METTL3-based epitranscriptomic editing screening identifies functional m6A sites in cancers

Abstract

N⁶-methyladenosine (m⁶A) represents the most abundant internal RNA modification and a key regulator of gene expression. Although individual m⁶A regulators and sites have been linked to cancer, their transcriptome-wide functional landscape remains undefined. Here we developed an epitranscriptomic screening platform based on targeted m⁶A deposition to identify functional modifications in prostate and lung cancer models. The unbiased screens uncovered 222 m⁶A sites that modulate cell proliferation, predominantly in a cell-type-specific manner. Among them, an m⁶A site within CHD9 emerged as a potent tumor-suppressive modification in prostate cancer. Deposition of m⁶A at this site increased CHD9 protein abundance, suppressed cell proliferation and attenuated xenograft growth. Mechanistically, m⁶A at CHD9 enhances translation through YTHDF1 and YTHDF3, promoting CHD9-MYBBP1A interaction in the nucleoplasm, sequestrating MYBBP1A from the nucleolus and activating CDKN1A (p21)-associated tumor-suppressive signaling. Collectively, our study establishes a scalable framework for functional mapping of the m⁶A epitranscriptome and uncovers a mechanistic link between CHD9 m⁶A modification and tumor suppression, paving the way for systematic exploration of other RNA modifications in cancer.