hsa-miR-125a-3p是RhoA-肌动蛋白途径的负性调节因子

hsa- miRNA- 125a- 3p Is a Negative Regulator of Rho A- actomyosin Pathway

为了探讨hsa-miR-125a-3p调控肺癌细胞侵袭的作用机制,分别检测了转染正义和反义hsa-miR-125a-3p的人肺癌A549细胞中Rho A mRNA和蛋白水平表达情况,结果显示,两种方式处理的细胞中Rho A mRNA均无明显变化,但Rho A蛋白浓度分别下降和增加;再用包含Rho A 3’-未翻译区(3’-UTR)荧光素酶报告基因与hsa-miR-125a-3p mimics共转染发现,当hsa-miR-125a-3p异常表达时,报告基因活性降低。进一步的实验显示,hsa-miR-125a-3p下调可诱导A549细胞侵袭;Rho抑制剂CT04阻断Rho A后,转染反义或正义hsa-miR-125a-3p的A549细胞侵袭率均无变化;转染正义的hsa-miR-125a-3p的A549细胞中活化Rho A(Rho A-GTP)、Rho A和肌动蛋白丝聚集表达均降低;相反,当转染反义的hsa-miR-125a-3p时,Rho A-GTP、Rho A和肌动蛋白丝聚集表达均增强。上述研究表明,hsa-miR-125a-3p可能通过Rho A-肌动蛋白途径影响肺癌细胞系A549的侵袭能力。

To study how hsa - miR - 125a - 3p enhances the invasive ability of lung cancer cells, we detected RhoA mRNA and protein by RT - PCR and Western blot respectively after overexpression of hsa - miR - 125a -3p by transfection of sense - miR - 125a - 3p or knockdown of hsa - miR - 125a - 3p by trandfection of antisense - miR- 125a- 3p. The resulted showed that RhoA mRNA remained unchanged, but RhoA protein levels are decreased and increased respectively. Luciferase reporter constructs containing the RhoA 3＇ UTR demonstrated reduced reporter activity after ectopic expression of hsa - miR - 125a -3p. Furthermore, A549 ceils demonstrated reduced invasion capacity after treatment with the Rho inhibitor CT04 by Transwell. Ultimately, an increased intracelluar concentration of RhoA - GTP protein and RhoA protein in A549 cells leads to high accumulation of actin fil- aments when antisense -miR -125a -3p was transfected. Contrarily, decreased intracelluar concentration of RhoA - GTP protein and RhoA protein in A549 cells leads to low accumulation of actin filaments when sense - miR - 125a-3p was transfected. In conclusion, hsa- miR- 125a- 3p regulated the invasion of A549 ceUs probably through the RhoA- actomyosin pathway.