Objective: To determine the inhibitory effect of miRNA-381 on renal carcinoma invasion and to explore the underlying mechanisms.
Methods: After up-regulation of miRNA-381, the inhibitory effect of miR-381 on cell invasion was investigated. We screened the target genes of miRNA-381 in a database (starBase) through combination of five programs including targetscan, picTar, RNA22, PITA and miRanda. Then, the predicted targeting genes were verified by the dual luciferase reporter assay. We also examined the expression of miRNA-381 and its target genes in renal cancer cells and tissues.
Results: Transfection and up-regulation of miRNA-381 resulted in a significant decrease in trans-membrane cell numbers and the ability of renal cell invasion. Bioinformatics analysis showed that CREB binding protein (CBP), β-catenin and lymphoid enhancer binding factor-1 (LEF-1) were the potential targets of miRNA-381. In the luciferase reporter gene system, co-transfection of miRNA-381 with the 3'UTR of wild-type target gene led to a significant decrease in luciferase activity. The expression of miRNA-381 was decreased in various renal cancer cells, and it was particularly lower in highly metastatic cell lines (786-OHM). On the contrary, the expression levels of miRNA-381 target genes (CBP, β-catenin and LEF-1) were significantly increased in cells and tissues.
Conclusion: MiRNA-381 can inhibit cell invasion in renal cancer by block the function of CBP, β-catenin and LEF-1.
目的：研究miRNA-381对肾癌侵袭的抑制作用，并探索其机制。方法：上调miRNA-381表达后，观察miRNA-381对肾癌细胞侵袭的抑制作用。通过miR数据库starBase研究miRNA-381靶基因，再通过双荧光素酶实验验证。分析miRNA-381及相应靶基因在细胞和组织水平的表达丰度。结果：转染miRNA-381后，其表达增加，跨膜细胞数目显著下降，细胞侵袭力受抑制。生物信息学分析显示miRNA-381靶基因为CREB绑定蛋白(CREB binding protein，CBP)、β-catenin和淋巴增强因子-1(lymphoid enhancer binding factor-1，LEF-1)；包含野生型靶基因3'-非编码区载体组的荧光素酶活性明显降低。实时定量PCR显示肾癌细胞中miRNA-381低表达，在高转移潜能肾癌细胞株786-OHM中表达更低；相反，在细胞、组织水平miRNA-381靶基因CBP，β-catenin与LEF-1表达均增高。结论：MiRNA-381可抑制肾癌细胞的侵袭能力，其机制可能是通过CBP，β-catenin和LEF-1实现的。.