<em>N</em>⁶-Methyladenosine Landscape of Glioma Stem-Like Cells: METTL3 Is Essential for the Expression of Actively Transcribed Genes and Sustenance of the Oncogenic Signaling

Abstract

Despite recent advances in N6-methyladenosine (m6A) biology, the regulation of crucial RNA processing steps by the RNA methyltransferase-like 3 (METTL3) in glioma stem-like cells (GSCs) remains obscure. An integrated analysis of m6A-RIP (RNA immunoprecipitation) and total RNA-Seq of METTL3-silenced GSCs identified that m6A modification in GSCs is principally carried out by METTL3. The m6A-modified transcripts showed higher abundance compared to non-modified transcripts. Further, we showed that the METTL3 is essential for the expression of GSC-specific actively transcribed genes. Silencing METTL3 resulted in the elevation of several aberrant alternative splicing events. We also found that putative m6A reader proteins play a key role in the RNA stabilization function of METTL3. METTL3 altered A-to-I and C-to-U RNA editing events by differentially regulating RNA editing enzymes ADAR and APOBEC3A. Similar to protein-coding genes, lincRNAs (long intergenic non-coding RNAs) with m6A marks showed METTL3-dependent high expression. m6A modification of 3′UTRs appeared to result in a conformation-dependent hindrance to miRNA binding to their targets. The integrated analysis of the m6A regulome in METTL3-silenced GSCs showed global disruption in tumorigenic pathways that are indispensable for GSC maintenance and glioma progression. We conclude that METTL3 plays a vital role in many steps of RNA processing and orchestrates successful execution of oncogenic pathways in GSCs.

Keywords

• METTL3
• m⁶A
• GSCs
• glioblastoma
• histone activation
• splicing
• RNA editing
• m⁶A reader