Silencing of hsa_circ_0004771 inhibits proliferation and induces apoptosis in breast cancer through activation of miR-653 by targeting ZEB2 signaling pathway.

Rong Xie,Hui Jiang,Xianmin Zhu,Jing Tang

doi:10.1042/bsr20181919

Abstract

Background: Circular RNAs (circRNAs) have been reported as the competing endogenous RNAs (ceRNAs) to sponge microRNAs (miRNAs) implicating in the initiation and progression of breast cancer. However, the functions of circRNAs in breast cancer have not been completely clarified. In the present study, we aimed to identify differentially expressed circRNAs in breast cancer tumor tissues, and their roles and downstream targets were investigated in the progression of breast cancer. Methods: High-throughput circRNA sequencing was performed to detect the differentially expressed circRNAs. The CCK-8 and flow cytometry were performed to measure the cell viability and apoptosis in breast cancer cells. Gene and protein expression were assayed by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and Western blotting, respectively. Results: hsa_circ_0004771 and Zinc finger E-box binding homeobox 2 (ZEB2) expression levels were up-regulated and positively correlated in breast cancer tumor tissues. In addition, the expression levels of miR-653 were reduced in breast cancer tumor tissues. We also found that hsa_circ_0004771 functioned as a sponge of miR-653 to inhibit its expression. miR-653 as a post-transcriptional regulator down-regulated the expression of ZEB2 by binding to its 3′-UTR. Interestingly, a significant inverse correlation was observed between miR-653 and hsa_circ_0004771 or ZEB2 expression in breast cancer tumor tissues. Knockdown of hsa_circ_0004771 and ZEB2 served as equally authentic of miR-653 mimics to induce growth inhibition and apoptosis in breast cancer cells. Conclusion: Hsa_circ_0004771/miR-653/ZEB2 regulatory feedback revealed a new molecular mechanism in the pathogenesis of breast cancer, which might provide novel therapeutic targets for the treatment of breast cancer.

Highlights

Circular RNAs as a class of endogenous noncoding RNA are predominantly generated in eukaryotes from four cyclized models, including back-spliced exons, circular intronic RNAs, exon–intron circular RNA (circRNA) and intergenic circRNAs and characterized by single-stranded, covalently closed circular molecules without 5 –3 polarity and polyadenylated tail, reflecting that circRNAs can tolerate the digestion of exonucleases [1,2]
To validate the results of high-throughput sequencing, we selected top five circRNAs for further assessment using reverse transcription-quantitative polymerase chain reaction (RT-qPCR), and the findings demonstrated that RT-qPCR results were consistent with sequencing data that top five circRNAs were markedly increased in breast cancer tumor tissues compared with corresponding nontumorous tissues (Figure 1B–F)
We found that hsa circ 0004771 functioned as a sponge of miR-653 to inhibit its expression. miR-653 as a post-transcriptional regulator down-regulated the expression of Zinc finger E-box binding homeobox 2 (ZEB2) by binding to its 3 -UTR

Summary

Introduction

Circular RNAs (circRNAs) as a class of endogenous noncoding RNA are predominantly generated in eukaryotes from four cyclized models, including back-spliced exons, circular intronic RNAs, exon–intron circRNAs and intergenic circRNAs and characterized by single-stranded, covalently closed circular molecules without 5 –3 polarity and polyadenylated tail, reflecting that circRNAs can tolerate the digestion of exonucleases [1,2]. Functional studies have been validated that circRNAs perform multiple functions, including the competing endogenous RNAs (ceRNAs) to sponge microRNAs (miRNAs), interaction with RNA-binding proteins to regulate cell cycle and proliferation and the management of gene transcription and protein translation [9,10,11,12,13]. We aimed to identify differentially expressed circRNAs in breast cancer tumor tissues, and their roles and downstream targets were investigated in the progression of breast cancer. Results: hsa circ 0004771 and Zinc finger E-box binding homeobox 2 (ZEB2) expression levels were up-regulated and positively correlated in breast cancer tumor tissues. Conclusion: Hsa circ 0004771/miR-653/ZEB2 regulatory feedback revealed a new molecular mechanism in the pathogenesis of breast cancer, which might provide novel therapeutic targets for the treatment of breast cancer

Objectives

Methods

Results

Conclusion