CAS directly interacts with vinculin to control mechanosensing and focal adhesion dynamics

Abstract

Focal adhesions are cellular structures through which both mechanical forces and regulatory signals are transmitted. Two focal adhesion-associated proteins, Crk-associated substrate (CAS) and vinculin, were both independently shown to be crucial for the ability of cells to transmit mechanical forces and to regulate cytoskeletal tension. Here, we identify a novel, direct binding interaction between CAS and vinculin. This interaction is mediated by the CAS SRC homology 3 domain and a proline-rich sequence in the hinge region of vinculin. We show that CAS localization in focal adhesions is partially dependent on vinculin, and that CAS–vinculin coupling is required for stretch-induced activation of CAS at the Y410 phosphorylation site. Moreover, CAS–vinculin binding significantly affects the dynamics of CAS and vinculin within focal adhesions as well as the size of focal adhesions. Finally, disruption of CAS binding to vinculin reduces cell stiffness and traction force generation. Taken together, these findings strongly implicate a crucial role of CAS–vinculin interaction in mechanosensing and focal adhesion dynamics.Electronic supplementary materialThe online version of this article (doi:10.1007/s00018-013-1450-x) contains supplementary material, which is available to authorized users.

Identification of Novel Crk-associated Substrate (p130Cas) Variants with Functionally Distinct Focal Adhesion Kinase Binding Activities

“…These cells grew in tight clusters, whereas cells expressing the most studied Cas1A or the other variants showed a high degree of cell scattering. The partial deletion of the SH3 domain in Cas1B corroborates the importance of the SH3 domain in establishing a polarized phenotype associated with cell motility and FA formation (1,42,43). Importantly, reconstitution of Cas1B in Cas Ϫ/Ϫ cells significantly reduced cell migration and invasion when compared with Cas1A cells.…”

The role of focal adhesion anchoring domains of CAS in mechanotransduction

“…Alternatively, it was shown that the interaction of the CAS-SH3 domain with its substrates can be regulated by phosphorylation on Tyr1291213. This enables dynamically regulated changes in CAS targeting to focal adhesions, which might be critical for proper subsequent signaling.…”

Vinculin in cell–cell and cell–matrix adhesions

“…Finally, a third possibility is phosphorylation enhances mechanical activation and vice versa [20]. Consistent with this notion, stretching uncovers tyrosine phosphorylation sites in other proteins (i.e., p130 Cas) [23, 24] as well as vinculin binding sites in talin [25]. Thus, vinculin activation is likely to be more sophisticated than the combinatorial activation or bundle inversion models predict (Fig.…”