FTO-Driven m6A Demethylation of TNC Promotes Epithelial-Mesenchymal Transition and Gastric Cancer Metastasis

Background/Objectives: Gastric cancer (GC) remains a leading cause of cancer-related mortality, with metastasis being the primary driver of poor prognosis. The extracellular matrix glycoprotein tenascin-C (TNC) is implicated in tumor progression and metastasis, yet its regulatory mechanisms in GC remain poorly understood. Here, we identify RNA N6-methyladenosine (m6A) demethylase fat mass and obesity-associated protein (FTO) as a key modulator of TNC expression, promoting epithelial-mesenchymal transition (EMT) and GC metastasis. Methods: Integrating The Cancer Genome Atlas Stomach Adenocarcinoma (TCGA-STAD) and Gene Expression Omnibus (GEO) datasets (GSE246567, GSE181840, GSE163126, GSM4837878), we demonstrate that TNC is significantly upregulated in GC and correlates with worse clinical outcomes. Functional assays both in vitro and in vivo reveal that TNC knockdown suppresses GC cell proliferation, migration, invasion, and tumor metastasis, while overexpression of TNC rescues these effects. Mechanistically, FTO selectively demethylates TNC mRNA, reducing m6A modification at the 3' untranslated region, thereby enhancing TNC mRNA stability and expression. RNA pull-down assays identify YTH domain family protein 2 (YTHDF2) as the m6A reader that recognizes the methylated site on TNC 3'UTR, and YTHDF2 knockdown partially rescues TNC expression and stability upon FTO loss. m6A RNA immunoprecipitation (MeRIP) and dual-luciferase reporter assays confirm FTO's direct regulation of TNC via m6A demethylation. Furthermore, TNC promotes EMT by activating the phosphoinositide 3-kinase (PI3K)-AKT (protein kinase B) signaling pathway, as demonstrated by rescue experiments using the AKT activator SC79 and the PI3K inhibitor LY294002. Results: High FTO and TNC expression levels in clinical tissue samples correlate with poor patient survival. Conclusions: These findings reveal that the FTO/TNC/EMT axis represents a previously unrecognized epigenetic mechanism driving GC metastasis and suggest that targeting FTO-mediated m6A modification may provide a novel therapeutic strategy for GC patients with high metastatic potential.