miR-218 调控FAK-Slit/ Bmi-1-TGF-β 信号通路抑制脑胶质瘤增殖的机制研究

胶质瘤治疗的重要策略之一是抑制其增殖。我们证实，miR-218通过下调Bmi1抑制胶质瘤增殖。但是，过表达miR-218抑制胶质瘤增殖的能力远远超过Bmi1 siRNA，提示Bmi1只可能是其靶基因之一，还需进一步阐明miR-218的分子机制。蛋白质谱技术联合RIP-chip对miR-218靶基因的筛选显示，FAK、TGF-β亦可能是其靶基因。我们的预实验显示，FAK调控Bmi1，可能是Slit2/Bmi1通路的上游分子，而TGF-β是Slit2/Bmi1的下游分子。我们提出理论假说，miR-218多环节调控FAK-Slit/Bmi1-TGF-β信号通路抑制胶质瘤细胞增殖。本研究将行定点突变、ChIP、EMSA等实验研究FAK对Bmi1的调控机制，开展干预、拮抗实验研究miR-218多环节调控FAK-Slit/Bmi1-TGF-β通路抑制胶质瘤增殖，将为胶质瘤治疗选择新靶点提供理论指引。

As to glioma therapy, one of the key factors is inhibition of glioma proliferation. Recent studies in our lab showed that, miR-218 inhibits glioma invasion, migration, and proliferation as well as self-renewal of cancer stem-like cells by targeting the polycomb group gene Bmi1. However, transfection with the miR-218-expressing vector in U251MG cells inhibited glioma cell proliferation much better than those observed after knockdown of Bmi1 by siRNA.The data are to be discussed, and it is proposed that the molecular mechanism of miR-218 should be further elucidated, and Bmi1 might only be one of the target genes of miR-218. Further targeting and screening of miR-218 target genes with RIP-chip and iTRAQ indicated that the genes of FAK and TGF-β might also be the candidates for miR-218. The gene of FAK regulates expression of Bmi1 in gliomas, which indicates that FAK is an upstream regulatory molecule of Slit/Bmi1 signaling pathway. Additionally, TGF-β is seen as the downstream molecule of Slit/ Bmi1 pathway. Based on these results, we propose a new hypothesis that miR-218 inhibits glioma cell proliferation through regulating multiple target genes in FAK-Slit/Bmi1-TGF-β signaling pathway. To test such a hypothesis, we will use site-directed mutagenesis，ChIP, and EMSA to study the transcriptional regulatory mechanism of FAK on Bmil. And then we will use gain-of-function and loss-of-function approaches to investigate the influence of miR-218 on glioma cell proliferation through directly regulating multiple target genes in vitro and in vivo. This study will provide a theoretical basis for the application of miR-218 to new targets in gene therapy of gliomas.