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Arecoline‐induced phosphorylated p53 and p21 WAF1 protein expression is dependent on ATM/ATR and phosphatidylinositol‐3‐kinase in clone‐9 cells
Author(s) -
Chou WenWen,
Guh JinnYuh,
Tsai JungFa,
Hwang ChiChing,
Chiou SheanJaw,
Chuang LeaYea
Publication year - 2009
Publication title -
journal of cellular biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.028
H-Index - 165
eISSN - 1097-4644
pISSN - 0730-2312
DOI - 10.1002/jcb.22137
Subject(s) - arecoline , microbiology and biotechnology , pi3k/akt/mtor pathway , kinase , phosphorylation , cancer research , phosphatidylinositol , clone (java method) , chemistry , biology , biochemistry , signal transduction , dna , receptor , muscarinic acetylcholine receptor
Betel‐quid use is associated with liver cancer whereas its constituent arecoline is cytotoxic, genotoxic, and induces p53‐dependent p21 WAF1 protein expression in Clone‐9 cells (rat hepatocytes). The ataxia telangiectasia mutated (ATM)/rad3‐related (ATR)‐p53‐p21 WAF1 and the phosphatidylinositol‐3‐kinase (PI3K)‐mammalian target of rapamycin (mTOR) pathways are involved in the DNA damage response and the pathogenesis of cancers. Thus, we studied the role of ATM/ATR and PI3K in arecoline‐induced p53 and p21 WAF1 protein expression in Clone‐9 cells. We found that arecoline (0.5 mM) activated the ATM/ATR kinase at 30 min. The arecoline‐activated ATM/ATR substrate contained p‐p53Ser15. Moreover, arecoline only increased the levels of the p‐p53Ser6, p‐p53Ser15, and p‐p53Ser392 phosphorylated p53 isoforms among the known isoforms. ATM shRNA attenuated arecoline‐induced p‐p53Ser15 and p21 WAF1 at 24 h. Arecoline (0.5 mM) increased phosphorylation levels of p‐AktSer473 and p‐mTORSer2448 at 30–60 min. Dominant‐negative PI3K plasmids attenuated arecoline‐induced p21 WAF1 , but not p‐p53Ser15, at 24 h. Rapamycin attenuated arecoline‐induced phosphrylated p‐p53Ser15, but not p21 WAF1 , at 24 h. ATM shRNA, but not dominant‐negative PI3K plasmids, attenuated arecoline‐induced p21 WAF1 gene transcription. We conclude that arecoline activates the ATM/ATR‐p53‐p21 WAF1 and the PI3K/Akt‐mTOR‐p53 pathways in Clone‐9 cells. Arecoline‐induced phosphorylated p‐p53Ser15 expression is dependent on ATM whereas arecoline‐induced p21 WAF1 protein expression is dependent on ATM and PI3K. Moreover, p21 WAF1 gene is transcriptionally induced by arecoline‐activated ATM. J. Cell. Biochem. 107: 408–417, 2009. © 2009 Wiley‐Liss, Inc.