LIMD1 phase separation contributes to cellular mechanics and durotaxis by regulating focal adhesion dynamics in response to force.

Cells sense and respond to extracellular mechanical cues through cell-matrix adhesions. Interestingly, the maturation of focal adhesions (FAs) is reciprocally force dependent. How biomechanical cues dictate the status of cell motility and how FAs spatial temporally coordinate force sensing and self-organization remain enigmatic. Here, we identify that LIMD1, a member of the LIM domain scaffolding proteins, undergoes force-sensitive condensation at the FAs. We also unveil that the multivalent interactions of LIMD1 intrinsically disordered region (IDR) and the LIM domains concertedly drive this phase transition under the regulation of phosphorylation. Intriguingly, formation of condensed LIMD1 protein compartments is sufficient to specifically enrich and localize late FA proteins. We further discover that LIMD1 regulates cell spreading, maintains FA dynamics and cellular contractility, and is critical for durotaxis-the ability of cells to crawl along gradients of substrate stiffness. Our results suggest a model that recruitment of LIMD1 to the FAs, via mechanical force triggered inter-molecular interaction, serves as a phase separation hub to assemble and organize matured FAs, thus allowing for efficient mechano-transduction and cell migration.