m6A 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 N6-methyladenosine (m6A) residues—a reversible base modification of unknown function in long non-coding RNAs. We show that m6A 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 m6A-methylation complex and recruit it to specific sites in RNA. This results in the methylation of adenosine nucleotides in adjacent m6A consensus motifs. Furthermore, we show that knockdown of RBM15 and RBM15B, or knockdown of methyltransferase like 3 (METTL3), an m6A methyltransferase, impairs XIST-mediated gene silencing. A systematic comparison of m6A-binding proteins shows that YTH domain containing 1 (YTHDC1) preferentially recognizes m6A residues on XIST and is required for XIST function. Additionally, artificial tethering of YTHDC1 to XIST rescues XIST-mediated silencing upon loss of m6A. These data reveal a pathway of m6A formation and recognition required for XIST-mediated transcriptional repression. The methylation of adenosine residues on the long non-coding RNA XIST is essential for X-chromosome transcriptional repression during female mammalian development. The long non-coding RNA (lncRNA) known as XIST is required for the translational silencing of genes on one of the two X chromosomes during female mammalian development. Here, Samie Jaffrey and colleagues show that XIST is highly methylated, containing at least 78 N6-methyladenosine (m6A) residues throughout the length of the RNA. The XIST-binding protein RBM15 is shown to recruit the WTAP–METTL3 RNA m6A methyltransferase complex to XIST, and the nuclear m6A reader YTHDC1 is shown to activate gene-silencing mechanisms.