Cheng-cao Sun¹, Shu-jun Li², Dejia Li²

¹Department ofOccupational and Environmental Health, School of Public Health, WuhanUniversity, ²Department ofOccupational and Environmental Health, Wuhan University

Objective:Hsa-miRNA-206 (miR-206), highly expressedinskeletal muscle, has recently been discovered to have anticancer propertiesindifferent tissues. However, the role of miR-206 on lung cancer is still ambiguous.In this study, we investigated the role of miR-206 on the development of lungcancer. Method: QRT-PCR is used to examine the expression ofmiR-206 in lung cancer tissues and its pair-matched normal lungtissues. Western-blot and immunofluorescence as well as qRT-PCR are usedto access the expression of oncogene c-MET, Bcl2, cyclin D1, cyclin D2,p57, MMP-9. Cell growth was measured using CCK8 assay, colony formation andBrdu immunofluorescence assay. Wound healing assay in vitroand transwell migration/invasion Assay are used to access the role ofmiR-206 on non-small lung cancer cell metastasis. Hoechst 33342staining, caspase-3 activity assay, annexin V-FITC/PI analysisand flow cytometry are used to messure the role of miR-206 onNSCLS cell apoptosis. Luciferase reporter assay is used to examinethe directly inhibitory role of miR-206 on 3'-UTR of MET and BCL2. Result: Theresults indicated that miR-206 expression was suppressed in lung cancer tissuesand very low levels were found in non-smallcell lung cancer (NSCLS) cellliness. Transient transfection of miR-206 into cultured A549 and SK-MES-1 cellsled to significant decrease in cell growth, migration, invasion and colonyformation, and promoted cell apoptosis. Using bioinformatics, we identifiedputative miR-206 binding sites within the 3'-untranslatedregion (3'-UTR) of thehuman c-Met and Bcl2 mRNA. The expression of c-Met and Bcl2 proteins were shownto be down-regulated after treated with miR-206 by subsequent Western blot andqRT-PCR analysis. Conversely, up-regulation of c-Met and Bcl2 were confirmedintissue samples of human lung cancer, with its level inversely correlated withmiR-206 expression. In addition, miR-206 also decreased the gene expression ofMMP-9, CCND1 and CCND2 while increased the gene expression of p57 (Kip2) inA549 andSK-MES-1 cells. Conclusion: Taken together, our resultsdemonstrated that miR-206 suppressed c-Met and Bcl2 expression in NSCLS andcould function as a potent tumor suppressor in c-Met/Bcl2-over expressingtumors. Inhibition of miR-206 function could contribute to aberrantcellproliferation, migration, invasion and apoptosis, leading to NSCLS development.

Key Words: hsa-miRNA-206  c-Met  Bcl2