Suprastructural specificity of integrin-mediated cell-matrix-interactions: Relation to cartilage homeostasis and osteoarthritis
Final Report Abstract
Molecular cartilage collagens, but not their fibrils, are binding substrates for α1β1-, α2β1-, and α10β1-integrins. This implies that the biological importance of integrin-binding to collagens is unrelated to mechanoreception, at least in chondrocytes. Possible functions of integrin-binding include the recognition of collagenous proteins after tissue degradation during wound healing or remodelling, early stages of fibrillogenesis or even to very loosely packed fibrils arising in other scenarios. Previously, the recombinant mini-collagen has served a great deal to elucidate α2β1 integrin function on cells after it has been immobilized to surfaces. This mini-collagen FC3 is a monovalent and soluble α2β1 integrin ligand. This is in contrast to collagen-I which forms supramolecular complexes. Moreover, being insoluble, it forms a solid phase border to the surrounding solvent phase from which cells approach the collagen fibrils with their collagen receptors, among them α2β1 integrin. Monomeric FC3 in solution antagonistically blocks α2β1 integrin-triggered signaling in platelets. By immobilizing the monovalent, monomeric FC3 to a surface, it becomes a member of physically connected linkage group, similar to the polyvalent supramolecular aggregates of collagen and agonistically triggers platelet signaling. By reducing the surface/linkage group into the colloidal/nanometer range, we discovered that about 120 integrins must be clustered within an area of about 15700 nm2 to elicit an α2β1 integrin signal within platelets. This can be achieved by coating 100 nm beads with FC3 molecules at a density of about 12100 µm-2, representing an intermolecular distance of around 9-10 nm. Platelet activation decreases with increasing intermolecular distances of integrin binding sites and eventually ceases at distances beyond 40 nm. Such a nano-scale array of α2β1 integrin binding sites within a linkage group exists only within supramolecular aggregates of collagen, whereby most likely the lateral association of collagen molecules is more relevant for integrin-mediated cell signaling than their staggered array with a D-distance of 67 nm. However, this lateral presentation of integrin binding sites has its limitation as well. Only FC3-covered beads of a diameter of 100 nm, but not of 500nm, elicit full platelet activation. This is in line with the observation that thick collagen-containing fibrils bind their corresponding integrin receptors poorly. Only thinner collagen fibrils in a diameter range of below 500 nm, which occur in tissue during wound healing, remodelling and early stage fibrillogenesis, are good substrate for integrin binding and signalling. Moreover, our results suggest that the diameter of collagen-containing fibrils likely is another very important physiological parameter which determines integrin-mediated adhesion, mechanotransduction, and signalling. To be an agonistic integrin-ligand, the collagen-containg fibrils must contain nano-arrays of α2β1 integrin binding sites which must cover a certain optimum contact area (between 15700 nm2 and 392700 nm2) and must have a certain density of binding sites (i.e. below a certain intermolecular distance), so that more than 120 integrins can cluster in a closest array in the platelet membrane. Then, signaling will be triggered by the integrins. The α2β1 integrin signaling is distinct from signaling via the other collagen receptor GPVI on platelets both kinetically and by using a different tuning of the signaling molecules, FAK and Src.
Publications
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(2012) Integrin alpha7beta1 is a redox-regulated target of hydrogen peroxide in vascular smooth muscle cell adhesion. Free Radic Biol Med 53: 521-531
de Rezende FF, Martins Lima A, Niland S, Wittig I, Heide H, Schroder K, Eble JA
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(2013) A novel flow cytometry-based platelet aggregation assay. Blood 121: e70-80
De Cuyper IM, Meinders M, van de Vijver E, de Korte D, Porcelijn L, de Haas M, Eble JA, Seeger K, Rutella S, Pagliara D, Kuijpers TW, Verhoeven AJ, van den Berg TK, Gutierrez L
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(2014) Collagen XXII binds to collagen-binding integrins via the novel motifs GLQGER and GFKGER. Biochem J 459: 217-227
Zwolanek D, Veit G, Eble JA, Gullberg D, Ruggiero F, Heino J, Meier M, Stetefeld J, Koch M
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(2014) Preserved bioactivity and tunable release of a SDF1-GPVI bi-specific protein using photo-crosslinked PEGda hydrogels. Biomaterials 35: 7180-7187
Schesny MK, Monaghan M, Bindermann AH, Freund D, Seifert M, Eble JA, Vogel S, Gawaz MP, Hinderer S, Schenke-Layland K
