病毒白细胞介素-6对抑癌基因p16启动子区甲基化及蛋白表达的影响

目的观察病毒白细胞介素-6（viral interleukin-6，vIL-6）对抑癌基因p16启动子区甲基化及蛋白表达的影响。方法以vIL-6真核表达质粒转染293T细胞（转染组）为实验，空载体转染的293T细胞（空载体组）和未转染的293T细胞（未转染组）为对照，3组均采用Western blot法检测DNA甲基转移酶1的表达情况，甲基化特异性PCR法检测p16启动子区甲基化状态，荧光定量RT—PCR和Western blot法检测p16基因的表达。结果转染组DNA甲基转移酶1蛋白表达较未转染组和空载体组增加；转染组p16基因启动子区出现甲基化改变；转染组p16 mRNA表达量明显低于未转染组和空载体组，差异有统计学意义（P〈O．05）；转染组P16蛋白表达较未转染组和空载体组降低。结论vIL-6可上调DNA甲基转移酶1表达，导致抑癌基因p16启动子区甲基化，抑制p16基因表达；vIL-6所致的DNA甲基化改变，可能是其致瘤机制之一。

Influence of viral interleukin-6 on DNA methylation of tumor suppressor gene p16 promoter and its protein expression

Objective To observe the effect of viral interleukin-6 （vIL-6） on the methylation status of tumor suppressor gene p16 promoter and its protein expression. Methods The eukaryotic expression plasmid carrying vIL-6 gene was transfected into 293T ceils （transfection group）. And 293T cells were transfected with empty vectors （empty vectors group） and the untransfected 293T cells were as controls （untransfection group）. The expression of DNA methyltransferase （DNMT）-I was identified with Western blot. The methylation status of p16 gene promoter was analyzed with methylation-specific PCR. The p16 gene expression was detected with quantitative RT-PCR and Western blot. Results The expression of DNMT-1 proteins increased in transfection group in comparison with untransfection group. Methylation of p16 gene promoter occurred in the 293T cells transfected with the vIL-6 gene. The expression of p16 mRNA decreased in the transfeetion group in comparison with the other two groups, which showed a significant difference （P~0.05）. The expression of P16 protein also decreased in the transfection group in comparison with the other two groups. Conclusion vIL-6 can upregulate the expression of DNMT-1, induce hypermethylation of p16 gene promoter, and decrease p16 gene expression. DNA methylation abnormalities induced by vIL-6 may be one of the mechanisms of oncogenesis.