NONO regulates m5C modification and alternative splicing of PTEN mRNAs to drive gastric cancer progression.

Abstract

Background: The effect of m⁵C modification on oncogene mRNAs has been well studied, while little is known about its influence on mRNAs of tumor suppressor genes (TSGs). Early studies showed PTEN, a key TSG, undergoes alternative splicing (AS) in cancers, however, the underlying regulatory mechanism remains elusive.

Methods: We analyzed tissue microarrays and transcriptomic data derived from gastric cancer, with an emphasis on RNA splicing and m⁵C regulators. To unravel the role of NONO in GC, we employed RNA sequencing, RNA-Bis-Seq, RNA immunoprecipitation, RNA in situ hybridization, and Minigene reporter assay with NONO knockdown cells. The clinical relevance was validated using CDX models and human tissue microarrays.

Results: Analysis of publicly available datasets and immunohistochemistry assay of tissue microarrays containing 40 GC tissues showed NONO was upregulated in GC and contributed to poor prognosis. In vitro and in vivo experiments indicated a positive regulatory role of NONO in terms of cell proliferation, migration, and invasion of GC. Mechanically, NONO interacted directly with PTEN pre-mRNA and recruited the RNA m⁵C methyltransferase NSUN2 via RNA-recognition motif (RRM) domains, altering the mRNA methylation pattern across PTEN pre-mRNA. The oncogenic role of NONO/NSUN2/PTEN axis in GC progression was further confirmed with pre-clinical experiments and clinical data.

Conclusion: Here, we revealed NONO-regulated AS of PTEN mRNA in an m⁵C-dependent manner, resulting in the downregulation of PTEN expression in gastric cancer (GC).This study unveils a novel regulatory mechanism of tumor suppressor gene inactivation mediated by m⁵C modification and related alternative splicing in cancer.