MiR-675 is frequently overexpressed in gastric cancer and enhances cell proliferation and invasion via targeting a potent anti-tumor gene PITX1

Abstract

Background: Gastric cancer (GC) is a common malignancy around the world. Irregular expression of microRNAs (miRNAs) contributes to the progression of malignancies. Our study illustrated that miR-675 facilitates GC proliferation and invasion via targeting paired-like homeodomain transcription factor 1 (PITX1) and promoting epithelial-mesenchymal transition (EMT) as well as Wnt/β-catenin signaling pathway.

Methods: We collected the RNA-seq data of GC and normal stomach tissues from TCGA database to analyze the expression of miR-675 and PITX1. Kaplan-Meier plotter on line tool was used to analyze the association between miR-675 or PITX1 expression and the overall survival of GC patients. The biological function of miR-675 in GC cells was evaluated via altering its expression using miR-675 agomiR or antamiR. Dual-luciferase reporter assay was applied for verifying whether miR-675 could direct bind to 3'UTR of PITX1. Rescue assays were applied for characterizing the effects of miR-675/PITX1 axis on GC growth and invasion. Western blot was performed to evaluate the protein expression levels of PITX1, EMT-related and Wnt signaling-related proteins.

Results: Our results showed that miR-675 is up-regulated and predictive of worse prognosis in GC patients. Overexpression of miR-675 in AGS cells notably promoted cell proliferation, migration and invasion, whilst down-regulation of miR-675 in SGC-7901 cells gained the opposite results. PITX1 is down-regulated in GC and identified as a direct target of miR-675. Overexpression of PITX1 in AGS cells reverses cell viability and invasion that enhanced by miR-675 up-regulation. Conversely, depletion of PITX1 in SGC-7901 cells rescues cell viability and invasion that inhibited by miR-675 down-regulation. Western bolt results revealed that miR-675 positively regulated EMT and Wnt/β-catenin signaling pathway in GC cells via targeting PITX1.

Conclusions: Our study emphasized the functional mechanism of miR-675 in GC and intimated that miR-675/PITX1 axis possibly affects proliferative and invasive properties of GC cells via regulating EMT and Wnt/β-catenin signaling pathway. Furthermore, miR-675 and PITX1 may be served as early diagnostic markers as well as therapeutic targets for GC.

Keywords: EMT; Gastric cancer; Invasion; PITX1; miR-675.