MicroRNA-154 inhibits the growth and metastasis of gastric cancer cells by directly targeting MTDH

Abstract

MicroRNAs (miRNAs) are a group of non-protein-coding, highly conserved single-stranded RNA molecules. The abnormal expression of miRNAs has been demonstrated to have an important function in the carcinogenesis and progression of gastric cancer. microRNA-154 (miR-154) has been reported to be downregulated in non-small cell lung, colorectal and prostate cancer. However, the expression and roles of miR-154 in gastric cancer remain to be established. The present study measured the expression levels of miR-154 in gastric cancer tissues and cell lines. miR-154 was found to be significantly downregulated in gastric cancer tissues and cell lines. In addition, functional studies indicated that the overexpression of miR-154 inhibited the proliferation, migration and invasion of gastric cancer cells. Using TargetScan, a dual luciferase reporter assay, reverse transcription-quantitative polymerase chain reaction and western blot analysis, metadherin (MTDH) was revealed as a novel miR-154 target. In addition, knocking down MTDH lead to a similar effect as overexpressing-154 in gastric cells. The present findings indicate that miR-154 was downregulated in gastric cancer, and inhibited tumor behaviors of gastric cancer cells partially through the downregulation of MTDH. Therefore, the miR-154/MTDH axis may be a novel therapeutic to treat patients with gastric cancer.

Keywords: gastric cancer; growth; metadherin; metastasis; microRNA-154.

Figure 1.

Relative expression of miR-154 in GC. (A) Relative expression of miR-154 in GC tissues and matched non-neoplastic gastric tissues. (B) miR-154 was downregulated in the four GC cell lines (SGC-7901, AGS, MKN-1 and BGC-823) compared with that in the normal gastric epithelium GES-1 cell line. *P<0.05 compared with their respective controls. miR-154, microRNA-154; GC, gastric cancer.

Figure 2.

Over-expression of miR-154 inhibited the tumor behavior of GC cells. (A) Reverse transcription-quantitative polymerase chain reaction was performed to measure relative miR-154 expression in GC cells following transfection with miR-154 mimics or NC. (B) Cell proliferation assays were performed to evaluate the proliferation of SGC-7901 and MKN-1 cells following transfection with miR-154 mimic or NC. (C) Cell migration and invasion assays were performed in SGC-7901 and MKN-1 cells following transfection with miR-154 mimic or NC (magnification, × 200). *P<0.05 compared with their respective controls. miR-154, microRNA-154; GC, gastric cancer; NC, negative control.

Figure 3.

miR-154 directly targeted MTDH. (A) The putative miR-154 binding sites in the 3′UTR of MTDH was shown. Mutation was generated on the MTDH 3′UTR sequence in the complementary site for the seed region of miR-154. (B) The relative luciferase activities of the PGL3-MTDH-3′UTR Wt were significantly decreased when miR-154 mimics was co-transfected. However, the luciferase activities of PGL3-MTDH-3′UTR Mut were unaffected by co-transfection with miR-154 mimics. (C) The expression of MTDH mRNA was determined by reverse transcription-quantitative polymerase chain reaction in GC cells following transfection with miR-154 mimics or NC. (D) The expression levels of MTDH protein were measured by western blot analysis in GC cells following transfection with miR-154 mimics or NC. β-actin was used as a control. *P<0.05 compared with their respective controls. miR-154, microRNA-154; MTDH, metadherin, 3′UTR, 3β untranslated region; Wt, wild type; Mut, mutant; GC, gastric cancer; NC, negative control.

Figure 4.

Knockdown of MTDH inhibited proliferation, migration and invasion of GC cells. (A) Expression levels of the MTDH protein were measured by western blot analysis following transfection with the MTDH siRNA or siRNA control. β-actin was used as a control. (B) MTDH siRNA inhibited the proliferation of GC cells. (C) MTDH siRNA inhibited the migration and invasion capacity of GC cells. *P<0.05 compared with their respective controls. MTDH, metadherin; GC, gastric cancer; siRNA, small interfering RNA; ctrl, control.