Histol Histopathol (1997) 12 : 233-240 Histology and 001: 10.14670/HH-12.233 Histopathology http://www.hh.um.es From Cell Biology to Tissue Engineering Invited Review Roles of integrins in fibronectin matrix assembly C.Wu Department of Biochemistry and Molecular Biology, Samuel C. Johnson Medical Research Center, Mayo Clinic Scottsdale, 13400 E. Shea Blvd, Scottsdale, USA Summary. F ibro nectin (Fn ) ma trix assem b ly is a in man). Only with wounding or during he mostasis is Fn dynamic cellular process in whi ch the soluble dimeri c Fn expressed locall y and deposited into extracellular matrix. molecules are assembled into insoluble, d isul fide bond Cell s not onl y respond to signals fro m Fn matrix but also stabil ized fi brillar polymeric matrix. Fn matrix assembly actively partic ipate in the fo rmati on of a Fn ma tri x. requires s pec ific Fn binding integrin s . Severa l Fn Recent studies suggest that specific cell surface Fn binding integrins that are capable of mediating Fn matrix binding integrins play important roles in initiation and assembly have been ident ified . They include a5B I , regulati on of fibronectin matrix assembly. In this review, a llbB3 and avB3 integrins. Cell s regula te the matrix I will provide a brief overview of recent advances in our assembly process not onl y by controlling cell surface unde rsta nd ing of the ro les of integrins in Fn matri x expression level of the Fn binding integrins but also by assembly. mo dul a ting F n binding a nd cy tos ke le to n b inding acti vit ies of the integrins. A major cha lle nge of future Fn domains involved in Fn matrix assembly studies is to delineate the signal transducti on pathway that regulates Fn matrix assembly. Fns a re modular, multido ma in g lycopro te ins of approx imately 500,000 daltons consisting of repeating Key words: Integrins, Fibronectins, Extracellular matri x homologies of three types (Type I, II and III ) (Fig. I) assembly, Signal transducti on (Hynes, 1990). Each F n molecule contains two similar di sul fide bonded subunits with d iffe re nces created by RNA splic ing, a nd by g lycosy lation a nd othe r post­ Introduction translati onal modificati ons (Hynes, 1990). The disul fide­ bonded dimer structure of Fn is c ri tical fo r Fn matri x Fibronectins (Fns) are cell adhesive g lycoproteins assembly (Schwarzbauer, 199 1; Ichihara et al. , 1995). In tha t pl ay c riti cal ro les in m a ny bi o log ical a nd addition , the first five type I re peats (29 kDa Matri x pathological processes including embryogenesis, wound Assembly Site, Fig . I ) (McKeown-Longo and Mosher, healing, metastasis, fibrosis a nd thro mbosis (Hynes, 1985; Quade and McDonald , 1988; Schwarzbauer, 199 1; 1990). During organogenesis, Fns are major constituents Sott il e a nd Wiley, 1994), the first ty pe III re peat of the extracellular matrix. Inactivation of the mouse Fn (Homophilic Interacti ve Site , Fig. I) (Chernousov et al. , gene caused early e mbryonic lethality (George e t al. , 199 1; Mori a and Ruoslahti , 1992), and the arg-gly-asp 1993) , de mo ns trating tha t Fn is ind is pe nsable for (RGD) containing integrin binding domain are important verte bra te e mbryogenesis. F n matrix assembly is a fo r Fn matrix assembly (McDonald, 1988; McDonald et dynamic cellular process in which the soluble dimeric Fn al. , 1987; Mosher et al. , 1992; Hocking et al. , 1996). The molecules are assembled into in soluble , disul fi de bond first five type I re peats of Fn that apparently fold to form stabili zed fibrillar polyme ri c matri x (McDonald , 1988; a high affinity binding site fo r specific components on Mosher et al. , 1992). Matri x Fn promotes cell adhesion, the surface of matrix fo rming cell s (Limper et al. , 199 1; mi grati on and di ffe re ntiati on , and thus is the primary Sottile et al. , 199 1; Moon et al. , 1994). Fn fragments that func ti onal form o f the protein. The assembly o f a Fn inc lude thi s d o m a in inhibit m a tri x forma tio n b y matrix is both temporall y and spati all y regulated in vivo. f ibro blasts ( Mc Keow n- Lo ngo a nd M osher, 1985; For example, in adult mammals, most Fn is synthesized McDonald et al. , 1987 ; Quade and McDonald , 1988), by the li ver a nd exists primaril y as a soluble pl asma resulting in a specific phenotype characteri zed by short prote in present at hi gh concentration (about 300 mg/ml stitches of cell surface Fn associated with a5BI integrin receptors. The first five type I repeats of Fn are also Offprint requests to: Chuanyue Wu , Ph .D., Department of Cell Biology, required fo r de novo assembly of recombinant Fns into Volker Hall, 1670 University Boulevard, The University of Alabama at the extracellular matrix (Schwarzbauer, 199 1; Sottile and Birm ingham. Birmingham , AL 35294, USA Wiley, 1994). Antibodies recogni zing the first type III 234 /ntegrins in matrix assembly repeat inhibit Fn binding and matrix assembly by Fn-binding integrins fibroblasts (Chernousov et a!., 1991). However, recombinant Fn lacking the first type III repeat can be Integrins are aB heterodimeric transmembrane assembled into the matrix (Schwarzbauer, 1991; glycoproteins that interact with extracellular (or other Ichihara-Tanaka et aI., 1992), suggesting that this site is cell surface) molecules and cytoplasmic molecules not absolutely required for Fn matrix assembly. The first including cytoskeletal and catalytic signaling proteins five type I repeats bind to the first several type III (Ruoslahti, 1988; Hynes, 1992; Schwartz et aI., 1995). repeats and it has been proposed that this interaction Integrins mediate cell-cell and cell-extracellular matrix may be involved in regulation of Fn-Fn interactions interactions and have been implicated in many biological (Aguirre et a!., 1994). Indeed, a specific peptide and pathological processes including embryogenesis, sequence derived from the first type m repeat induces wound healing, inflammation and cancer. Several disulfide crosslinking of Fn into high molecular weight integrins, including aSB1, a4BI , avBI , aIIbB3 and multimers in vitro (Moria and Ruoslahti, 1992; Moria et avB3, bind Fns. Most of the Fn-binding integrins (aSB1 , a!., 1994) . However, attempts to demonstrate the avBI, alIbB3 and avB3) recognize the RGD containing presence of a Fn fragment equivalent to this peptide cell binding domain. In addition to the common RGD under physiological condition have been unsuccessful sequence, distinctive sequences (synergy regions) (Moria et aI., 1994). In addition, it has been shown that located amino-terminal to the RGD sequence also the heat-denatured first type III module, but not the contribute to the binding of Fn to aSB1 integrin (Aota et native form of the module, binds to the 70 kDa amino aI., 1991, 1994) and alIbB3 integrin (Bowditch et a I. , terminal fragment of Fn (Hocking et aI. , 1994). These 1991 , 1994). In contrast to aSBI, avBI, alIbB3 and results suggest that during Fn matrix assembly in avB3, a4B1 integrin recognizes the CS I (or V2S) site cultured cells and in vivo, one or more earlier events are within the alternatively spliced IIICS (or V) region of Fn required to unmask putative homophilic binding sites on (Fig. 1) (Wayner et aI., 1989; Guan and Hynes, 1990; Fn. One of the earliest events in Fn matrix assembly by Mould et aI. , 1990). cu ltured cells appears to be the binding of Fn to cell The ligand binding affinity of a given integrin may surface mediated by interactions between the RGD vary depending on its cellular environment. For containing cell binding domain of Fn and specific Fn example, aSBI integrin binds Fn with high affinity binding integrins (Wu et a!., 1993, 1995c, 1996; (Kd~O.1 p.M) in CHO cells but it binds Fn with low Hocking et aI. , 1996). Antibodies to the RGD containing affinity (Kd >1 pM) in human KS62 erythroleukemia cell binding domain and Fn fragments containing the cell cells (O'Toole et aI. , 1994). The effect of cellular binding domain inhibit Fn matrix assembly by environment on integrin ligand binding affinity is fibroblasts, resulting in fewer fibrils that are of normal dependent on specific types of integrins, as another Fn­ length, consistent with a role for this site in initiation of binding integrin, alIbB3, has low affinity ligand binding Fn matrix assembly in these cells (McDonald et aI., in CHO cells (O'Toole et aI., 1994). This is similar to 1987; Roman et aI., 1989). aIIbB3 integrin on resting circulation platelets, which Homophilic Interactive Site cao- SS EDlIl-8 Illes S S or V -aac.l..l..fnL Heparin I Collagen Binding Heparin II ~ Domain o Type I (45 aa) Homologous o Type II (60 aa) Repeats Type III (90 aa) Fig. 1. Fn o domain structure. A Variably Spliced U 235 Integrins in matrix assembly also has a low affinity binding state. The ligand binding Integrin-cytoplasmic molecule interactions affinity of integrins can be controlled from within the cells ("inside-ollt signaling" ) (Ginsberg et aI., 1992; Tntegrins are capable of interacting with various Hynes, 1992; Schwartz et aI., 1995). Recent studies have cytoskeletal and catalytic signaling proteins molecules demonstrated that the integrin cytoplasmic domains (Fig. 2). The major cytoskeleton-binding sites have been mediate inside-out signal transduction. One of the best located to the 13 cytoplasmic domain of integrins (Sastry studied integrins in term of inside-out signal and Horwitz, 1993).