Super-enhancer-dependent transcriptional and m⁶A-mediated post-transcriptional regulation cooperatively drive lncRNA H19 to promote ESCC progression

Epigenetic dysregulation mediated by long non-coding RNAs (lncRNAs), super-enhancers (SEs), and m⁶A modification remains incompletely characterized in esophageal squamous cell carcinoma (ESCC). ChIP-qPCR, dual-luciferase reporter assays and CRISPRi were performed to investigate the transcriptional regulation mechanism of H19 by super-enhancer. The functions and downstream signal transduction mechanisms of the H19 in ESCC were explored by a series of in vitro and in vivo assays, including RNA stability, RIP-qPCR, and MeRIP-qPCR. Furthermore, single-cell RNA sequencing (scRNA-seq) and pseudotime trajectory analysis were used to analyze its correlation with ESCC patient survival. We identified H19, a super-enhancer-driven lncRNA, as a critical oncogenic factor in ESCC, with its overexpression significantly correlating with poor patient prognosis. Mechanistically, the master transcription factor KLF5 binds the super-enhancer region of H19 locus to drive its transcriptional activation. Post-transcriptionally, H19 interacts with the m⁶A methylation reader protein IGF2BP2, which stabilizes H19 in an m⁶A-dependent manner mediated by METTL3. The resulting H19/IGF2BP2 complex upregulates the oncogenic Tweety family member 3 (TTYH3) by enhancing mRNA stability. Functional assays revealed that the H19/IGF2BP2-TTYH3 axis facilitates ESCC malignant progression, while the IGF2BP2 inhibitor CWI1-2 suppresses tumorigenesis. ScRNA-seq combined with pseudotime trajectory analysis further established a significant positive correlation between METTL3/IGF2BP2/TTYH3 expression and malignant phenotypes in ESCC. The H19/IGF2BP2-TTYH3 axis drives ESCC progression, providing valuable prognostic biomarkers and promising therapeutic targets for ESCC treatment.