Journal Article

Resveratrol causes Cdc2-tyr15 phosphorylation via ATM/ATR–Chk1/2–Cdc25C pathway as a central mechanism for S phase arrest in human ovarian carcinoma Ovcar-3 cells

Alpna Tyagi, Rana P. Singh, Chapla Agarwal, Sunitha Siriwardana, Robert A. Sclafani and Rajesh Agarwal

in Carcinogenesis

Volume 26, issue 11, pages 1978-1987
Published in print November 2005 | ISSN: 0143-3334
Published online June 2005 | e-ISSN: 1460-2180 | DOI: http://dx.doi.org/10.1093/carcin/bgi165
Resveratrol causes Cdc2-tyr15 phosphorylation via ATM/ATR–Chk1/2–Cdc25C pathway as a central mechanism for S phase arrest in human ovarian carcinoma Ovcar-3 cells

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Resveratrol is one of the most extensively studied cancer chemopreventive agents; however, its mechanisms of action are not completely understood. Here, we observed that resveratrol induces S phase arrest via Tyr15 phosphorylation of Cdc2 in human ovarian carcinoma Ovcar-3 cells. Overexpression of Cdc2AF, a mutant resistant to Thr14 and Tyr15 phosphorylation, ablated resveratrol-induced S phase arrest. Further upstream, we observed that resveratrol causes phosphorylation of cell division cycle 25C (Cdc25C) tyrosine phosphatase via the activation of checkpoint kinases Chk1 and Chk2, which in turn were activated via ATM (ataxia telangiectasia mutated)/ATR (ataxia telangiectasia-Rad3-related) kinase in response to DNA damage, as resveratrol also increased phospho-H2A.X (Ser139), which is known to be phosphorylated by ATM/ATR in response to DNA damage. The involvement of these molecules in resveratrol-induced S phase was also supported by the studies showing that addition of ATM/ATR inhibitor caffeine reverses resveratrol-caused activation of ATM/ATR–Chk1/2 as well as phosphorylation of Cdc25C, Cdc2 and H2A.X, and S phase arrest. In additional studies assessing whether observed effects of resveratrol are specific to Ovcar-3 cells, we observed that it also induces S phase arrest and H2A.X (Ser139) phosphorylation in other ovarian cancer cell lines PA-1 and SKOV-3, albeit at different levels; whereas, resveratrol showed only marginal S phase arrest in normal human foreskin fibroblasts with undetectable level of phospho-H2A.X (Ser139). These findings for the first time identify that resveratrol causes Cdc2-tyr15 phosphorylation via ATM/ATR–Chk1/2–Cdc25C pathway as a central mechanism for DNA damage and S phase arrest selectively in ovarian cancer cells, and provide a rationale for the potential efficacy of ATM/ATR agonists in the prevention and intervention of cancer.

Keywords: ATM, ataxia telangiectasia-mutated; ATR, ataxia telangiectasia-Rad3-related; Cdc25C, cell division cycle 25C; Chk, checkpoint kinase; H2A.X, histone 2AX; GFP, green fluorescent protein; tTA, tetracycline transactivator; FACS, fluorescence activated cell sorting; HFF, human foreskin fibroblast; PBS, phosphate-buffered saline.

Journal Article.  7109 words.  Illustrated.

Subjects: Clinical Cytogenetics and Molecular Genetics

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