Distinct and compensatory roles of STAG1 and STAG2 in post-mitotic genome refolding

Abstract The three-dimensional architecture of the eukaryotic genome is largely shaped by the cohesin complex, which contains either Stag1 or Stag2 subunits. Although both subunits contribute to chromatin organization, their specific functions in de novo loop formation during post-mitotic genome refolding remain elusive. Here, we leverage the mitosis-to-G1 transition to dissect their individual roles. We found that Stag1 depletion has a negligible impact on post-mitotic genome restructuring or transcription reactivation. In contrast, Stag2 orchestrates chromatin remodeling in a manner that is both cell cycle stage-specific and chromatin context-dependent. During early-G1, Stag2 preferentially associates with euchromatin, where it drives the rapid formation of small chromatin loops. This facilitates prompt promoter-enhancer (P-E) contact formation and enables efficient transcription activation. As the nuclear concentration of Stag2 increases by late-G1, it progressively suppresses large loops, likely due to its shorter chromatin residence time and its potential to competitively displace the more extrusion-capable Stag1-associated cohesin. Mechanistically, Stag2-mediated loop extrusion is constrained by CTCF-bound barriers, rather than by genomic travel distance. Although Stag2 associates rapidly with euchromatin in early-G1, its recruitment to heterochromatin is delayed until late-G1. Simultaneous depletion of both Stag proteins results in a synergistic loss of virtually all structural loops and a more severe disruption of transcription than that caused by individual deletions. Together, these results establish Stag2 as the principal regulator of post-mitotic genome reorganization among Stag paralogs, mediating spatiotemporal control of chromatin architecture, while Stag1 provides compensatory support to ensure functional robustness.