GAB2蛋白促进SK-Mel28黑色素瘤细胞转移机制研究

发布时间：2018-04-18 05:34

本文选题：GAB + 黑色素瘤　；参考：《中华肿瘤防治杂志》2017年14期

【摘要】：目的恶性黑色素瘤主要由皮肤黑色素细胞的恶性转化引起。Grb2协同结合蛋白2(Grb2-associated binding protein-2,GAB2)是一个脚手架蛋白,通过招募受体酪氨酸激酶等膜受体进行信号传递,已经证实GAB2蛋白与多种肿瘤的发生和转移有关,但GAB2蛋白抑制黑色素瘤的机制还不清楚。本研究通过沉默GAB2基因对SK-Mel28细胞转移能力的影响,探讨GAB2基因促进SK-Mel28黑色素瘤细胞转移的细胞信号转导通路变化。方法采用RNA干扰技术沉默GAB2基因表达。采用Matrigel法检测SK-Mel28细胞在沉默GAB2基因表达后48h对细胞转移能力的影响;采用细胞划痕实验检测SK-Mel28细胞在沉默GAB2基因表达后细胞迁移能力的改变;蛋白质印迹法检测PI3K和MAPK信号通路的关键蛋白、pAKT、pERK、pPDK1和β-catenin的蛋白表达水平。结果沉默GAB2基因可使SK-Mel28细胞的转移能力和迁移能力减弱,统计学分析显示,SK-Mel28细胞的转移能力降低了54.5%,与scramble RNA对照组比较差异有统计学意义,t=5.053,P0.001;SK-Mel28细胞在转染GAB2siRNA 18h后细胞的迁移能力减弱,与scramble RNA对照组比较迁移距离减少了43.0%,差异有统计学意义,t=3.877,P=0.001。SK-Mel28黑色素瘤细胞的GAB2蛋白表达水平降低后,PI3K信号通路的关键分子pAKT和pPDK1表达下调,而T-AKT和T-PDK1无明显变化,细胞膜表面粘附因子β-catenin蛋白表达下降。结论 GAB2蛋白的表达水平与SK-Mel28黑色素瘤的转移相关,其机制与GAB2蛋白调控下游的PI3K信号通路并抑制转移相关因子β-catenin有关。
[Abstract]:Objective malignant melanoma is mainly caused by malignant transformation of melanocytes. Grb2 cobinding protein 2(Grb2-associated binding protein-2 GAB2) is a scaffold protein, which is transmitted by recruitment of receptor tyrosine kinase and other membrane receptors.It has been proved that GAB2 protein is involved in the occurrence and metastasis of many kinds of tumors, but the mechanism of GAB2 protein inhibiting melanoma is unclear.In this study, the effect of silencing GAB2 gene on the metastatic ability of SK-Mel28 cells was studied to investigate the changes of signal transduction pathway of SK-Mel28 melanoma cells promoted by GAB2 gene.Methods GAB2 gene expression was silenced by RNA interference technique.Matrigel assay was used to detect the effect of SK-Mel28 cells on the ability of cell metastasis after GAB2 gene expression was silenced for 48 hours, and cell scratch assay was used to detect the change of cell migration ability of SK-Mel28 cells after silencing the expression of GAB2 gene.The protein expression levels of pAKTT pERKN pPDK1 and 尾 -catenin in PI3K and MAPK signaling pathway were detected by Western blotting.Results silencing GAB2 gene decreased the metastasis and migration ability of SK-Mel28 cells.Statistical analysis showed that the metastatic ability of SK-Mel28 cells decreased by 54.5%, which was significantly lower than that of scramble RNA control group. The migration ability of SK-Mel28 cells was decreased after transfection of GAB2siRNA for 18 h.Compared with the control group of scramble RNA, the migration distance was decreased by 43.0. The difference was statistically significant. The expression of GAB2 protein in melanoma cells was significantly decreased, but the expression of pAKT and pPDK1, the key molecules of PI3K signaling pathway, was down-regulated, but T-AKT and T-PDK1 did not change significantly.The expression of adhesion factor 尾-catenin on cell membrane was decreased.Conclusion the expression of GAB2 protein is related to the metastasis of SK-Mel28 melanoma, and its mechanism is related to the regulation of downstream PI3K signal pathway by GAB2 protein and the inhibition of metastasis related factor 尾 -catenin.
【作者单位】：天津大学生命科学学院微生物与免疫学教研室;天津市肿瘤医院血液科;
【基金】：天津市应用基础与前沿技术研究计划(13JCYBJC40800)
【分类号】：R739.5

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