SF3B4 Regulated Alternative Splicing that Produced an Oncogenic Isoform of HOXA11-AS to Promote Glycolytic Reprogramming and ESCC Progression

Abstract

Background Dysregulated splicing factors (SFs) and aberrant alternative splicing (AS) events are involved in tumor progression. However, the AS landscape underlying SFs dysregulation and the further signal transduction network were unraveled in esophageal squamous cell carcinoma (ESCC). This study revealed the biological function of splicing factor 3b subunit 4 (SF3B4) in non-coding RNA AS and glycolytic reprogramming, and proposed a novel diagnostic pannel and therapeutic targets for ESCC. Methords The expression, diagnostic efficiency and prognostic value of SF3B4 were investigated by bioinformatics, real-time fluorescent quantitative PCR and immunohistochemistry assays. The biological functions of SF3B4 in ESCC were analyzed in vivo and in vitro by loss-of-function studies. RNA sequencing, minigene reporter, RNA immunoprecipitation and correlation analysis were performed to elucidate SF3B4-regulated AS isoforms and SF3B4-interaction motif. Seahorse metabolism assays and high performance liquid chromatography-mass spectrometry analysis were conducted to explore the potential molecular mechanism of SF3B4 and downstream AS isoforms in driving ESCC development. Results SF3B4 was significantly up-regulated in ESCC and facilitated cell proliferation, survival, cycle progression and cisplatin resistance. Mechanically, SF3B4 increased proportion of the tumorigenic splicing isoform (HOX-L) of long noncoding RNA homeobox A11 antisense (HOXA11-AS), which resulted in enhanced glycolysis and elevated transcription of glycolytic enzyme PKM2, ENO1, HK2, GLUT1, LDHA and PGK1, through promoting phosphorylation of β-catenin at serine 675 and activation of Wnt pathway. Remarkably, inhibition of glycolysis reversed the malignant phenotype induced by the SF3B4-HOX-L axis. Moreover, the RNA levels of SF3B4 and HOX-L were positively correlated with ESCC tumor volume, and high SF3B4 expression demonstrated significant poor survival for ESCC patients. Additionally, the combination of SF3B4 and HOXA11-AS expression also showed good diagnostic performance. Conclusions These findings highlighted the oncogenic role of the SF3B4-HOX-L- Wnt-β-catenin-glycolytic enzyme axis in ESCC development, and proposed SF3B4 and HOX-L splicing isoform as novel therapeutic targets and diagnostic biomarkers for ESCC.