An amphipathic helix of vinexin α is necessary for substrate stiffness-dependent conformational change in vinculin

Extracellular matrix (ECM) stiffness regulates various cell behaviors, including cell differentiation, proliferation, and migration. Vinculin and vinexin α, both of which localize to focal adhesions, cooperatively function as mechanosensors of ECM stiffness. On a rigid ECM, vinexin α interacts with vinculin and induces a conformational change in vinculin to an ‘open’ form, which promotes nuclear localization of Yes-associated protein (YAP) and transcriptional coactivator with a PDZ-binding motif (TAZ). However, the detailed mechanism by which vinexin α induces the conformational change in vinculin has not been revealed. Here, we identify an amphipathic helix named H2 as a novel vinculin binding site in vinexin α. H2 helix interacts with the vinculin D1b subdomain and promotes the formation of a talin-vinculin-vinexin α ternary complex. Mutations in the H2 region not only impair the ability of vinexin α to induce the ECM stiffness-dependent conformational change in vinculin but also to promote nuclear localization of YAP/TAZ on rigid ECM. Altogether, these results demonstrate that the H2 helix in vinexin α plays a critical role in ECM-stiffness-dependent regulation of vinculin and cell behaviors.