Essential role of METTL3-mediated m6A modification in glioma stem-like cells maintenance and radioresistance

Oncogene. 2018 Jan 25;37(4):522-533. doi: 10.1038/onc.2017.351. Epub 2017 Oct 9.

Abstract

Despite advances in biology and therapeutic modalities, existence of highly tumorigenic glioma stem-like cells (GSCs) makes glioblastomas (GBMs) invincible. N6-methyl adenosine (m6A), one of the abundant mRNA modifications catalyzed by methyltransferase-like 3 and 14 (METTL3/14), influences various events in RNA metabolism. Here, we report the crucial role of METTL3-mediated m6A modification in GSC (neurosphere) maintenance and dedifferentiation of glioma cells. METTL3 expression is elevated in GSC and attenuated during differentiation. RNA immunoprecipitation studies identified SOX2 as a bonafide m6A target of METTL3 and the m6A modification of SOX2 mRNA by METTL3 enhanced its stability. The exogenous overexpression of 3'UTR-less SOX2 significantly alleviated the inhibition of neurosphere formation observed in METTL3 silenced GSCs. METTL3 binding and m6A modification in vivo required intact three METTL3/m6A sites present in the SOX2-3'UTR. Further, we found that the recruitment of Human antigen R (HuR) to m6A-modified RNA is essential for SOX2 mRNA stabilization by METTL3. In addition, we found a preferential binding by HuR to the m6A-modified transcripts globally. METTL3 silenced GSCs showed enhanced sensitivity to γ-irradiation and reduced DNA repair as evidenced from the accumulation of γ-H2AX. Exogenous overexpression of 3'UTR-less SOX2 in METTL3 silenced GSCs showed efficient DNA repair and also resulted in the significant rescue of neurosphere formation from METTL3 silencing induced radiosensitivity. Silencing METTL3 inhibited RasV12 mediated transformation of mouse immortalized astrocytes. GBM tumors have elevated levels of METTL3 transcripts and silencing METTL3 in U87/TIC inhibited tumor growth in an intracranial orthotopic mouse model with prolonged mice survival. METTL3 transcript levels predicted poor survival in GBMs which are enriched for GSC-specific signature. Thus our study reports the importance of m6A modification in GSCs and uncovers METTL3 as a potential molecular target in GBM therapy.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • 3' Untranslated Regions / genetics
  • Adenosine / analogs & derivatives
  • Adenosine / metabolism
  • Animals
  • Brain / cytology
  • Brain / pathology
  • Brain / surgery
  • Brain Neoplasms / genetics*
  • Brain Neoplasms / pathology
  • Brain Neoplasms / radiotherapy
  • Brain Neoplasms / surgery
  • Carcinogenesis / genetics
  • Cell Dedifferentiation / genetics
  • Cell Dedifferentiation / radiation effects
  • Cell Line, Tumor
  • Cell Proliferation / genetics
  • DNA Repair / radiation effects
  • ELAV-Like Protein 1 / metabolism
  • Female
  • Gene Expression Regulation, Neoplastic
  • Glioblastoma / genetics*
  • Glioblastoma / pathology
  • Glioblastoma / radiotherapy
  • Glioblastoma / surgery
  • Humans
  • Methyltransferases / genetics
  • Methyltransferases / metabolism*
  • Mice
  • Mice, Nude
  • Mutagenesis, Site-Directed
  • Neoplastic Stem Cells / pathology*
  • Neoplastic Stem Cells / radiation effects
  • RNA, Messenger / metabolism
  • Radiation Tolerance / genetics
  • SOXB1 Transcription Factors / genetics*
  • SOXB1 Transcription Factors / metabolism
  • Spheroids, Cellular / radiation effects
  • Xenograft Model Antitumor Assays

Substances

  • 3' Untranslated Regions
  • ELAV-Like Protein 1
  • ELAVL1 protein, human
  • RNA, Messenger
  • SOX2 protein, human
  • SOXB1 Transcription Factors
  • N-methyladenosine
  • Methyltransferases
  • METTL3 protein, human
  • Adenosine