A conserved noncoding locus modulates random monoallelic Xist expression across a TAD boundary

Topologically associating domains (TADs) are thought to restrict the action of distal cis-regulatory elements, such as enhancers, to target gene promoters within the same TAD. Here we discover that a cis-regulatory element of Xist is located almost 200kb away from its promoter and across a TAD boundary. This element, the promoter of a noncoding RNA, Linx, opposes Xist up-regulation in cis and influences the choice of X chromosome to be inactivated. Even though Linx lies in the same TAD as Xist’s antisense Tsix locus, we show that the repressive action of Linx on Xist is independent of Tsix. Furthermore, this Xist regulatory action is independent of Linx transcript or of its transcription. Finally, we demonstrate that Linx cis-regulatory repressive element is well conserved in mammals, unlike Tsix, suggesting that it represents an ancestral mechanism for random monoallelic Xist expression. Our study uncovers a novel regulatory axis for X-chromosome inactivation and a new class of cis-regulatory effects that rely on TAD partitioning to modulate developmental decisions. Our dataset includes 8 RNA-seq samples, 22 5C-samples, 20 Capture-C samples and 6 ATAC-seq samples. These include at least two biological replicates for each condition. Control/reference samples were included and processed in parallel: "wildtype" genotype to compare with deletions/inversions, and "day 0 (undifferentiated)" to compare with differentiated samples.