Transcription can directly remodel chromatin architecture around activated genes.

(A) Pooled Capture Hi-C interaction maps for an approximately 600 kb region around the Nfatc3 gene are shown for DN3 and DP cells, above the differential heat map, where DP-enriched interactions are displayed in red and DN3-enriched interactions in blue. The arrows indicate the DP-specific domain. Positions of genes are shown below the plots. (B) Insulation scores for the same genomic region as in (A), computed at 2 kb resolution with an 80-kb (40 bin) window for DN3 (blue) and DP (red) cells. (C) ChIP-seq profiles (normalized as counts per million reads) for the same genomic region as in (A), for DN3 (blue) and DP (red) cells. Top to bottom: RNA polymerase II, CTCF. Red stripe indicates DP-specific domain, with increased insulation score maximum, increased RNA polymerase II loading, and negligible changes in CTCF binding. (D) Pooled Capture Hi-C interaction maps for the same genomic region as in (A) are shown for untreated ESCs and ESCs with ectopic activation of Nfatc3, above the differential heat map. Arrow indicates position of strengthened domain brought about by Nfatc3 induction. (E) Insulation scores for the same genomic region as in (A), computed at 2 kb resolution with an 80-kb (40 bin) window for untreated (green) and Nfatc3-induced (black) ESCs. Dark stripe indicates position of reinforced domain with corresponding increase in insulation score maximum. (F) Pooled Capture Hi-C interaction maps for an approximately 600 kb region around the Il17rb gene are shown for ESCs and DN3 cells, above the differential heat map, where DN3-enriched interactions are displayed in red and ESC-enriched interactions in blue. The arrows indicate the thymocyte-specific domain, encompassing Il17rb and Chdh. Positions of genes are shown below the plots. (G) Insulation scores for the same genomic region as in (F), computed at 2 kb resolution with an 80-kb (40 bin) window for ESCs (green), DN3 (blue) and DP (red) cells. (H) CTCF ChIP-seq profiles (normalized as counts per million reads) for the same genomic region as in (F), for ESCs (green) and DN3 cells (blue). Yellow stripe indicates thymocyte-specific domain. (I) Bar charts showing normalized Nfatc3 and Il17rb expression levels in untreated and CRISPRa-treated cells determined by qRT-PCR (2 biological replicates). (J) Pooled Capture Hi-C interaction maps for the same genomic region as in (F) are shown for untreated ESCs and ESCs with ectopic activation of Il17rb, above the differential heat map. Arrow indicates position of strengthened domain, and rectangle shows reduction in interdomain interactions brought about by Il17rb induction. (K) Insulation scores for the same genomic region as in (F), computed at 2 kb resolution with an 80-kb (40 bin) window for untreated (green) and Il17rb-induced (black) ESCs. Yellow stripe indicates position of new domain, containing new insulation score minima flanking an insulation score local maximum. Source data can be found in S1 Data. CRISPRa, CRISPR activation; DN3, double negative; DP, double positive; ESC, embryonic stem cell.