Organization, dynamics and mechanoregulation of integrin-mediated cell-ECM adhesions

Abstract

The ability of animal cells to sense, adhere to and remodel their local extracellular matrix (ECM) is central to control of cell shape, mechanical responsiveness, motility and signalling, and hence to development, tissue formation, wound healing and the immune response. Cell-ECM interactions occur at various specialized, multi-protein adhesion complexes that serve to physically link the ECM to the cytoskeleton and the intracellular signalling apparatus. This occurs predominantly via clustered transmembrane receptors of the integrin family. Here we review how the interplay of mechanical forces, biochemical signalling and molecular self-organization determines the composition, organization, mechanosensitivity and dynamics of these adhesions. Progress in the identification of core multi-protein modules within the adhesions and characterization of rearrangements of their components in response to force, together with advanced imaging approaches, has improved understanding of adhesion maturation and turnover and the relationships between adhesion structures and functions. Perturbations of adhesion contribute to a broad range of diseases and to age-related dysfunction, thus an improved understanding of their molecular nature may facilitate therapeutic intervention in these conditions.

Figure 1 |. Integrin conformational dynamics at integrin-mediated adhesions.

a | Cartoon of the top view of a spread polarised cell with IACs and actin structures indicated. b | Depiction of key steps in the formation and maturation of IACs from initial nascent adhesions to mature FA. c | General schematic depicting the three major conformations of integrin – bent closed (BC), extended closed (EC) and extended open (EO) – along with key interacting proteins and sites of force application. α-subunit is shown in blue and β-subunit in red. Inhibitors (such as filamin^,, ICAP-1, or SHARPIN) binding to α or β cytoplasmic tails are depicted as yellow circles while activating proteins such as talin and kindlin that directly or indirectly link to the cytoskeleton are shown in green. d | Reaction scheme showing the kinetics of integrin and ligand binding taking into account integrin conformational change – based on results in .

Figure 2 |. Cataloging the Molecular components of IACs.

a | Literature-curated adhesome is assembled from the analysis of published studies describing IAC locations and protein-protein interactions. b | Experimentally-defined adhesomes are obtained from mass spectrometry-based proteomic analysis using various enrichment strategies, such as hypotonic shock, chemical crosslinking, or proximity biotinylation. Enriched IAC-containing fractions are then subjected to tryptic digestion and mass spectrometric analysis. c | Proximity biotinylation enables inference of protein interactions networks. Bait moiety such as BirA*, a promiscuous biotin ligase from Escherichia coli, can be fused to a ‘bait’ protein that localize to IACs. Incubation with biotin results in BirA*-catalyzed biotinylization of nearby ‘prey’ protein within 10–15 nm radius. Biotinylated ‘prey’ proteins can then be affinity-purified and subjected to mass spectrometric analysis. d | Core consensus adhesome. Meta-analysis of literature-curated adhesome and experimentally-defined adhesomes, together with proximity biotinylation data, have largely converged on a set of core consensus adhesome components. Protein-protein interactions are depicted by black lines. Color-coding indicate distinct modular sub-networks.

Figure 3 |. Molecular organisation of IACs.

a - b | Lateral composite nanocluster organisation. In mature FAs, extensive granularity and occasional alignments of nanoclusters into linear chains along actin templates can be observed in certain cell types^,. For integrin α5β1, active and inactive integrin segregate into distinct nanoclusters suggesting that nanoclusters could function as discrete units. Single-molecule analysis of integrin motion provides a consistent picture whereby integrin can diffuse freely through IACs. Integrin is immobilised in distinct loci which are comparatively enriched in IACs. c | Multilaminar architecture of FA. Schematic diagram depicting vertical organisation of various IAC components that have been analysed by SRM studies^,,,. Integrins are depicted in clusters of inactive (bent-closed), active (extended-open), as well as in tilted orientation as inferred from fluorescence anisotropy measurements. d | Podosomes or invadopodia are arrays of typically centrally located IAC structures, consisting of actin-rich core that protrude against and degrade ECM, surrounded by ring-like plaques containing IAC components. e | Vertical organisation of podosome ring featuring polarised orientation of talin, and similar organisation of paxillin and vinculin as in FA. f | Comparison of SRM-based spatial organization with sub-networks of IAC components identified by proteomic analysis (Fig. 2d). A putative IAC compartment anchored by KANK proteins that may potentially interface with microtubules is also depicted.

Figure 4 |. Molecular-scale mechanoregulation of IACs.

a | Mechanotransduction by force-dependent conformational changes. Several IAC proteins such as talin, vinculin, RIAM, α-actinin, KANK1 and filamin A contain domains that can be partially unfolded under piconewton physiological forces^,–. For example, talin R3 domain is a 4-bundle of α-helix that contains a binding site for RIAM. RIAM can bind talin under low intramolecular tension but upon the increase in intramolecular tension, R3 undergoes partial unfolding that displaces RIAM and in turn exposes a binding site to vinculin. b | Integrin-Talin-Actin as the structural and mechanical ‘backbone’ of the IACs. In mature focal adhesions, talin adopt a vertically polarized orientation with N-terminal FERM domain engaging integrin b cytoplasmic tail. Actin binding sites 2 and 3 provide direct linkage to the F-actin cytoskeleton while indirect cross-linking is mediated by vinculin. c | Molecular clutch model of IAC mechanotransduction. Clutch molecules are capable of binding to both integrin and actin filaments. If molecular clutch is disengaged (middle panel), rearward force generated by actin polymerization in lamellipodia is primarily channeled to actin retrograde flow. Upon molecular clutch engagement (bottom panel), actin retrograde flow is mechanically coupled into traction force and support leading edge protrusion. d | Molecular-scale force transmission in IACs. Schematic diagram of force transmission in integrin-talinvinculin-actin complexes. Single-integrin force vector is primarily co-aligned with the F-actin orientation, potentially implicating a significant degree of tilting and stretching of the integrin ectodomain^,. Integrin force is thought to be transmitted via talin, which may serve as a force aggregator as reflected by the intramolecular force gradient due to multiple vinculin and actin binding sites^,. Load-bearing by integrin and talin are variable with high-load bearing molecules (>10 pN) comprising a subpopulation, dependent on vinculin^,. Thus at steady-state only a subset of integrin-talin-vinculin-actin complexes in IACs are fully tensioned.