The FTO/miR-181b-3p/ARL5B signaling pathway regulates cell migration and invasion in breast cancer

Abstract

Background: N6-methyladenosine (m⁶ A) RNA modification has been demonstrated to be a significant regulatory process in the progression of various tumors, including breast cancer. Fat mass and obesity-associated (FTO) enzyme, initially known as the obesity-related protein, is the first identified m⁶ A demethylase. However, the relationship between FTO and breast cancer remains controversial. In this study, we aimed to elucidate the role and clinical significance of FTO in breast cancer and to explore the underlying mechanism.

Methods: We first investigated the expression of FTO in breast cancer cell lines and tissues by quantitative reverse transcription-PCR (qRT-PCR), Western blotting, and immunohistochemistry. Wound healing assay and Transwell assay were performed to determine the migration and invasion abilities of SKBR3 and MDA-MB453 cells with either knockdown or overexpression of FTO. RNA sequencing (RNA-seq) was conducted to decipher the downstream targets of FTO. qRT-PCR, luciferase reporter assay, and Western blotting were employed to confirm the existence of the FTO/miR-181b-3p/ARL5B axis. The biological function of ADP ribosylation factor like GTPase 5B (ARL5B) in breast cancer cells was evaluated by wound healing assay and Transwell invasion assay.

Results: High FTO expression was observed in human epidermal growth factor receptor 2 (HER2)-positive breast cancer, predicting advanced progression (tumor size [P < 0.001], nuclear grade [P = 0.001], peritumoral lymphovascular invasion [P < 0.001), lymph node metastasis [P = 0.002], and TNM stage [P = 0.001]) and poor prognosis. Moreover, FTO promoted cell invasion and migration in vitro. Mechanistically, RNA-seq and further confirmation studies suggested that FTO up-regulated ARL5B by inhibiting miR-181b-3p. We further verified that ARL5B also displayed carcinogenic activity in breast cancer cells.

Conclusion: Our work demonstrated the carcinogenic activity of FTO in promoting the invasion and migration of breast cancer cells via the FTO/miR-181b-3p/ARL5B signaling pathway.

Keywords: ARL5B; Fat mass and obesity-associated (FTO) enzyme; breast cancer; disease-free survival; human epidermal growth factor receptor 2; m6A modification; metastasis; miR-181b-3p; overall survival.