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Human replication protein Cdc6 prevents mitosis through a checkpoint mechanism that implicates Chk1
Lorena Clay-Farrace1, Cristina Pelizon, David Santamaria
1MRC Cancer Cell Unit, Hutchison/MRC Research Centre, Hills Road, Cambridge CB2 2XZ, UK. lc321@cam.ac.uk
The EMBO Journal
|January 30, 2003
Summary
Human Cdc6 (HuCdc6) protein, known for DNA replication, also blocks entry into mitosis. This cell cycle regulation involves the Chk1 kinase pathway, ensuring DNA replication completion before cell division.
Area of Science:
- Cell Biology
- Molecular Biology
- Genetics
Background:
- Replication protein Cdc6/Cdc18 is crucial for DNA replication initiation and cell cycle progression in yeasts.
- In metazoans, Cdc6's role was primarily linked to S phase entry, not mitotic regulation.
Purpose of the Study:
- To investigate the role of human Cdc6 (HuCdc6) in regulating the onset of mitosis.
- To elucidate the molecular mechanisms by which HuCdc6 influences cell cycle progression into mitosis.
Main Methods:
- Overexpression of HuCdc6 in G(2) phase cells.
- Co-expression experiments with active Cyclin B/CDK1, Cdc25B, and Cdc25C.
- Utilizing Chk1 kinase inhibitor UCN-01.
- Detection of Chk1 phosphorylation.
Main Results:
- HuCdc6 overexpression in G(2) phase cells prevents entry into mitosis.
- This mitotic block is rescued by co-expressing active Cyclin B/CDK1, Cdc25B, or Cdc25C.
- HuCdc6-induced G(2) arrest is overcome by a Chk1 inhibitor, indicating involvement of the Chk1 checkpoint pathway.
- Phosphorylation of Chk1 was observed upon HuCdc6 overexpression in G(2).
Conclusions:
- Human Cdc6 (HuCdc6) plays a significant role in regulating the G(2)/M transition, extending beyond its known function in S phase.
- HuCdc6 can activate a checkpoint pathway involving Chk1, potentially ensuring complete DNA replication before mitosis.
- The phosphorylation status of HuCdc6 may regulate its ability to inhibit mitotic entry.