1. Cell cycle-dependent phosphorylation of the large subunit of replication factor C (RF-C) leads to its dissociation from the RF-C complex.
- Author
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Munshi A, Cannella D, Brickner H, Salles-Passador I, Podust V, Fotedar R, and Fotedar A
- Subjects
- Animals, CDC2-CDC28 Kinases metabolism, COS Cells, Cell Cycle, Centrifugation, Density Gradient, Cyclin B metabolism, Cyclin E metabolism, Cyclin-Dependent Kinase 2, DNA-Binding Proteins metabolism, Electrophoresis, Polyacrylamide Gel, Epitopes, G2 Phase, HeLa Cells, Humans, Mitosis, Phosphorylation, Precipitin Tests, Protein Binding, Protein Structure, Tertiary, Recombinant Proteins chemistry, Recombinant Proteins metabolism, Replication Protein C, S Phase, Time Factors, DNA-Binding Proteins chemistry
- Abstract
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.
- Published
- 2003
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