METTL3-Mediated m6A Modification of COL5A2 Inhibits Ferroptosis Through an IGF2BP3-dependent Mechanism in Gastric...

IntroductionMETTL3, the core enzyme of m6A methylation, influences gastric cancer (GC) progression, but its role in ferroptosis remains unclear. This study investigated the regulatory role of METTL3 in GC ferroptosis and its underlying mechanism via the downstream target gene collagen type V alpha 2 chain(COL5A2).MethodGC cell lines (AGS, MKN-45) and normal GES-1 cells were used. Ferroptosis was induced by erastin and RSL-3. METTL3 overexpression/knockdown models were constructed to assess ferroptosis indicators, including lipid reactive oxygen species(ROS), malondialdehyde(MDA), Fe²⁺, glutathione(GSH) and malignant phenotypes. m6A sites were predicted and validated by MeRIP-qPCR. m6A readers were knocked down to identify effectors, and mRNA stability was evaluated via RIP and actinomycin D assays. The focal adhesion kinase(FAK)/mitogen-activated protein kinase(MAPK)/extracellular signal-regulated kinase(ERK)pathway and ferroptosis-related proteins were detected by Western blot, with pathway involvement confirmed using the FAK inhibitor PF-573228. To validate the in vitro findings, a nude mouse xenograft tumor model was established.ResultsMETTL3 was highly expressed in GC cells and suppressed by ferroptosis inducers. METTL3 overexpression inhibited ferroptosis (reducing ROS, MDA, Fe²⁺, increasing GSH) and promoted proliferation and migration. COL5A2 was positively correlated with METTL3. MeRIP-qPCR confirmed that METTL3 mediates m6A modification of COL5A2's 3'-UTR. RIP and ActD assays demonstrated that the m6A reader insulin-like growth factor 2 mRNA-binding protein 3(IGF2BP3) maintains COL5A2 mRNA stability. IGF2BP3 knockdown promoted ferroptosis and apoptosis, which were reversed by COL5A2 overexpression. Furthermore, COL5A2 overexpression activated the FAK/MAPK/ERK pathway, upregulated GPX4, and inhibited ferroptosis; these effects were counteracted by the FAK inhibitor. In vivo, METTL3 overexpression promoted tumor growth, reduced 4-HNE levels, and regulated ferroptosis and FAK/MAPK/ERK pathway proteins, all of which were reversed by COL5A2 knockdown.ConclusionMETTL3 upregulates COL5A2 expression through m6A modification, with IGF2BP3 maintaining COL5A2 mRNA stability. This subsequently activates the FAK/MAPK/ERK signaling pathway, inhibits ferroptosis, and promotes GC progression. The METTL3/IGF2BP3/COL5A2 axis represents a promising therapeutic target for GC treatment.