Research

Cells receive and transmit mechanical and biochemical information through contacts they make with other cells and the extracellular matrix (ECM). External and internal cues drive cell and tissue behaviors such as differentiation, morphogenesis, and migration, which play a role in physiological and pathophysiological processes. In particular, an incomplete understanding of how cell adhesions sense, integrate, and respond to physical signals is impeding efforts in developing therapies for mechanosensitive diseases or generating functional engineered tissues.

Cell-ECM adhesion

Integrin-based adhesions, commonly referred to as focal adhesions (FAs), mediate linkages between the ECM and the force generating actomyosin cytoskeleton. Cell-ECM adhesion plays a major role in many disease states, including fibrosis and cancer metastasis. FAs consist of numerous proteins that serve structural and signaling functions. A core mechanical linkage within the FA is between integrins, talin, and actin, which can be further reinforced by the localization of vinculin. This intact mechanical connection is critical for cells to adequately sense and respond to the physical cues of the ECM.

A number of techniques have been developed to measure the forces that are being generated at FAs. A common method for studying the forces that a whole cell exerts on a substrate is traction force microscopy (TFM). Different TFM methods exist that focus on different aspects of force generation, some of which are easier to implement than others. Another method to measure forces at FAs within cells are FRET-based tension sensors that are genetically encoded within a specific protein. These sensors have been placed within multiple FA proteins (as well as cell-cell adhesion proteins) and have allowed quantification of forces that a single protein experiences. By integrating the information obtained using these and other methods, we can begin to understand how forces are being transmitted between the inside and outside of the cell.

One of the main proteins implicated in transferring forces from the ECM to the cytoskeleton is vinculin. The vinculin head domain is responsible for association with critical FA components, including talin. A major function of the vinculin tail domain is to bind and bundle actin filaments. Vinculin has been previously shown to be critical for many FA functions, such as FA maturation and traction force exertion, and is sensitive to mechanical loading. Using a FRET-based tension sensor placed inside of vinculin, it has been demonstrated that vinculin load is sensitive to cell confinement, adopting a spatial pattern of load in response to different geometries.

Additionally, the novel combination of FRET and FRAP has allowed for measurements of force-sensitive dynamics of vinculin. Vinculin can adopt a force-stabilized state, in which vinculin under high load has slower dynamics than when under low load, at a single FA level. Force-stabilization may occur via a catch-bond discovered to exist between vinculin and actin. The force-stabilized state of vinculin is dependent on the ability of vinculin to bind talin and actin, and on ROCK-mediated contractility. Finally, the force-stabilized state of vinculin is necessary for effective haptotaxis on fibronectin.

There is still much to understand about cell-ECM adhesion, including what other proteins are involved in force transmission, how these forces are being converted into biochemical signals that dictate cell fate, and what components are useful therapeutic targets.

Cell-Cell adhesion

Cells adhere to each other through a few different structures, one of which is the cadherin-mediated adherens junction (AJ). Cell-cell adhesion plays a key role in many morphogenetic and migratory events during development, as well as in disease states such as cardiovascular disease and cancer metastasis. Similarly to FAs, AJs consist of numerous proteins that serve structural and signaling functions. A core mechanical linkage within the AJ is between cadherin, a-catenin, and actin, which can be further reinforced by the localization of vinculin. An intact connection between cells is important for tissue-level coordination.

One important context within which to study cell-cell adhesion is during collective cell migration. During this process, cells must coordinate their behavior to move together, which involves mechanical and biochemical signaling at AJs. Because AJs are coupled to the actomyosin cytoskeleton, much of this signaling also impacts cytoskeletal organization. During many migratory processes, including wound healing, AJs and the actomyosin cytoskeleton are remodelled to form a supracellular actomyosin cable at the leading edge that is anchored by reinforced AJ structures.

It has been demonstrated that removal of AJs from the leading edge of a collectively migrating group of cells is necessary for formation of the supracellular actomyosin cable. Removal of AJs may allow for extra space to assemble new structures or induce localized signaling that promotes actomyosin assembly. Alternatively, the AJ material that is removed from the leading edge may be the same material that accumulates at former tricellular junctions, which may promote assembly and coupling of the cable. The actomyosin cable itself has unique dynamics, with both actin and myosin being stabilized under the higher tension sustained in comparison to stationary tissue. However, actin and myosin dynamics are not completely coupled, with a pool of actin being unaffected by myosin perturbation.

Many questions remain about the nature of cell-cell adhesion. In the context of collective cell migration, it is unclear what mechanical or biochemical signals drive reorganization of the AJs and promote formation of the actomyosin cable. In other contexts, it is unknown how cells convert mechanical stimuli from neighboring cells into biochemical signals, how far into a tissue signals can be transmitted along AJs, or the nature of the feedback between cell-cell adhesion and cell fate.