In the present study, we developed a talin tension sensor (TS) and explored the role of mechanical force across talin in integrin-mediated adhesion and mechanotransduction.Ĭonstruction and characterization of a talin TS How this dynamic assembly mediates mechanotransduction is therefore a key question.ĭevelopment of a method to measure forces across specific molecules using a fluorescence resonance energy transfer (FRET) pair connected to a calibrated spring demonstrated directly that vinculin in FAs is under mechanical tension ( Grashoff et al., 2010). The integrin- and F-actin bonds between vinculin and talin must therefore be dynamic, with rapid association and dissociation, to mediate force transmission, the so-called FA clutch. In focal adhesions (FAs) near cell edges, actin flows rearward over the immobile integrins, with talin and vinculin moving rearward at intermediate rates. The force-transmitting linkages between integrins and actin are dynamic, with F-actin flowing over the adhesions under the force exerted by both actin polymerization and myosin-dependent filament sliding ( Case and Waterman, 2015). Mechanosensing through integrins is important in development and numerous diseases including cancer, hypertension, and fibrosis ( Orr et al., 2006 Butcher et al., 2009). These effects include modulation of ECM production by matrix stiffness and externally applied forces. For example, cells sense the mechanical stiffness of the ECM and modulate their own contractility, signaling, and gene expression programs accordingly, a property termed stiffness sensing ( Humphrey et al., 2014). The mechanosensitivity of integrin-mediated adhesions allows tissues to tune their function and gene expression to mechanical cues in the environment ( Orr et al., 2006 Costa et al., 2012). Talin deletion in several organisms yields phenotypes that are similar to deletion or mutation of the integrins themselves, consistent with its essential role ( Monkley et al., 2000 Brown et al., 2002 Cram et al., 2003). When talin is under mechanical tension, these domains can unravel to allow binding of the vinculin head domain, which reinforces the linkage to actin through an ABS in the vinculin tail. The talin rod domain also contains multiple binding sites for vinculin, which are buried within 4- and 5-α-helical bundles. Talin contains three F-actin–binding sites (ABSs), with the far C-terminal–binding site in the rod domain, ABS3, generally thought to be the most important.
The N-terminal FERM (or head) domain of talin binds directly to integrin β subunit cytoplasmic domains and is required for conformational activation of integrins to bind ECM proteins with high affinity. Integrins connect the ECM to the actin cytoskeleton through a complex set of linkages in which the cytoskeletal protein talin plays a prominent role ( Ziegler et al., 2008 Calderwood et al., 2013). Overall, these results shed new light on talin function and constrain models for cellular mechanosensing. These results indicate that central versus peripheral adhesions must be organized and regulated differently, and that ABS2 and ABS3 have distinct functions in spatial variations and stiffness sensing. However, differential stiffness sensing by talin requires ABS2 but not vinculin or ABS3.
The difference between central and peripheral adhesions requires ABS3 but not vinculin or ABS2. Unlike vinculin, talin is under lower tension on soft substrates.
Tension on talin is increased by vinculin and depends mainly on actin-binding site 2 (ABS2) within the middle of the rod domain, rather than ABS3 at the far C terminus. We find that talin in focal adhesions is under tension, which is higher in peripheral than central adhesions. Here, we report the development and validation of a talin tension sensor. Integrin-dependent adhesions are mechanosensitive structures in which talin mediates a linkage to actin filaments either directly or indirectly by recruiting vinculin.
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