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抑癌基因PTEN编码产物具有双专一性磷酸酶活性 ,并与细胞骨架张力蛋白同源。PTEN可参与粘着斑的形成和解聚而影响细胞迁移。现用PTEN表达质粒转染SMMC 772 1肝癌细胞 ,研究SMMC 772 1细胞运动能力的变化及PTEN与粘着斑激酶 (FAK)酪氨酸磷酸化水平之间的关系。PTEN过表达能够显著抑制细胞在Fn基质上的活动 :细胞在Fn基质上的迁移下降了 35 % ;在 30min和 6 0min两个时间点 ,Fn基质上细胞铺展分别降低了 2 9%和 2 6 % ;而在多聚赖氨酸基质上细胞铺展并没有变化。运用免疫沉淀和Western印迹方法 ,分析FAK及其酪氨酸磷酸化水平 ,发现PTEN过表达不影响FAK表达 ,但显著降低Fn诱导的FAK酪氨酸磷酸化水平 ,两者水平呈负相关。流式细胞仪分析细胞周期结果表明 ,PTEN抑制细胞 ,S期细胞下降了 16 %。上述结果提示 ,PTEN抑制肝癌细胞迁移铺展和增殖 ;PTEN对细胞运动的影响可能通过调节FAK酪氨酸磷酸化水平而实现。
The tumor suppressor PTEN encoding product has bispecific phosphatase activity and is homologous to the cytoskeletal tension protein. PTEN can participate in the formation and disaggregation of adhesion spots and affect cell migration. The current PTEN expression plasmids were transfected into SMMC 772 1 hepatoma cells to study the changes of motor ability of SMMC 772 1 cells and the relationship between PTEN and focal adhesion kinase (FAK) tyrosine phosphorylation. Overexpression of PTEN significantly inhibited the activity of the cells on the Fn matrix: the migration of the cells on the Fn matrix decreased by 35%; at 30min and 60min, the cell spreading on the Fn matrix decreased by 29% and 26% %; While in the polylysine matrix cell spreading did not change. Immunoprecipitation and Western blotting were used to analyze FAK and tyrosine phosphorylation levels. It was found that overexpression of PTEN did not affect FAK expression, but significantly decreased Fn-induced tyrosine phosphorylation of FAK. The levels of FAK and tyrosine phosphorylation were negatively correlated. Flow cytometry analysis of cell cycle results showed that PTEN inhibited cells, S-phase cells decreased by 16%. The above results suggest that PTEN can inhibit the migration and proliferation of hepatocellular carcinoma cells. The effect of PTEN on cell motility may be mediated by the regulation of tyrosine phosphorylation of FAK.