Mounting evidence has implicated the RNA m6A methylation catalyzed by METTL3 in a wide range of physiological and pathological processes, including tumorigenesis. The detailed m6A landscape and molecular mechanism of METTL3 in prostate cancer (PCa) remains ill-defined. We find that METTL3 is overexpressed in PCa and correlates with worse patient survival. Functional studies establish METTL3 as an oncoprotein dependent on its m6A enzymatic activity in both AR+ and AR- PCa cells. To dissect the regulatory network of m6A pathway in PCa, we map the m6A landscape in clinical tumor samples using m6A-seq and identify genome-wide METTL3-binding transcripts via RIP-seq. Mechanistically, we discover RRBP1 as a direct METTL3 target in which METTL3 stabilizes RRBP1 mRNA in an m6A-dependent manner. RRBP1 positively correlates with METTL3 expression in PCa cohorts and exerts an oncogenic role in aggressive PCa cells. Leveraging the 3D structural protein-protein interaction between METTL3 and METTL14, we successfully develop two potential METTL3 peptide inhibitors (RM3 and RSM3) that effectively suppress cancer cell proliferation in vitro and tumor growth in vivo. Collectively, our study reveals a novel METTL3/m6A/RRBP1 axis in enhancing aggressive traits of PCa, which can be therapeutically targeted by small-peptide METTL3 antagonists.
