EB病毒潜伏膜蛋白1通过Survivin介导细胞增殖和抑制细胞凋亡

EB Virus Latent Membrane Protein 1 Initiates Cells Proliferation and Inhibits Apoptosis via Survivin Triggering Signal-Pathway

利用间接免疫荧光、基因转染、抗体剔除 (Ab knock out)、细胞平板集落形成、流式细胞术以及半胱氨酸天冬酰胺酶 (caspase3)活性检测等方法 ,从survivin核移位、Rb磷酸化、细胞周期演进、细胞克隆形成和细胞凋亡等方面 ,探讨EB病毒潜伏膜蛋白 1(LMP1)调控细胞增殖和细胞凋亡双重效应的分子机制 .结果发现 ,LMP1表达介导survivin核移位 ,促进细胞Rb磷酸化增加 ,S期细胞数显著增加 ;LMP1通过survivin促进细胞克隆形成 .用Ab knock out阻断survivin核移位和survivin反义核酸抑制survivin表达时 ,Rb磷酸化水平降低 ,S期细胞减少 ,抑制LMP1介导的细胞增殖 ,活化细胞caspase 3,诱导细胞凋亡 .结果提示 。

To probe the molecular mechanism of EB virus latent membrane protein 1 triggering pleiotropic biological effect of apoptosis and cell proliferation, the sub cellular distribution of survivin, the phosphorylation level of Rb, cell cycle, apoptosis and cell proliferation were detected using indirect immunofluoresence assay, gene transfection, Ab knock out, colony formation, flow cytometry, caspase 3 assay and Western blotting. The results showed that the expression of survivin and Rb phosphorylation was higher than that without LMP1 expression and LMP1 expression triggered the translocation of survivin into the nucleus and Rb phosphorylation. The phosphorylation level of Rb was decreased when anti sense survivin was induced into Tet on LMP1 HNE2 by the gene transfection. The number of cells in S stage of cell cycle was decreased when survivin translocation into nucleus was blocked using Ab knock out and transcription of survivin was blocked using gene transfection. Low level expression of LMP1 could increase cell proliferation, which could be inhibited by anti sense survivin. Survivin triggered by LMP1 blocked caspase 3 activation and inhibited apoptosis. All these results indicated that LMP1 promoted Rb phosphorylation, cell cycle entry and triggered cell proliferation via activating survivin expression. The expression of survivin triggered by LMP1 could promote cell proliferation and inhibit cell apoptosis.