Circ_0007444 Promotes the Proliferation and Migration of MDA-MB-231 and MCF-7 Cells via the miR-6838-5p/CEP55 Axis
YE Yu, ZHANG Jiawen, ZHU Jianqin, HUA Jingwen, SUN Xiaochun*
This study aims to clarify the molecular mechanism by which circ_0007444 regulates the progression of breast cancer, providing new directions for the clinical diagnosis, treatment and prognosis of breast cancer. Through bioinformatics analysis, differentially expressed circRNAs in breast cancer tissues were screened. The circular structure of circ_0007444 was verified by Sanger sequencing, RNase R experiment and agarose gel electrophoresis. The expression level of circ_0007444 in breast cancer tissues and their matched adjacent normal tissues, normal breast epithelial cells MCF-10A and breast cancer cell lines (MDA-MB-231 and MCF-7) was detected by RT-qPCR (real-time fluorescence quantitative PCR). The localization of circ_0007444 in MDA-MB-231 and MCF-7 cells was determined by RNA FISH experiment. The effects of transfection of si-circ_0007444 on cell proliferation and migration ability were evaluated by plate clone formation assay and Transwell assay. Three public databases (Starbase, CircBank and Circular RNA Interactome) were used to predict and screen the miRNAs targeted by circ_0007444. The targeting interaction between circ_0007444 and miR-6838-5p was verified by dual-luciferase reporter system, and the effects of co-transfection of si-circ_0007444 and miR-6838-5p inhibitor on the proliferation and migration ability of breast cancer cells were explored by plate clone formation assay and Transwell assay. The downstream target genes regulated by miR-6838-5p were predicted by databases (Starbase, miRWalk, miRTar- Base, GEPIA) and the intersection was taken. The expression levels of the downstream target CEP55 (centrosomal protein 55) in MCF-10A, MDA-MB-231 and MCF-7 were detected by RT-qPCR and Western blot. The mRNA and protein expression levels of CEP55 after knockdown and overexpression of miR-6838-5p were verified by RT-qPCR and Western blot. The effects of co-transfection of OE-CEP55 and si-circ_0007444 on the proliferation and migration of breast cancer cells were explored by plate clone formation assay and Transwell assay. The results showed that circ_0007444 was significantly highly expressed in breast cancer cells. Cell function experiments confirmed that knockdown of circ_0007444 could inhibit cell proliferation activity and migration ability. Mechanism studies showed that circ_0007444 promoted breast cancer progression by sponging miR-6838-5p and up-regulating CEP55. In conclusion, this study first clarified the regulatory mode of the circ_0007444/miR-6838-5p/CEP55 axis in breast cancer progression, providing experimental evidence for the development of circular RNA-based molecular targeted therapy.
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