Xist Repeat A coordinates an assembly of SR proteins to recruit SPEN and induce gene silencing

The lncRNA Xist silences gene expression by recruiting the protein SPEN through its 5′-proximal Repeat A domain. How Repeat A recruits SPEN, how SPEN enacts silencing, and why Repeat A is required for Xist biogenesis in addition to silencing remain unclear. We find that sequences in Repeat A critical for SPEN recruitment, silencing, and Xist biogenesis directly bind SR-rich splicing factors and not SPEN. SRSF1, one such factor, is necessary and sufficient for SPEN recruitment to Repeat A. SPEN and SR protein-binding motifs in Repeat A enable Xist ’s association with many proteins, including those involved in m ⁶ A methylation, RNA turnover, and transcriptional elongation. Our results reveal an unexpectedly essential role for splicing factors in coordinating silencing by SPEN and suggest a unifying model for the origin of Repeat A, the function of SPEN, and their roles in Xist biogenesis and silencing.