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Updated: Jun 1, 2026

Combining Mitotic Cell Synchronization and High Resolution Confocal Microscopy to Study the Role of Multifunctional Cell Cycle Proteins During Mitosis
08:33

Combining Mitotic Cell Synchronization and High Resolution Confocal Microscopy to Study the Role of Multifunctional Cell Cycle Proteins During Mitosis

Published on: December 5, 2017

Nuclear Chk1 prevents premature mitotic entry

Makoto Matsuyama1, Hidemasa Goto, Kousuke Kasahara

  • 1Division of Biochemistry, Aichi Cancer Center Research Institute, Chikusa-ku, Nagoya 464-8681, Japan. minagaki@aichi-cc.jp

Journal of Cell Science
|June 2, 2011
PubMed

Insights

Nuclear Chk1 (Checkpoint kinase 1) regulates the timing of mitotic entry, while centrosomal staining was found to be a false positive. This clarifies Chk1

Area of Science:

  • Cell cycle regulation
  • Molecular biology
  • Cancer research

Background:

  • Checkpoint kinase 1 (Chk1) is known to inhibit cyclin-B1-Cdk1 activation, preventing premature mitosis.
  • The precise location where Chk1 exerts its inhibitory function (nucleus or centrosome) before the G2-M transition is debated.

Purpose of the Study:

  • To investigate the specific role of Chk1 localization in regulating the timing of mitotic entry.
  • To determine whether Chk1 functions in the nucleus or centrosome before the G2-M transition.

Main Methods:

  • Utilized a specific anti-Chk1 antibody (DCS-310) and conditional Chk1 knockout mouse embryonic fibroblasts.
  • Employed anti-Myc antibody in Chk1(+/myc) human colon adenocarcinoma (DLD-1) cells.
  • Combined protein array and RNAi technologies to identify cross-reacting proteins.
  • Generated Chk1 mutants with altered subcellular localization (nuclear vs. centrosomal).

Main Results:

  • The anti-Chk1 antibody DCS-310 showed cross-reactivity with Ccdc-151 at the centrosome.
  • Conditional Chk1 knockout reduced nuclear staining but not centrosomal staining with DCS-310.
  • Chk1 was detected in the nucleus, not the centrosome, in DLD-1 cells.
  • Nuclear Chk1 localization delayed mitotic entry, whereas centrosomal Chk1 had minimal impact.

Conclusions:

  • Nuclear Chk1, not centrosomal Chk1, is crucial for the correct timing of mitotic entry.
  • The anti-Chk1 antibody DCS-310 can lead to misinterpretation of Chk1 localization at the centrosome due to cross-reactivity.

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