Cell cycle-dependent phosphorylation of the large subunit of replication factor C (RF-C) leads to its dissociation from the RF-C complex.
The five subunit replication factor C (RF-C) complex plays a critical role in DNA elongation. We find that the large subunit of RF-C (RF-Cp145) is phosphorylated in vivo whereas the smaller RF-C subunits are not phosphorylated. The phosphorylation of endogenous RFCp145 is modulated in a cell cycle-dependent manner. Phosphorylation is maximal in G2/M and is inhibited by an inhibitor of cyclin-dependent kinases. Phosphorylation of purified recombinant RF-C complex in vitro reveals that RF-Cp145 is preferentially phosphorylated by cdc2-cyclin B but not by cdk2-cyclin A or cdk2-cyclin E. In vitro phosphorylation of RF-C complex by cdc2-cyclin B kinases leads to dissociation of phosphorylated RFCp145 from the RF-C complex. Using different approaches we demonstrate that phosphorylated RFCp145 is indeed dissociated from RF-Cp40 and RF-Cp37 in vivo. These results suggest that destabilization of the RF-C complex by CDKs may inactivate the RF-C complex at the end of S phase.
Munshi
Anil
A
Sidney Kimmel Cancer Center, San Diego, California 92121, USA.
Cannella
Dominique
D
Brickner
Howard
H
Salles-Passador
Isabelle
I
Podust
Vladimir
V
Fotedar
Rati
R
Fotedar
Arun
A
D000818:Animals; D042846:CDC2-CDC28 Kinases / Q000378:metabolism; D019556:COS Cells; D002453:Cell Cycle; D002499:Centrifugation, Density Gradient; D019926:Cyclin B / Q000378:metabolism; D019927:Cyclin E / Q000378:metabolism; D051357:Cyclin-Dependent Kinase 2; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D000939:Epitopes; D016195:G2 Phase; D006367:HeLa Cells; D006801:Humans; D008938:Mitosis; D010766:Phosphorylation; D011233:Precipitin Tests; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D051818:Replication Protein C; D016196:S Phase; D013997:Time Factors
D019926:Cyclin B; D019927:Cyclin E; D004268:DNA-Binding Proteins; D000939:Epitopes; D011994:Recombinant Proteins; D042846:CDC2-CDC28 Kinases; C495894:CDK2 protein, human; D051357:Cyclin-Dependent Kinase 2; D051818:Replication Protein C