Synergistic control of cell adhesion by integrins and syndecans

Abstract

The ability of cells to adhere to each other and to their surrounding extracellular matrices is essential for a multicellular existence. Adhesion provides physical support for cells, regulates cell positioning and enables microenvironmental sensing. The integrins and the syndecans are two adhesion receptor families that mediate adhesion, but their relative and functional contributions to cell-extracellular matrix interactions remain obscure. Recent advances have highlighted connections between the signalling networks that are controlled by these families of receptors. Here we survey the evidence that synergistic signalling is involved in controlling adhesive function and the regulation of cell behaviour in response to the external environment.

Figure 1. Domain structures of integrin and syndecan-4, transmembrane matrix receptors

Integrins consist of α,β heterodimers, comprising large extracellular and short cytoplasmic domains^, . The extracellular domain structure of each subunit is conserved between isoforms, with the exception of a subset of α-subunits (α₁, α₂, α₁₀, α₁₁, α_X, α_M, α_L, α_D and α_E) that include an inserted “A-domain” within the ligand binding pocket. The cytoplasmic interactions of integrins are mostly mediated by the β-subunit tail, most notably by recruitment of the cytoskeletal protein talin. Syndecans exist as homodimers and bind to the matrix through glycosaminoglycan chains substituted at 3-5 positions on the extracellular domain. The short cytoplasmic domains can be subdivided into two conserved regions, C1 and C2, that bind a Src/Fyn tyrosine kinase complex and PDZ-domain-containing proteins, respectively, and a central variable region. The variable region confers specific properties on each syndecan, and most notably is a PKCα binding site in syndecan-4.

Figure 2. Adhesion formation is dependent on engagement of syndecan-4

Unlike cells plated onto fibronectin (a), cells plated onto an integrin ligand spread but fail to form FA (b) unless stimulated with an antibody against syndecan-4 (c) or a syndecan-binding fragment of fibronectin (d). Adhesion contact formation, in response to a syndecan-4 ligand, can be blocked by disrupting expression of syndecan-4 (e) or PKCα (f), highlighting the specific role of syndecan-4 in adhesion contact formation and the immediate relationship with classical adhesion signalling pathways. Images portray fixed fibroblasts stained for the adhesion marker vinculin (green) and actin (red), and arrows indicate focal adhesions.

Figure 3. Protein kinase signalling depends on synergy between integrin and syndecan-4 and regulates endocytosis

Syndecan-4 regulates PKCα and tyrosine kinases, FAK and Src, in synergy with α₅β₁ integrin. Activation of PKCα depends on direct association (solid arrows) with the syndecan-4 cytoplasmic domain and PIP₂, and can be prevented by the phosphorylation of syndecan-4 by PKCδ. Both PKCα and FAK play key roles in cell migration due to the regulation of clathrin-dependent endocytosis of α₅β₁-integrin, which is mediated by another syndecan-4-binding protein, dynamin. The contributions of PKCα and FAK to integrin endocytosis are long-range effects (dotted arrows), and do not depend on direct association between α₅β₁-integrin and syndecan-4. Endocytosis of the syndecan itself depends on association between the C terminus of the syndecan cytoplasmic domain and one of the paired PDZ domains of the small scaffolding protein syntenin.

Figure 4. Adhesion-dependent GTPase signalling depends on synergy between integrin and syndecan-4

Efficient activation of the cytoskeletal regulators Rac and Rho, in response to fibronectin, depends on engagement of syndecan-4 by the ECM. Direct association (solid arrows) between PKCα and the cytoplasmic domain of syndecan-4 is necessary for GTP-loading of both Rac and Rho, and the activation of Rac also depends on the PDZ-binding motif of syndecan-4. ECM engagement of integrins causes redistribution of cholesterol into detergent insoluble membrane microdomains, and is necessary for the redistribution of GTP-Rac to the membrane where it associates with downstream effectors, such as PAK. It is the localised activation of Rac and Rho that initiates signalling cascades (dotted arrows) that result in the organisation of the actin cytoskeleton.

Figure 5. Interplay between β3 integrin, VEGFR and syndecan function in vascularisation

Perturbing α_vβ₃ function in vivo can have remarkably different effects on pathological angiogenesis (A) depending on the nature of the perturbation and the cell type expressing the integrin (tumour (Blue) vs. host (Red)). Antagonism of α_vβ₃, or expression of mutant β₃ subunits with compromised signalling capabilities, inhibits angiogenesis (seemingly through the modulation of different pathways), whilst β₃-integrin-deficient mice exhibit enhanced pathological angiogenesis. Further characterisation of these phenomena has demonstrated an important link between α_vβ₃ function and VEGF/VEGFR signalling. In vitro studies suggest that syndecan-1 has the capacity to differentially regulate this α_vβ₃ and VEGF/VEGFR signalling network. There would also appear to be fundamental differences between α_vβ₃-dependent regulation of pathological angiogenesis (A) and developmental neovascularisation (B); inhibition of β₃ ligand-binding can disrupt developmental vascularisation whereas perturbation of β₃ expression or signalling has no effect. Syndecan-1 (and syndecan-4) deficient mice, like β₃-integrin null mice, appear developmentally normal, suggesting that they do not exhibit significant developmental angiogenic defects.

Figure 6. Models of the complexity of integrin-syndecan synergy in vivo

In each model, depending on the cellular context, molecules A, B, C and D may correspond to different proteins (see Key) A) Ternary complex formation. Syndecan associates with cell surface receptor and its ligand (either soluble or membrane-bound), and generates intracellular signals. ECM engagement of integrins, and possibly syndecans, is required to potentiate the syndecan-mediated signals. B) Multimeric complex formation. Cell surface receptors for chemoattractive, repulsive or growth factors are central to the formation of complexes with their ligands, syndecans, integrins and ECM molecules. Cooperative intracellular signals are generated from each receptor, initiating a coordinated cellular response. C) Syndecan-mediated ligand-presentation in trans. Syndecan on an adjacent cell binds to and localises a soluble ligand, to allow interaction with its receptor (possibly in complex with an integrin). Such interactions could initiate both syndecan and integrin signalling events. Integrin and syndecan activity in this model may or may not require association with ECM molecules D) Syndecan-mediated ligand-presentation in cis. Syndecans, viewed as pioneering molecules, bind to and localise chemoattractive/repulsive/growth factors, to allow interaction with their receptors and integrins on the same cell. Collaborative intracellular signals may be elicited from both syndecans and integrins in such a model, but may also require interaction with ECM molecules