Stress fiber traction force reshapes chromatin accessibility and YAP binding to direct diverse transcriptional programs in mesenchymal stem cells.

Stress fiber-generated traction forces critically regulate mesenchymal stem cell (MSC) behavior, yet how mechanical cues are integrated across transcriptional programs remains unclear. Here, we attenuated actomyosin contractility in human MSCs and performed parallel Assay for Transposase-Accessible Chromatin with high-throughput sequencing (ATAC-seq), YAP-targeted Cleavage Under Targets and Tagmentation sequencing (CUT&Tag) and RNA-seq profiling. We show that reduced stress fiber traction force selectively reorganizes chromatin accessibility into coherent functional modules, resulting in diverse transcriptional programs. The mechanosensitive co-activator YAP functions as a parallel force-responsive regulatory layer coordinating with chromatin accessibility changes. Integration of chromatin accessibility, YAP occupancy, and transcriptomic profiles reveals pathway-specific regulatory responses, identifying focal adhesion and PI3K-Akt signaling as central mechanosensitive pathways coordinated across layers. Together, these findings establish a modular framework for force-dependent gene regulation, demonstrating how mechanical signals are integrated across epigenomic and transcriptional networks to shape MSC transcriptional programs.