In vitro study of piwi interaction RNA-31106 promoting breast carcinogenesis by regulating METTL3-mediated m6A RNA methylation.

Abstract

Breast cancer is the most common gynecological malignancy and the leading cause of cancer-related deaths in women. P-element induced wimpy testis (PIWI)-interacting RNAs (piRNAs) are novel non-coding RNAs whose abnormal expressions have been closely associated with multiple cancers. This study explored the roles and possible mechanisms of piRNA-31106 in breast cancer. The expression of piRNA-31106 in breast cancer tissues and cells was detected by reverse transcription polymerase chain reaction (RT-PCR). The pcDNA vector containing piRNA-31106 (pcDNA-piRNA-31106) and a short hairpin (sh)RNA containing piRNA-31106 (shRNA-piRNA-31106) were used to interfere with piRNA-31106 expression in breast cancer cells. The effects on cell proliferation, apoptosis/cell cycle, invasion, and metastasis were detected via Cell Counting Kit-8 (CCK-8), flow cytometry, transwell assays, and scratch tests, respectively. The protein expressions of murine double minute 2 (MDM2), cyclin-dependent kinase 4 (CDK4), and cyclinD1 were detected by Western blot analysis. The N6-methyladenosine (m6A) RNA methylation level and the binding relationship between piRNA-31106 and METTL3 were analyzed. The role of METTL3 in the regulation of breast cancer by piRNA-31106 was further analyzed by using small interfering (si)RNA targeting METTL3. PiRNA-31106 was highly expressed in breast cancer tissues and cell lines MDA-MB-231 and MCF-7. Overexpression of piRNA-31106 promoted the viability, invasion, and migration of breast cancer, inhibited apoptosis, and promoted the expressions of MDM2, CDK4, and cyclinD1. Inhibition of piRNA-31106 showed the opposite effect. In addition, piRNA-31106 promoted the m6A methylation levels and facilitated methyltransferase-like 3 (METTL3) expression in MDA-MB-231 and MCF-7 cells. RNA immunoprecipitation (RIP) assays confirmed the binding relationship between piRNA-31106 and METTL3. Further experiments demonstrated that si-METTL3 could inhibit the regulatory effects of piRNA-31106 on breast cancer. PiRNA-31106 was significantly highly expressed in breast cancer and could promote breast cancer progression by regulating METTL3-mediated m6A RNA methylation.