Jpx RNA regulates CTCF anchor site selection and formation of chromosome loops

Chromosome loops shift dynamically during development, homeostasis, and disease. CTCF is known to anchor loops and construct 3D genomes, but how anchor sites are selected is not yet understood. Here we unveil Jpx RNA as a determinant of anchor selectivity. Jpx RNA targets thousands of genomic sites, preferentially binding promoters of active genes. Depleting Jpx RNA causes ectopic CTCF binding, massive shifts in chromosome looping, and downregulation of >700 Jpx target genes. Without Jpx, thousands of lost loops are replaced by de novo loops anchored by ectopic CTCF sites. Although Jpx controls CTCF binding on a genome-wide basis, it acts selectively at the subset of developmentally sensitive CTCF sites. Specifically, Jpx targets low-affinity CTCF motifs and displaces CTCF protein through competitive inhibition. We conclude that Jpx acts as a CTCF release factor and shapes the 3D genome by regulating anchor site usage. Overall design: We performed Jpx CHART-seq in undifferentiated (d0) and differentiating (days 4 and 7 of differentiation) mouse ES cells. Jpx antisense (AS) CHART and no-RNase H CHART served as negative control experiments. We conducted RNA-seq, CTCF ChIP-seq, and RAD21 ChIP-seq in control or Jpx-depleted d7 mouse ES cells. In situ Hi-C experiments were performed in d7 (control and Jpx-depleted) mouse ES cells.