Involvement of ATM/ATR-p38 MAPK cascade in MNNG induced G1-S arrest.
OBJECTIVE
To understand the effect of low concentration of N-methyl-N'-nitro-nitrosoguanidine (MNNG), which is a widely distributed environmental mutagen and carcinogen especially for human gastric cancer, on DNA damage and to study its possible pathway in regulating cell cycle arrest.


METHODS
The DNA damage effect was measured by Comet assay. A specific phospho-(Ser/Thr) ATM/ATR substrate antibody was used to detect the damage sensor by Western blot. p38 kinase activity was measured by direct kinase assay, and immunoprecipitation for the possible connection between ATM/ATR and p38 MAPK. Flow cytometry analysis and p38 MAPK specific inhibitor SB203580 were combined to detect the possible cell cycle arrest by p38 MAPK.


RESULTS
With the same low concentration MNNG exposure (0.2 microM 2.5 h), Comet assays indicated that strand breaks accumulated, Western blot and kinase assay showed ATM/ATR and p38 kinase activated, immunoprecipitation showed phospho-ATM/ATR substrate antibody combined with both p38 MAPK antibody and phospho-p38 MAPK antibody. p38 MAPK pathway was involved in the G1-S arrest.


CONCLUSIONS
Activation of ATM/ATR by MNNG induced DNA damage leads to activation of p38 MAPK, which involves in the G1 checkpoint in mammalian cells.
Zhu
Ke-Qing
KQ
Department of Pathology, School of Medicine, Zhejiang University, Hangzhou 310031, Zhejiang Province, China.
Zhang
Suo-Jiang
SJ
D000818:Animals; D064007:Ataxia Telangiectasia Mutated Proteins; D018797:Cell Cycle Proteins / Q000378:metabolism*; D002522:Chlorocebus aethiops; D004268:DNA-Binding Proteins; D016193:G1 Phase / Q000187:drug effects* / Q000502:physiology; D008769:Methylnitronitrosoguanidine / Q000494:pharmacology*; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D016196:S Phase / Q000187:drug effects* / Q000502:physiology; D025521:Tumor Suppressor Proteins; D014709:Vero Cells; D048051:p38 Mitogen-Activated Protein Kinases
D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D025521:Tumor Suppressor Proteins; D008769:Methylnitronitrosoguanidine; C576324:ATM protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; D017346:Protein Serine-Threonine Kinases; D020928:Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases