Cell Adhesion to Fibrillin-1 Molecules and Microfibrils Is Mediated by α5β1 and αvβ3 Integrins

Abstract

Fibrillins are the major glycoprotein components of microfibrils that form a template for tropoelastin during elastic fibrillogenesis. We have examined cell adhesion to assembled purified microfibrils, and its molecular basis. Human dermal fibroblasts exhibited Arg-Gly-Asp and cation-dependent adhesion to microfibrils and recombinant fibrillin-1 protein fragments. Strong integrin ␣ 5 ␤ 1 interactions with fibrillin ligands were identified, but integrin ␣ v ␤ 3 also contributed to cell adhesion. Fluorescence-activated cell sorting analysis confirmed the presence of abundant ␣ 5 ␤ 1 and some ␣ v ␤ 3 receptors on these cells. Adhesion to microfibrils and to Arg-Gly-Aspcontaining fibrillin-1 protein fragments induced signaling events that led to cell spreading, altered cytoskeletal organization, and enhanced extracellular fibrillin-1 deposition. Differences in cell shape when plated on fibrillin or fibronectin implied substrate-specific ␣ 5 ␤ 1 -mediated cellular responses. An Arg-Gly-Asp-independent cell adhesion sequence was also identified within fibrillin-1. Adhesion and spreading of smooth muscle cells on fibrillin ligands was enhanced by antibody-induced ␤ 1 integrin activation. A375-SM melanoma cells bound Arg-Gly-Asp-containing fibrillin-1 protein fragments mainly through ␣ v ␤ 3 , whereas HT1080 cells used mainly ␣ 5 ␤ 1 . This study has shown that fibrillin microfibrils mediate cell adhesion, that ␣ 5 ␤ 1 and ␣ v ␤ 3 are both important but cell-specific fibrillin-1 receptors, and that cellular interactions with fibrillin-1 influence cell behavior.Fibrillins are large cysteine-rich glycoproteins and the major constituents of fibrillin-rich microfibrils of the extracellular matrix (ECM) 1 (1, 2). They are multidomain molecules, containing 47 epidermal growth factor (EGF)-like domains and 8-cysteine (TB) modules. Fibrillin-1 and fibrillin-2 are both expressed during fetal development, but fibrillin-1 isoform is by far the most abundant isoform in adult tissues (3, 4). The possibility that fibrillins interact with cell receptors was suggested by electron microscopy of vascular and other tissues showing juxtaposition of extracellular microfibrils and cellular dense plaques (5) and by the discovery that fibrillins contain the Arg-Gly-Asp (RGD) putative cell attachment motif (6). Integrins are ␣␤ heterodimeric transmembrane receptors that mediate cell adhesion to the ECM, usually through the characteristic RGD motif, and have widespread essential functions in development, tissue organization, and the immune system (7). Integrin-mediated cell interactions regulate cell adhesion and migration and initiate signaling pathways that lead to reorganization of actin cytoskeleton and cellular proliferative and secretory responses.Fibrillin-1 contains one RGD sequence in the fourth TB module (6). Fibrillin-2 contains two RGD sequences, one in the same position as the fibrillin-1 RGD sequence in the fourth TB module and the other in the third TB module where it is surrounded by hydrophobic amino acids (8). Fibrillin-3 also ...

New insights into elastic fiber assembly

“…Fibrillin-1 isolated from human fibroblasts appears as linear fibrils with a beaded periodicity (Sakai et al, 1991). Fibrillins have ArgGly-Asp (RGD) sequences that interact with integrins (Pfaff et al, 1996;Sakamoto et al, 1996;Bax et al, 2003) and heparin-binding domains that interact with cell surface heparan sulfate proteoglycans (Tiedemann et al, 2001;Ritty et al, 2003), suggesting that fibrillins directly signal cells through these receptors. Furthermore, in vivo assembly of fibrillin may require cell-surface receptors in a similar manner to fibronectin (Wu et al, 1995).…”

Tissue elasticity and the ageing elastic fibre

“…iii In the core of the mature elastic fibre, ultrastructural analyses reveal twisted rope-like structures of highly cross-linked elastin (Ronchetti et al 1998). Fibrillin microfibrils are mostly located at the microfibril periphery, where they can interact with cellular integrins via an RGD site on fibrillin-1 (Bax et al 2003). b The structure of tropoelastin consists of alternating hydrophobic and cross-linking domains.…”

“…Each domain is stabilised by three cys-cys disulphide bonds (indicated in blue on the ribbon model of two contiguous fibrillin-1 cbEGF domains) and by a single bound Ca 2+ (Downing et al 1996;Wess et al 1998). For interpretation of coloured elements within this figure please refer to the electronic version of the manuscript fibrillin microfibrils communicate with cells via an RGD (Arg-Gly-Asp) recognition site, which is bound by α 5 β 1 and α v β 3 integrins (Bax et al 2003). The importance of fibrillin in maintaining tissue function is highlighted by the severe ocular, skeletal and cardiovascular pathologies experienced by individuals suffering from Marfan syndrome, a heritable connective tissue disorder caused by mutations in fibrillin-1 (Robinson and Booms 2001).…”

Xenopus fibrillin is expressed in the organizer and is the earliest component of matrix at the developing notochord‐somite boundary

“…One role is to multimerize into microfibrils, which can perform biomechanical roles in the body (Robinson and Godfrey, 2000). Fibrillins 1 and 2 have been shown to be ligands for both integrin and proteoglycan cell surface receptors (Pfaff et al, 1996;Sakamoto et al, 1996;Ritty et al, 2003;Bax et al, 2003;Cain et al, 2005). These interactions likely mechanically link cells to fibrillin containing matrix (Czirok et al, 2004), and signaling functions through these receptors are possible based on analogy with other matrix molecules such as laminins (Hopker et al, 1999).…”