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PMID: 15632059 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

Chk1-dependent S-M checkpoint delay in vertebrate cells is linked to maintenance of viable replication structures.

Molecular and cellular biology ·Vol. 25 ·No. 2 ·2005-01-00 ·Pages 563-74

Zachos G, Rainey MD, Gillespie DA

Abstract

We investigated mitotic delay during replication arrest (the S-M checkpoint) in DT40 B-lymphoma cells deficient in the Chk1 or Chk2 kinase. We show here that cells lacking Chk1, but not those lacking Chk2, enter mitosis with incompletely replicated DNA when DNA synthesis is blocked, but only after an initial delay. This initial delay persists when S-M checkpoint failure is induced in Chk2-/- cells with the Chk1 inhibitor UCN-01, indicating that it does not depend on Chk1 or Chk2 activity. Surprisingly, dephosphorylation of tyrosine 15 did not accompany Cdc2 activation during premature entry to mitosis in Chk1-/- cells, although mitotic phosphorylation of cyclin B2 did occur. Previous studies have shown that Chk1 is required to stabilize stalled replication forks during replication arrest, and strikingly, premature mitosis occurs only in Chk1-deficient cells which have lost the capacity to synthesize DNA as a result of progressive replication fork inactivation. These results suggest that Chk1 maintains the S-M checkpoint indirectly by preserving the viability of replication structures and that it is the continued presence of such structures, rather than the activation of Chk1 per se, which delays mitosis until DNA replication is complete.

MeSH Terms
Animals Antineoplastic Agents/metabolism Aphidicolin/metabolism CDC2 Protein Kinase/metabolism Cell Line, Tumor Checkpoint Kinase 1 Checkpoint Kinase 2 Chickens DNA Replication Gene Targeting Mitosis/physiology Nocodazole/metabolism Nucleic Acid Conformation Phosphorylation Protein Kinase Inhibitors/metabolism Protein Kinases/genetics,metabolism Protein Serine-Threonine Kinases/genetics,metabolism Staurosporine/analogs & derivatives,metabolism
Chemicals
Antineoplastic Agents Protein Kinase Inhibitors Aphidicolin 7-hydroxystaurosporine Protein Kinases Checkpoint Kinase 2 Checkpoint Kinase 1 Protein Serine-Threonine Kinases CDC2 Protein Kinase Staurosporine Nocodazole
Authors & Affiliations
3 authors, click to expand affiliations / ORCID
Zachos George
Beatson Institute for Cancer Research, Garscube Estate, Switchback Rd., Glasgow G61 1BD, United Kingdom.
Rainey Michael D
Gillespie David A F
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Article Info
Journal
Molecular and cellular biology
Abbr.
Mol Cell Biol
ISSN
0270-7306
DOI
10.1128/MCB.25.2.563-574.2005
Published
2005-01-00
Pages
563-74
Language
English
Region
United States
NLM ID
8109087
PMCID
PMC543419
Subset
IM
External Links
PubMed source DOI: 10.1128/MCB.25.2.563-574.2005 PMC Full Text View journal details
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