Kindlin Regulates Mechanosensitive Activation and Adhesion Assembly of Integrin beta6

Abstract Inside‐out and outside‐in activations of integrin both play important roles in adhesion formation and cell motility. While such mechanisms of RGD‐binding integrin, including integrin β1 and β3, are well documented, mechanosensitive regulations of integrin β6 remain largely unknown. Using traction force‐free RGD‐membrane as the model system, integrin β6 and viscous RGD ligands initially colocalize, assemble micrometer‐sized clusters (15‐min), and then gradually dissipate (60‐min). Domain‐swapping approaches are used and find that integrin β6 chimera with integrin β1's cytoplasmic tail can establish the persistent spatial clustering, suggesting the involvement of inside‐out signaling. In vitro, pulldown and microscale thermophoresis experiments reveal that cytoplasmic adapter kindlin2 is the key factor and specifically exhibits weak association with integrin β6. Such kindlin2 deficiency can in turn be restored by additional kindlin2 or the introduction of PIPK1 that promotes kindlin2 recruitment to the plasma membrane. Dissipated integrin β6 is found in non‐active conformation. Strengthening the interaction between kindlin2 and integrin β6 stabilizes integrin β6 in active conformation and is necessary to promote the migration of AsPC‐1 cells on the soft PDMS. Thus, kindlin‐mediated inside‐out activation of integrin β6 bypasses extracellular force‐dependent signaling and regulates the mechanosensitive adhesion assembly and cell migration on the compliant substrate.