Abstract
<title>Abstract</title> <p>Replication stress triggers extensive remodelling of nuclear architecture to safeguard genome stability, but how this reorganization is coordinated remains unclear. Here we identify TopBP1 condensates as central regulators of nuclear actin remodelling during this response. Sustained replication stress promotes TopBP1 condensate formation in S phase at both the nuclear interior and the nuclear periphery. We show that these condensates drive nuclear F-actin assembly through ATR-mTOR-N-WASP-dependent nucleation, while separately stabilizing actin filaments through an mTOR-dependent, ATR-independent mechanism involving cofilin inhibition. Nuclear F-actin, in turn, is required for proper TopBP1 condensate positioning, revealing a reciprocal relationship between actin network organization and condensate localization. At the nuclear periphery, TopBP1 condensates further require Lamin B1 and stabilize Emerin, coupling nuclear envelope integrity to the spatial organization of DNA repair. Notably, this TopBP1-dependent nuclear actin programme is enhanced in oxaliplatin-resistant colorectal cancer cells, where it promotes survival under chemotherapy. Our findings establish TopBP1 condensates as key architectural hubs that integrate replication stress signaling with nuclear actin dynamics and envelope organization, and identify TopBP1 condensation as a potential therapeutic target in chemo-resistant cancer.</p>