Induction of endomitosis-like event in HeLa cells following CHK1 inhibitor treatment

Hisao Homma; Hitomi Nojima; Atsushi Kaida; Masahiko Miura

doi:10.1016/j.bbrc.2019.09.046

Induction of endomitosis-like event in HeLa cells following CHK1 inhibitor treatment

Biochem Biophys Res Commun. 2019 Dec 3;520(2):492-497. doi: 10.1016/j.bbrc.2019.09.046. Epub 2019 Oct 11.

Authors

Hisao Homma¹, Hitomi Nojima², Atsushi Kaida¹, Masahiko Miura³

Affiliations

¹ Department of Oral Radiation Oncology, Division of Oral Health Sciences, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, 1-5-45 Yushima, Bunkyo-ku, Tokyo, 113-8549, Japan.
² Department of Oral Radiation Oncology, Division of Oral Health Sciences, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, 1-5-45 Yushima, Bunkyo-ku, Tokyo, 113-8549, Japan; Department of Oral and Maxillofacial Surgery, Division of Oral Health Sciences, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, 1-5-45 Yushima, Bunkyo-ku, Tokyo, 113-8549, Japan.
³ Department of Oral Radiation Oncology, Division of Oral Health Sciences, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, 1-5-45 Yushima, Bunkyo-ku, Tokyo, 113-8549, Japan. Electronic address: masa.mdth@tmd.ac.jp.

PMID: 31610912
DOI: 10.1016/j.bbrc.2019.09.046

Abstract

The effects of CHK1 inhibitor on cell cycle kinetics have not been fully investigated yet. In this study, we closely analyzed this kinetics using a CHK1 inhibitor (PF00477736) in HeLa cells expressing fluorescent ubiquitination-based cell cycle indicator (Fucci). This system allowed us to visualize cell cycle progression following CHK1 inhibitor treatment in real-time. FACS analysis showed that high levels of DNA damage as determined by γH2AX immunostaining was induced in S phase and that polyploid cells harboring the same levels of DNA damage appeared thereafter. Surprisingly, time-lapse imaging of Fucci fluorescence revealed that many cells entered M phase at once and exhibited prolonged mitosis; eventually progressing to G1 phase not accompanied by cytokinesis; this is an endomitosis-like event. Most of these cells then underwent S/G2 phases at least once, which corroborated the appearance of polyploid cells. However, a small fraction of cells with 2 N DNA content still remained 24 h after the treatment. When co-treated with MAD2 inhibitor, a core factor constituting spindle checkpoint, the 2 N DNA cell fraction disappeared and almost all cells exhibited endomitosis, leading to enhanced sensitivity. Detailed cell cycle analysis revealed that induction of an endomitosis-like event might be associated with CHK1 inhibitor-induced cell death in HeLa cells.

Keywords: CHK1; Endomitosis; Fluorescent ubiquitination-based cell cycle indicator (Fucci); MAD2; Polyploidy; Spindle checkpoint.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Benzodiazepinones / pharmacology*
Cell Cycle / drug effects
Cell Survival
Checkpoint Kinase 1 / antagonists & inhibitors*
Checkpoint Kinase 1 / metabolism*
DNA Damage
Fluorescence
HeLa Cells
Humans
M Phase Cell Cycle Checkpoints / drug effects
Mad2 Proteins / antagonists & inhibitors
Mad2 Proteins / metabolism
Mitosis / drug effects*
Phosphorylation / drug effects
Polyploidy
Protein Kinase Inhibitors / pharmacology*
Pyrazoles / pharmacology*
Time-Lapse Imaging

Substances

Benzodiazepinones
MAD2L1 protein, human
Mad2 Proteins
PF 00477736
Protein Kinase Inhibitors
Pyrazoles
CHEK1 protein, human
Checkpoint Kinase 1