
authors.initials: N
authors.lastname: Kakusho
authors.forename: Zhiying
authors.initials: Z
authors.lastname: You
authors.forename: Marlene
authors.initials: M
authors.lastname: Gharib
authors.forename: Brandon
authors.initials: B
authors.lastname: Wyse
authors.forename: Erin
authors.initials: E
authors.lastname: Drury
authors.forename: Michael
authors.initials: M
authors.lastname: Weinreich
authors.forename: Pierre
authors.initials: P
authors.lastname: Thibault
authors.forename: Alain
authors.initials: A
authors.lastname: Verreault
authors.forename: Hisao
authors.initials: H
authors.lastname: Masai
authors.forename: Krassimir
authors.initials: K
authors.lastname: Yankulov
mesh_terms: D000595:Amino Acid Sequence; D017489:CDC28 Protein Kinase, S cerevisiae / Q000378:metabolism*; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism; D002843:Chromatin / Q000378:metabolism*; D056504:Chromatin Assembly Factor-1 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D042002:Chromatin Assembly and Disassembly; D004261:DNA Replication; D020868:Gene Silencing; D008969:Molecular Sequence Data; D009154:Mutation; D010766:Phosphorylation; D018809:Proliferating Cell Nuclear Antigen / Q000378:metabolism; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism; D016196:S Phase; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D016615:Telomere / Q000235:genetics / Q000378:metabolism
chemical_list: D018797:Cell Cycle Proteins; D002843:Chromatin; D056504:Chromatin Assembly Factor-1; D018809:Proliferating Cell Nuclear Antigen; C533589:RLF2 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; C489404:CDC7 protein, S cerevisiae; D017346:Protein Serine-Threonine Kinases; D017489:CDC28 Protein Kinase, S cerevisiae
keywords: CAC1, the largest subunit of CAF-I; CAF-I, Chromatin Assembly Factor I; CDK, Cyclin-Dependent Kinase; Cdc28p; Cell cycle; Chromatin Assembly Factor I (CAF-I); Cyclin-Dependent Kinase 8 (CDK8); DDK, Dbf4-Dependent Kinase, Cdc7p-Dbf4p; Dbf4-Dependent Kinase (DDK); PCNA, Proliferating Cell Nuclear Antigen, POL30; PIP, PCNA Interaction Peptide; Proliferating Cell Nuclear Antigen (PCNA); TPE; Telomere Position Effect