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

Abstract

Despite advances in biology and therapeutic modalities, existence of highly tumorigenic glioma stem-like cells (GSCs) makes glioblastomas (GBMs) invincible. N6-methyl adenosine (mA), 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 mA 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 mA target of METTL3 and the mA 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 mA modification in vivo required intact three METTL3/mA sites present in the SOX2-3'UTR. Further, we found that the recruitment of Human antigen R (HuR) to mA-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 mA modification in GSCs and uncovers METTL3 as a potential molecular target in GBM therapy.

The dual role of N6‐methyladenosine modification of RNAs is involved in human cancers

“…However, recently, Visvanathan et al. have discovered powerful but opposite evidence that METTL3‐mediated m 6 A modification was required for GSCs maintenance 48. The underlying mechanism here is controversial with what we discussed above.…”

Profiling of m6A RNA modifications identified an age‐associated regulation of AGO2 mRNA stability

“…In addition, CDF analysis of AUF1 and HuR target mRNAs (Mukherjee et al., 2011; Yoon et al., 2014) from MeRIP‐seq and total RNA‐seq data has shown that AUF1 and HuR target RNAs are downregulated further than nontarget mRNAs once they are methylated (Figure 3f,g, Figures S2 and S3). These results imply possible involvement of AUF1 and HuR in the stability regulation of m6A‐modified RNA (Visvanathan et al., 2017). …”

“…Dynamic regulation of m6A modification and mRNA decay is also dictated by position‐specific m6A methylation within the mRNA, and presence/activity of m6A‐binding proteins within a given context (Alarcón et al., 2015; Wang & He, 2014; Wang et al., 2014, 2015). A possibility is that DROSHA mRNA methylation is maintained by the presence of m6A readers or m6A‐regulated factors such as HuR, FMR1, and G3BP, resulting in preservation of DROSHA mRNA levels during the aging process (Edupuganti et al., 2017; Visvanathan et al., 2017). This may explain why DROSHA mRNA levels did not change despite having higher methylation levels compared with AGO2 mRNA.…”

Methyltransferase like 3 promotes colorectal cancer proliferation by stabilizing CCNE1 mRNA in an m6A‐dependent manner

“…As the critical m6A methyltransferase, the role of METTL3 in tumours is controversial. Some studies reported that METTL3 played an oncogenic role in myeloid leukaemia, liver cancer, breast cancer, glioblastoma, bladder cancer and lung cancer . Other studies indicated that METTL3 played a tumour suppressor in renal cell carcinoma and glioblastoma .…”

“…23 Recent studies showed that m6A modification played essential roles in tissue development, stem cell formation and differentiation, 24,25 control of heat shock response 26 and circadian clock controlling, 27 as well as in tumours formation. [28][29][30][31][32][33][34][35] As the critical m6A methyltransferase, the role of METTL3 in tumours is controversial.…”