Synergistic phase separation of two pathways promotes integrin clustering and nascent adhesion formation.

Lindsay B Case,Milagros De Pasquale,Michael K Rosen,Lisa Henry

doi:10.7554/elife.72588

Lindsay B Case, Milagros De Pasquale + Show 2 more

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https://doi.org/10.7554/elife.72588

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Abstract

Integrin adhesion complexes (IACs) are integrin-based plasma-membrane-associated compartments where cells sense environmental cues. The physical mechanisms and molecular interactions that mediate initial IAC formation are unclear. We found that both p130Cas ('Cas') and Focal adhesion kinase ('FAK') undergo liquid-liquid phase separation in vitro under physiologic conditions. Cas- and FAK- driven phase separation is sufficient to reconstitute kindlin-dependent integrin clustering in vitro with recombinant mammalian proteins. In vitro condensates and IACs in mouse embryonic fibroblasts (MEFs) exhibit similar sensitivities to environmental perturbations including changes in temperature and pH. Furthermore, mutations that inhibit or enhance phase separation in vitro reduce or increase the number of IACs in MEFs, respectively. Finally, we find that the Cas and FAK pathways act synergistically to promote phase separation, integrin clustering, IAC formation and partitioning of key components in vitro and in cells. We propose that Cas- and FAK-driven phase separation provides an intracellular trigger for integrin clustering and nascent IAC formation.

Highlights

Integrin-mediated adhesion complexes (IACs) are plasma membrane-associated compartments that provide specific adhesion between cells and their surroundings (Case and Waterman, 2015; Chastney et al, 2021)
We propose that pCas- and FAK114 driven phase separation provides an intracellular trigger for integrin clustering and nascent adhesion formation. 117 RESULTS Multivalent interactions promote phase separation of p130Cas under physiologic conditions The adaptor protein Cas (p130Cas/BCAR1) is present in nascent adhesions (Donato et al, 2010) and has been implicated in promoting Integrin adhesion complexes (IACs) formation (Meenderink et al, 2010)
Droplets formed from 1 μM pCas, Nck, N-WASP, Focal Adhesion Kinase (FAK) and paxillin did not recruit β1 integrin (PC = 1.2), and this was not changed by addition of 1 μM talin head domain (talinH) (PC = 1.2). adding 1 μM kindlin, significantly increased integrin partitioning in droplets ((PC = 3.0, Fig. 3d)

Summary

Introduction

Integrin-mediated adhesion complexes (IACs) are plasma membrane-associated compartments that provide specific adhesion between cells and their surroundings (Case and Waterman, 2015; Chastney et al, 2021). we find that the Cas and FAK pathways act synergistically to promote phase separation, integrin clustering and nascent adhesion formation in vitro and in cells.

Results

Conclusion