METTL16 suppresses ferroptosis in cholangiocarcinoma by promoting ATF4 via m⁶A modification

Background: N6-methyladenosine (m⁶A) modification is the most common post-transcriptional modifications, which is critical for the metabolism of ferroptosis-related RNAs. Yet, the impact of m⁶A modification on ferroptosis in cholangiocarcinoma (CC) is far from clear. Methods: Public databases and tissue arrays were applied to explore the clinical relevance of METTL16 in CC. Then, the effects of METTL16 on growth and ferroptosis were studied in vitro and in vivo. Mechanistically, RNA-sequencing, methylated RNA immunoprecipitation, dual-luciferase reporter assays and RNA stability assays were used to identify the METTL16/ATF4 axis in ferroptosis in CC. Results: Clinically, we find that METTL16 is overexpressed and associated with a poor prognosis in patients with CC. Functionally, METTL16 protects against ferroptosis by maintaining mitochondrial homeostasis, including mitochondrial structure, membrane potential and energy products. It also decreases cellular metabolism of Fe²⁺ and lipid peroxide, thereby promoting cell growth in vitro and in vivo. Mechanistically, ATF4 is a novel target of METTL16 and METTL16 enhances the m6A level and expression of ATF4 mRNA by inhibiting its decay, which further prevented ferroptosis in CC via m6A modification. Conclusions: Our findings highlighted the role of METTL16/ATF4 in ferroptosis, which sheds light on potential therapeutic strategies for CC.