How tensins transform focal adhesions into fibrillar adhesions and phase separate to form new adhesion signalling hubs.

Tissues in our body are made up of cells and surrounding fibrillar material, the extracellular matrix or ECM. The cells attach to, sense and reorganise the ECM, an ability which is particularly important during the development of organisms, in diseases and in regeneration processes, which all require specific cellular responses to changing ECM environments. Cellular responses include changes in their ability to move (e.g. closing of wounds), changes in cell growth, and also in synthesising new or reshaping their old ECM environment. Many studies have focused on how cells sense the ECM, but we are still far from understanding how this information is translated into signals that promote specific cellular responses. Understanding this process is critically important if we want to get a step closer to treating the roots of diseases and promoting regeneration and recovery.

Cells sense their ECM environment by grabbing and pulling the neighbouring extracellular fibrillar material using proteins called integrins, which extend across the cell surface from inside to outside. Integrins both bind to the ECM externally and to proteins inside the cells. These proteins couple integrins to the actin cytoskeleton, a contractile network of intracellular fibres, which upon reorganisation induces pulling forces and cell movement. Our previous results show that two proteins that couple integrins to actin, called talin and vinculin, are central to sensing environmental changes. Recently we have shown that the association of talin with other proteins, tensins, is critical for ECM reorganisation. We now have further data showing that a gradual increase in the concentration of tensins in specific areas in cells can lead them to form little droplets or condensates. We hypothesise that these tensin condensates can also attract other proteins that have a critical role in the regulation of cell adhesion and cell migration.

Testing this hypothesis requires detailed information on how proteins bind to each other and how these impact on specific cellular functions. To overcome this gap in our understanding, we propose a joint effort of two laboratories that have vast expertise in the field of cell adhesion regulation but apply very different research methods. The Barsukov laboratory determines details of a protein structure, which is critical for the understanding how proteins bind to each other. Structural details help to design small changes (mutations) in proteins with the aim of experimentally blocking specific proteins interactions. The Ballestrem laboratory has developed powerful methods in cell biology and microscopy that monitor protein interactions in cells and investigate what role these interactions play in determining cell behaviour in the face of changing ECM environments. Both teams have already worked successfully together and revealed essential mechanisms that control cell-matrix sensing and ECM organisation.

The proposed research aims to understand (i) how tensins interact with talin and what role this interaction has in enriching tensins in cell-matrix adhesion sites that are involved in ECM reorganisation; (ii) how tensins condense to droplets that can attract other proteins to form molecular reservoirs that regulate cell adhesion and migration; (iii) how tensin binding to integrins is regulated and how this affects the binding strength of integrins to the ECM. Ultimately, the knowledge gained will open a pathway to the development of new ways to prevent diseases (e.g. cancer, fibrosis) and promote regeneration (wound healing).

Cells interact with the extracellular matrix (ECM) through transmembrane adhesion receptors (integrins) that are linked to signalling proteins that regulate cell migration and ECM remodelling/synthesis. The cell-matrix adhesion complexes can vary in their molecular composition and in their function. Published data suggest that adapter proteins such as talin that couples integrins to actomyosin localise to force exerting focal adhesions (FA), whereas tensins mark a subset of adhesions called fibrillar adhesions (FB) that are linked to the remodelling of the cell ECM. FBs originate from FAs and here we aim to understand how molecular interactions lead to this transition.

Tensins bind to integrins and we have recently shown the association of two members of the tensin protein family (tensin1 and 3) with talin. Our pilot data suggest that talin contains multiple tensin binding sites and that they have an important role in the enrichment of tensins in FBs. Additional data show that the local enrichment of tensins leads to condensates through liquid-liquid phase separation (LLPS). We hypothesise that tensin enrichment and engagement with subsets of integrins is critical for the formation of FBs and that LLPS forms condensates that can act as a reservoir of signalling proteins. The use of structural biology combined with cell biology and advanced fluorescence microscopy will enable us to gain in-depth knowledge of how the different interactions are accompanied by structural changes of proteins in time and space and what consequences this has for the development of specific adhesion sites and downstream signalling pathways.

Insights from these studies may ultimately lead to the development of new strategies to prevent disease and promote tissue regeneration.