Force-dependent vinculin binding to talin in live cells: a crucial step in anchoring the actin cytoskeleton to focal adhesions

Author:

Hirata Hiroaki12,Tatsumi Hitoshi3,Lim Chwee Teck14,Sokabe Masahiro123

Affiliation:

1. Mechanobiology Institute, National University of Singapore, Singapore;

2. Cell Mechanosensing Project, International Cooperative Research Project/Solution-Oriented Research for Science and Technology, Japan Science and Technology Agency, Nagoya, Japan;

3. Department of Physiology, Nagoya University Graduate School of Medicine, Nagoya, Japan; and

4. Department of Biomedical Engineering and Department of Mechanical Engineering, National University of Singapore, Singapore

Abstract

Mechanical forces play a pivotal role in the regulation of focal adhesions (FAs) where the actin cytoskeleton is anchored to the extracellular matrix through integrin and a variety of linker proteins including talin and vinculin. The localization of vinculin at FAs depends on mechanical forces. While in vitro studies have demonstrated the force-induced increase in vinculin binding to talin, it remains unclear whether such a mechanism exists at FAs in vivo. In this study, using fibroblasts cultured on elastic silicone substrata, we have examined the role of forces in modulating talin-vinculin binding at FAs. Stretching the substrata caused vinculin accumulation at talin-containing FAs, and this accumulation was abrogated by expressing the talin-binding domain of vinculin (domain D1, which inhibits endogenous vinculin from binding to talin). These results indicate that mechanical forces loaded to FAs facilitate vinculin binding to talin at FAs. In cell-protruding regions, the actin network moved backward over talin-containing FAs in domain D1-expressing cells while it was anchored to FAs in control cells, suggesting that the force-dependent vinculin binding to talin is crucial for anchoring the actin cytoskeleton to FAs in living cells.

Publisher

American Physiological Society

Subject

Cell Biology,Physiology

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