The M~6 A Methyltransferase METTL3 Promotes Bladder Cancer Progression Via AFF4/NF-kB/MYC Signaling Network

N6-methyladenosine(m~6A)is the most abundant modification in eukaryotic messenger RNAs(m RNAs),and plays important roles in many bioprocesses.However,its functions in bladder cancer(BCa)remain elusive.Here,we discovered that methyltransferase-like 3(METTL3),a major RNA N6-adenosine methyltransferase,was significantly up-regulated inhuman BCa.Knockdown of METTL3 drastically reduced BCa cell proliferation,invasion,and survival in vitro and tumorigenicity in vivo.On the other hand,overexpression of METTL3significantly promoted BCa cell growth and invasion.Through transcriptome sequencing,m~6A sequencing and m~6A methylated RNA immuno-precipitation quantitative reverse transcription polymerase chain reaction,we revealed the profile of METTL3-mediated m~6A modification in BCa cells for the first time.AF4/FMR2family member 4(AFF4),two key regulators of NF-?B pathway(IKBKB and RELA)and MYC were further identified as direct targets of METTL3-mediated m~6A modification.In addition,we showed that besides NF-?B,AFF4 binds to the promoter of MYC and promotes its expression,implying a novel multilevel regulatory network downstream of METTL3.Our results uncovered an AFF4/NF-?B/MYC signaling network operated by METTL3-mediated m6A modification and provided insight into the mechanisms of BCa progression.