IDH1-dependent m6A methylation defines transcriptomic heterogeneity in glioma.

Gliomas are biologically heterogeneous brain tumors with marked differences in clinical behavior based on the IDH1 mutation status. While epigenetic dysregulation is well characterized, the contribution of RNA modifications, particularly N6-methyladenosine (m6A), remains underexplored. Using direct RNA nanopore sequencing of patient-derived gliomas, we generated the first isoform-resolved m6A maps across IDH1-mutant and wild-type tumors. IDH1-mutant gliomas exhibited globally elevated m6A methylation, along with increased expression of methyltransferases (METTL3, METTL14) and stabilizing readers (YTHDF3). In contrast, wild-type glioblastomas showed enhanced expression of m6A erasers (ALKBH5, FTO) and RNA decay factors (YTHDF2). These subtype-specific differences in m6A architecture impacted transcript stability, isoform usage, and gene expression. Isoform-level analyses revealed stronger prognostic associations than gene-level parameter, including for IGF2BP2-202, PUF60-202, and GLUL-203. Our study establishes m6A as a critical, subtype-specific layer of RNA regulation in glioma with clinical and therapeutic implications.