N⁶-methylation of 2′-O-methyladenosine (Am) in RNA occurs in eukaryotic cells to generate N⁶,2′-O-dimethyladenosine (m⁶Am). Identification of the methyltransferase responsible for m⁶Am catalysis has accelerated studies on the function of m⁶Am in RNA processing. While m⁶Am is generally found in the first transcribed nucleotide of mRNAs, the modification is also found internally within U2 snRNA. However, the writer required for catalyzing internal m⁶Am formation had remained elusive. By sequencing transcriptome-wide RNA methylation at single-base-resolution, we identified human METTL4 as the writer that directly methylates Am at U2 snRNA position 30 into m⁶Am. We found that METTL4 localizes to the nucleus and its conserved methyltransferase catalytic site is required for U2 snRNA methylation. By sequencing human cells with overexpressed Mettl4, we determined METTL4’s in vivo target RNA motif specificity. In the absence of Mettl4 in human cells, U2 snRNA lacks m⁶Am thereby affecting a subset of splicing events that exhibit specific features such as 3′ splice-site weakness and an increase in exon inclusion. These findings suggest that METTL4 methylation of U2 snRNA regulates splicing of specific pre-mRNA transcripts.

中文翻译：

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METTL4 催化 U2 snRNA 中的 m6Am 甲基化以调节前 mRNA 剪接。

RNA中2'- O-甲基腺苷(Am)的N ⁶ -甲基化发生在真核细胞中以产生N ⁶ ,2'- O-二甲基腺苷(m ⁶ Am)。负责 m ⁶ Am 催化的甲基转移酶的鉴定加速了对 m ⁶ Am 在 RNA 加工中的功能的研究。虽然 m ⁶ Am 通常在 mRNA 的第一个转录核苷酸中发现，但在U2 snRNA内部也发现了这种修饰。然而，催化内部 m ⁶所需的作家Am 编队仍然难以捉摸。通过以单碱基分辨率对转录组范围内的 RNA 甲基化进行测序，我们将人类 METTL4 鉴定为直接将U2 snRNA位置 30处的 Am 甲基化为 m ⁶ Am 的作者。我们发现 METTL4 定位于细胞核，其保守的甲基转移酶催化位点是U2 snRNA甲基化所必需的。通过对过表达Mettl4 的人类细胞进行测序，我们确定了 METTL4 的体内靶标 RNA 基序特异性。在人类细胞中缺乏Mettl4的情况下，U2 snRNA缺乏 m ⁶Am 从而影响显示特定特征的剪接事件子集，例如 3' 剪接位点弱点和外显子包含的增加。这些发现表明U2 snRNA 的METTL4 甲基化调节特定前 mRNA 转录本的剪接。