组蛋白甲基转移酶SETD2对P53介导细胞凋亡的调控

摘要

Objective To investigate the role of the histone methyltransferase SETD2 on the regulation of P53-mediated apoptosis.Methods Amplified the Setd2 gene by PCR to recombine it into the eukaryotic expression vector.Using indirect immunofluorescence assay (ⅡF) to analyze the subcellular localization of Myc-tagged SETD2 protein.A glutathione Stransferase (GST) pull-down assay was performed to detect the direct interaction between P53 and SETD2 truncates.Flow cytometry (FCM) was used to study the effect of SETD2 on P53-dependent cell apoptosis.Results Setd2 gene was amplified by PCR and constructed into eukaryotic expression vector.SETD2 protein was mainly localized in the nucleus.GST pull-down assay showed that SETD2 interacted with P53 directly.The SET domain and the C-terminus of SETD2 mediated the interaction with P53.Overexpression of SETD2 enhanced the P53-promoted cell apoptosis.Conclusion These findings suggest that the histone methyltransferase SETD2 upregulate P53-dependent apoptosis.%目的 探讨组蛋白甲基转移酶SETD2对P53功能的调控.方法 PCR扩增人源SETD2并重组入真核表达载体;通过间接免疫荧光鉴定SETD2的亚细胞定位;通过GST Pull-down实验寻找P53结合SETD2的区域;流式细胞术分析SETD2对P53参与细胞凋亡的影响.结果 成功克隆了人源SETD2的全长,并将其成功构建到真核表达载体中;SETD2以胞核定位为主;体外直接相互作用实验证实SETD2和P53存在相互作用,SETD2分子中部的SET 结构域和C端都可以结合P53;流式细胞分析结果表明SETD2对于P53参与的细胞凋亡具有明显的促进作用.结论 本研究揭示P53正向调控因子SETD2对P53介导的细胞凋亡具有促进效应,从而将组蛋白表遗传学修饰与P53转录因子活性调控关联起来,研究结果对肿瘤、免疫等疾病的防治具有一定的科学意义.