United we stand: integrating the actin cytoskeleton and cell-matrix adhesions in cellular mechanotransduction

Abstract

Many essential cellular functions in health and disease are closely linked to the ability of cells to respond to mechanical forces. In the context of cell adhesion to the extracellular matrix, the forces that are generated within the actin cytoskeleton and transmitted through integrin-based focal adhesions are essential for the cellular response to environmental clues, such as the spatial distribution of adhesive ligands or matrix stiffness. Whereas substantial progress has been made in identifying mechanosensitive molecules that can transduce mechanical force into biochemical signals, much less is known about the nature of cytoskeletal force generation and transmission that regulates the magnitude, duration and spatial distribution of forces imposed on these mechanosensitive complexes. By focusing on cell-matrix adhesion to flat elastic substrates, on which traction forces can be measured with high temporal and spatial resolution, we discuss our current understanding of the physical mechanisms that integrate a large range of molecular mechanotransduction events on cellular scales. Physical limits of stability emerge as one important element of the cellular response that complements the structural changes affected by regulatory systems in response to mechanical processes.

Fig. 1.

Adhesion and actin organization at different stages of adhesion assembly. (A) Side view schematic. (1) The lamellipodium (LP) drives nascent adhesions assembly at the leading edge of the cell. (2) Nascent adhesions recruit different cytoplasmic proteins and strengthen their linkage to the actin cytoskeleton. (3) Focal-adhesion maturation occurs within the lamella (LM). (B) Top view schematic. The LP–LM boundary is associated with the generation of different contractile actomyosin bundles. Transverse arcs (TA) are parallel to the cell edge and the LP–LM boundary. Radial stress fibers (SF) are anchored in focal adhesions and oriented perpendicular to the cell edge. The locations of the types of focal adhesion described in (A) are indicated by (1) to (3).

Fig. 2.

Traction-force microscopy. (A) Actin and (B) focal adhesions in a U2OS cell visualized by GFP–actin and mApple–paxillin, respectively. The yellow dashed line in the actin image indicates the boundary between the lamellipodium and lamella, which contains transverse arcs and stress fibers. Whereas stress fibers are straight, peripheral bundles are invaginated as a result of the contractility within the cell body. Scale bar, 10 µm. (C) Forces exerted during traction-force microscopy. During traction-force microscopy, traction forces are reconstructed on the basis of substrate displacements that are tracked with fluorescent beads embedded in a soft elastic substrate. (D) Reconstructed traction stresses exerted on the underlying fibronectin-coated polyacrylamide substrate. Clearly, actin organization, the localization of adhesions marked by GFP–paxillin and the traction-force pattern show strong correlations with each other. In particular, regions of high forces correlate with the presence of focal adhesions.

Fig. 3.

Correlations between retrograde flow, adhesion size and traction forces during adhesion assembly. (A) Different kinds of traction force are generated in different actin modules. (1) During adhesion assembly, polymerization forces drive rapid retrograde flow of actin (red arrow) and associated adhesion proteins, and weakly ligated integrins (gray arrow). Low traction forces (black arrow, F) are exerted. (2) During adhesion stabilization, myosin stresses drive the flow of actin; retrograde movement of adhesions is reduced as they become engaged to the ECM and traction forces on the ECM build. (3) During adhesion maturation, actin retrograde flow is continuous as a stress fiber assembles at the adhesion plaque, focal adhesion proteins accumulate, adhesion elongates and traction forces increase. (B) The typical correlation between retrograde flow speed and traction force shows a biphasic form, with the peak separating two different types of friction regimes. (C) The typical correlation between focal adhesion size and traction force shows different regimes for small and large adhesions. In B and C, different regimes of the correlations are related to the different stages [(1), (2) and (3)] of the assembly process as shown in A. F, force; v, velocity.

Fig. 4.

Mechanisms of force-dependent protein recruitment to actin filaments. (A) Force-induced stretching of proteins (blue ellipse) results in the exposure of cryptic binding sites or increased affinity, thus, increasing the association of binding partners (orange). In this schematic, force is applied through F-actin (red filament) that is bound to rigid ECM (gray) through different binding partners. Examples include binding sites in talin and p130Cas. (B) Force might also decrease dissociation kinetics, for example for integrins and myosin, which have been shown to act as catch bonds. Here, this effect is shown schematically for integrins under tension. (C) The re-organization of structures under force might facilitate the recruitment of binding partners. In this example, actin filaments that are cross-linked at a large angle can only facilitate a small number of cross-links (green) between both filaments. Parallel filaments, however, allow the formation of a much higher number of cross-links to be formed between both filaments. (D) Under tension, actin filaments rupture and expose barbed ends, a process that leads to the recruitment of barbed-end binding proteins (blue), such as VASP, mDia or capping protein.