The dynamic N⁶-methyladenosine (m⁶A) modification of mRNA plays a role in regulating gene expression and determining cell fate. However, the functions of m⁶A mRNA modification in bladder cancer stem cells (BCSCs) have not been described. Here, we show that global RNA m⁶A abundance and the expression of m⁶A-forming enzyme METTL3 are higher in BCSCs than those in non-CSCs of bladder cancer (BCa) cells. The depletion of the METTL3 inhibited the self-renewal of BCSCs, as evidenced by decreased ALDH activity and sphere-forming ability. Mechanistically, METTL3 regulates the m⁶A modification and thereby the expression of AF4/FMR2 family member 4 (AFF4), knockdown of which phenocopies the METTL3 ablation and diminishes the tumor-initiating capability of BCSCs in vivo. AFF4 binds to the promoter regions and sustains the transcription of SOX2 and MYC which have critical biological functions in BCSCs. Collectively, our results demonstrate the critical roles of m⁶A modification in self-renewal and tumorigenicity of BCSCs through a novel signaling axis of METTL3-AFF4-SOX2/MYC.

中文翻译：

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m6A甲基化调节的AFF4促进膀胱癌干细胞的自我更新。

mRNA的动态N ^6-甲基腺苷（m ⁶ A）修饰在调节基因表达和决定细胞命运方面起作用。然而，尚未描述m ⁶ A mRNA修饰在膀胱癌干细胞（BCSC）中的功能。在这里，我们显示，BCSC中的全局RNA m ⁶ A丰度和m ⁶ A形成酶METTL3的表达高于膀胱癌（BCa）细胞的非CSC中的表达。METTL3的耗竭抑制了BCSCs的自我更新，这可以通过降低ALDH活性和形成球的能力来证明。机械上，METTL3调节m ⁶AF4 / FMR2家族成员4（AFF4）的修饰及其表达，其敲除显型复制METTL3消融并减弱体内BCSC的肿瘤启动能力。AFF4结合启动子区域并维持SOX2和MYC的转录，而SOX2和MYC在BCSC中具有关键的生物学功能。总的来说，我们的结果证明了通过METTL3-AFF4-SOX2 / MYC的新信号轴，m ⁶ A修饰在BCSCs自我更新和致瘤性中的关键作用。