Mismatch repair-mediated G2/M arrest by 6-thioguanine involves the ATR-Chk1 pathway.
DNA mismatch repair (MMR) deficiency in human cancers is associated with resistance to a spectrum of clinically active chemotherapy drugs, including 6-thioguanine (6-TG). We and others have shown that 6-TG-induced DNA mismatches result in a prolonged G2/M cell cycle arrest followed by apoptosis in MMR(+) human cancer cells, although the signaling pathways are not clearly understood. In this study, we found that prolonged (up to 4 days) treatment with 6-TG (3microM) resulted in a progressive phosphorylation of Chk1 and Chk2 in MMR(+) HeLa cells, correlating temporally with a drug-induced G2/M arrest. Transfection of HeLa cells with small interfering RNA (siRNA) against the ataxia telangiectasia-related (ATR) kinase or against the Chk1 kinase destroyed the G2/M checkpoint and enhanced the apoptosis following 6-TG treatment. On the other hand, the induction of a G2/M population by 6-TG was similar in ATM(-/-) and ATM(+) human fibroblasts, suggesting that the ATM-Chk2 pathway does not play a major role in this 6-TG response. Our results indicate that 6-TG DNA mismatches activate the ATR-Chk1 pathway in the MMR(+) cells, resulting in a G2/M checkpoint response
Yamane
Kazuhiko
K
Department of Radiation Oncology, Case Western Reserve University, Case Comprehensive Cancer Center/University Hospitals of Cleveland, Cleveland, OH 44106-4942, USA.
Taylor
Kerri
K
Kinsella
Timothy J
TJ
D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects; D064007:Ataxia Telangiectasia Mutated Proteins; D020137:Base Pair Mismatch / Q000235:genetics*; D015153:Blotting, Western; D018797:Cell Cycle Proteins / Q000378:metabolism*; D000071877:Checkpoint Kinase 1; D064447:Checkpoint Kinase 2; D004260:DNA Repair / Q000235:genetics*; D016195:G2 Phase / Q000187:drug effects* / Q000502:physiology; D006367:HeLa Cells; D006801:Humans; D020287:In Situ Nick-End Labeling; D008938:Mitosis / Q000187:drug effects* / Q000502:physiology; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000235:genetics; D015398:Signal Transduction; D013866:Thioguanine / Q000494:pharmacology*; D014162:Transfection
D000964:Antimetabolites, Antineoplastic; D018797:Cell Cycle Proteins; D034741:RNA, Small Interfering; D011494:Protein Kinases; D064447:Checkpoint Kinase 2; C576597:ATR protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; C000605867:CHEK1 protein, human; C578634:CHEK2 protein, human; D000071877:Checkpoint Kinase 1; D017346:Protein Serine-Threonine Kinases; D013866:Thioguanine