Zeng Xuehuo, Yan Tao, Schupp Jane E, Seo Yuji, Kinsella Timothy J
Department of Radiation Oncology, Case Western Reserve University and the Case Integrative Cancer Biology Program, Case Comprehensive Cancer Center/University Hospitals of Cleveland, Cleveland, Ohio 44106-6068, USA.
Clin Cancer Res. 2007 Feb 15;13(4):1315-21. doi: 10.1158/1078-0432.CCR-06-1517.
We investigate the roles of DNA mismatch repair (MMR) and p53 in mediating the induction of autophagy in human tumor cells after exposure to 6-thioguanine (6-TG), a chemotherapy drug recognized by MMR. We also examine how activation of autophagy affects apoptosis (type I cell death) after MMR processing of 6-TG.
Using isogenic pairs of MLH1(-)/MLH1(+) human colorectal cancer cells (HCT116) and MSH2(-)/MSH2(+) human endometrial cancer cells (HEC59), we initially measure activation of autophagy for up to 3 days after 6-TG treatment using LC3, a specific marker of autophagy. We then assess the role of p53 in autophagic signaling of 6-TG MMR processing using both pifithrin-alpha cotreatment to chemically inhibit p53 transcription and small hairpin RNA inhibition of p53 expression. Finally, we use Atg5 small hairpin RNA inhibition of autophagy to assess the effect on apoptosis after MMR processing of 6-TG.
We find that MMR is required for mediating autophagy in response to 6-TG treatment in these human tumor cells. We also show that p53 plays an essential role in signaling from MMR to the autophagic pathway. Finally, our results indicate that 6-TG-induced autophagy inhibits apoptosis after MMR processing of 6-TG.
These data suggest a novel function of MMR in mediating autophagy after a chemical (6-TG) DNA mismatch damage through p53 activation. The resulting autophagy inhibits apoptosis after MMR processing of 6-TG.
我们研究了DNA错配修复（MMR）和p53在介导人肿瘤细胞暴露于6-硫鸟嘌呤（6-TG，一种被MMR识别的化疗药物）后自噬诱导中的作用。我们还研究了自噬激活如何影响6-TG经MMR处理后的细胞凋亡（I型细胞死亡）。
使用MLH1(-)/MLH1(+)人结肠癌细胞（HCT116）和MSH2(-)/MSH2(+)人子宫内膜癌细胞（HEC59）的同基因细胞对，我们最初使用自噬的特异性标志物LC3测量6-TG处理后长达3天的自噬激活情况。然后，我们使用pifithrin-α共处理化学抑制p53转录以及用小发夹RNA抑制p53表达，来评估p53在6-TG MMR处理的自噬信号传导中的作用。最后，我们使用Atg5小发夹RNA抑制自噬，以评估对6-TG经MMR处理后细胞凋亡的影响。
我们发现MMR是介导这些人肿瘤细胞对6-TG处理产生自噬所必需的。我们还表明p53在从MMR到自噬途径的信号传导中起关键作用。最后，我们的结果表明6-TG诱导的自噬在6-TG经MMR处理后抑制细胞凋亡。
这些数据表明MMR在通过p53激活介导化学（6-TG）DNA错配损伤后的自噬中具有新功能。由此产生的自噬在6-TG经MMR处理后抑制细胞凋亡
