N(6)-adenosine-methyltransferase catalytic subunit METTL3
RNA N6-methyladenosine methyltransferase complex
m(6)A RNA methylation promotes XIST-mediated transcriptional repression.
The long non-coding RNA X-inactive specific transcript (XIST) mediates the transcriptional silencing of genes on the X chromosome. Here we show that, in human cells, XIST is highly methylated with at least 78 N<sup>6</sup>-methyladenosine (m<sup>6</sup>A) residues-a reversible base modification of unknown function in long non-coding RNAs. We show that m<sup>6</sup>A formation in XIST, as well as in cellular mRNAs, is mediated by RNA-binding motif protein 15 (RBM15) and its paralogue RBM15B, which bind the m<sup>6</sup>A-methylation complex and recruit it to specific sites in RNA. This results in the methylation of adenosine nucleotides in adjacent m<sup>6</sup>A consensus motifs. Furthermore, we show that knockdown of RBM15 and RBM15B, or knockdown of methyltransferase like 3 (METTL3), an m<sup>6</sup>A methyltransferase, impairs XIST-mediated gene silencing. A systematic comparison of m<sup>6</sup>A-binding proteins shows that YTH domain containing 1 (YTHDC1) preferentially recognizes m<sup>6</sup>A residues on XIST and is required for XIST function. Additionally, artificial tethering of YTHDC1 to XIST rescues XIST-mediated silencing upon loss of m<sup>6</sup>A. These data reveal a pathway of m<sup>6</sup>A formation and recognition required for XIST-mediated transcriptional repression.