Hyperosmotic stress induces a large-scale rewiring of 3D chromatin interactions [mAID2 Hi-C]

Cells rapidly adapt to hyperosmotic stress through coordinated molecular responses. To determine how three-dimensional (3D) chromatin structure contributes to this process, we profiled chromatin interactions, architectural protein occupancy, and transcriptional dynamics in human cells exposed to sorbitol-induced hyperosmotic stress. We performed time-resolved Hi-C to measure stress-induced remodeling of chromatin loops and domains, CUT&Tag to quantify CTCF, RAD21, YAP1, and H3K27ac occupancy at loop anchors, and RNA-seq to capture stress-responsive transcriptional programs. These data reveal global loss of pre-existing chromatin contacts and concurrent formation of de novo, transient loops enriched for retained CTCF and cohesin, as well as transcriptional responses that are temporally layered and largely decoupled from loop remodeling. This dataset provides a resource for investigating how nuclear architecture and transcriptional regulation respond to hyperosmotic stress. Overall design: eGFP-YAP HEK293 cells were serum-starved for 1 hour and then exposed to 200 mM sorbitol to induce hyperosmotic stress. Chromatin organization was profiled by in situ Hi-C at eight time points (0 min, 10 min, 30 min, 1 h, 3 h, 6 h, 12 h, and 24 h) to capture the formation, loss, and recovery of chromatin interactions. To assess the generality of chromatin remodeling, additional Hi-C experiments were performed in HEK293 wild-type and HCT116 cells with and without 1 hour of sorbitol treatment. To probe the requirement for architectural proteins, we also performed Hi-C in HCT116 mAID2-CTCF and mAID2-RAD21 degron cell lines treated with 1 µM 5-Ph-IAA to rapidly and reversibly degrade CTCF or RAD21 prior to sorbitol exposure. Transcriptional responses were measured by stranded RNA-seq across a time course (0 h, 1 h, 3 h, 6 h, 9 h, 12 h, and 24 h). To quantify changes in architectural protein occupancy, CUT&Tag for CTCF and RAD21 was performed in untreated and 1-hour sorbitol-treated HEK293 cells.