|Depamphilis, Melvin - NICHD, NIH, BETHESDA|
Submitted to: Molecular and Cellular Biology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: March 23, 2004
Publication Date: July 1, 2004
Citation: Li, C., Vassilev, A., DePamphilis, M.L. 2004. Role for Cdk1 (Cdc2)/Cyclin A in preventing the mammalian origin recognition complex's largest subunit (Orc1) from binding to chromatin during mitosis. Molecular and Cellular Biology. 24(13) p.5875-5886. Interpretive Summary: When cells growing and proliferating (dividing), the first and most important step in the process is to copy their entire genomic information. This step is called DNA replication. Animal cells have a very complex control system to ensure the integrity of the genomic information from generation to generation, DNA replication has to be once per cell cycle to make a full copy of genome, no more, nor less. Origin Recognition Complexes (ORC, which includes six proteins) are the proteins to control the start of DNA replication. The mechanisms that regulate the function of ORC is the subject of this study. When all six component of this protein complex bound to DNA, the replication started. When DNA replication finished, one of the ORC protein (Orc1) was released from DNA and this protein complex lost its function. We also found that Cdk1, a cell cycle-dependent protein kinase, selectively bound to Orc1 after DNA replication fully completed. This protein kinase modifies the Orc1 protein with a phosphoryl-group and prevent Orc1 premature rebinding of DNA, therefore, preventing the formation of functional ORC/chromatin complexes, and preventing DNA replication before new cell cycle started.
Technical Abstract: The eukaryotic origin recognition complex (ORC) selects the genomic sites where pre-replication complexes are assembled and DNA replication begins. ORC activity in mammalian cells is regulated during cell division by selectively releasing the Orc1 subunit in S-phase and then rebinding it to chromatin at the beginning of G1-phase. Premature rebinding of Orc1 in human cells is largely prevented by Orc1 degradation during S-phase, but in hamster cells, Orc1 is not degraded in vivo. Here we show that in hamster cells, Orc1 is selectively bound to Cdk1(Cdc2)/cyclin A protein kinase specifically during the G2/M period when Orc1 becomes hyperphosphorylated, and that premature rebinding of Orc1 is prevented during mitosis by a cyclin-dependent protein kinase activity. These and other data reveal that Cdk1/cyclin B induces mitosis while Cdk1/cyclin A inhibits assembly of functional ORC/chromatin complexes, the first step in initiating DNA replication.