title: A CDK-catalysed regulatory phosphorylation for formation of the DNA replication complex Sld2-Dpb11.
abstract: Phosphorylation often regulates protein-protein interactions to control biological reactions. The Sld2 and Dpb11 proteins of budding yeast form a phosphorylation-dependent complex that is essential for chromosomal DNA replication. The Sld2 protein has a cluster of 11 cyclin-dependent kinase (CDK) phosphorylation motifs (Ser/Thr-Pro), six of which match the canonical sequences Ser/Thr-Pro-X-Lys/Arg, Lys/Arg-Ser/Thr-Pro and Ser/Thr-Pro-Lys/Arg. Simultaneous alanine substitution for serine or threonine in all the canonical CDK-phosphorylation motifs severely reduces complex formation between Sld2 and Dpb11, and inhibits DNA replication. Here we show that phosphorylation of these canonical motifs does not play a direct role in complex formation, but rather regulates phosphorylation of another residue, Thr84. This constitutes a non-canonical CDK-phosphorylation motif within a 28-amino-acid sequence that is responsible, after phosphorylation, for binding of Sld2-Dpb11. We further suggest that CDK-catalysed phosphorylation of sites other than Thr84 renders Thr84 accessible to CDK. Finally, we argue that this novel mechanism sets a threshold of CDK activity for formation of the essential Sld2 to Dpb11 complex and therefore prevents premature DNA replication.
authors.affiliation: Division of Microbial Genetics, National Institute of Genetics, Research Organization of Information and Systems, Mishima, Shizuoka, Japan.
authors.forename: Yon-Soo
authors.initials: YS
authors.lastname: Tak
authors.forename: Yoshimi
authors.initials: Y
authors.lastname: Tanaka
authors.forename: Shizuko
authors.initials: S
authors.lastname: Endo
authors.forename: Yoichiro
authors.initials: Y
authors.lastname: Kamimura
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Araki
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D002384:Catalysis; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism*; D018844:Cyclin-Dependent Kinases / Q000378:metabolism*; D004261:DNA Replication; D008969:Molecular Sequence Data; D010766:Phosphorylation; D025941:Protein Interaction Mapping; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D017384:Sequence Deletion; D013912:Threonine / Q000235:genetics / Q000378:metabolism
chemical_list: D018797:Cell Cycle Proteins; C097185:DPB11 protein, S cerevisiae; C115043:SLD2 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D013912:Threonine; D018844:Cyclin-Dependent Kinases