Dualistic nature of adhesive protein function: fibronectin and its biologically active peptide fragments can autoinhibit fibronectin function.

Abstract

Fibronectin and certain polypeptide regions of this adhesive glycoprotein mediate cell attachment and spreading on various substrates. We explored the theoretical prediction that this adhesive protein could become a competitive inhibitor of fibronectin-mediated processes if present in solution at appropriately high concentrations. Fibronectin function was inhibited by purified plasma fibronectin at 5-10 mg/ml, by a 75,000-dalton cell-interaction fragment of the protein at 0.5-1 mg/ml, and even by two synthetic peptides containing a conserved, hydrophilic amino acid sequence at 0.1-0.5 mg/ml . Inhibition of fibronectindependent cell spreading was dose dependent, noncytotoxic, and reversible . It was competitive in nature, since increased quantities of substrate-adsorbed fibronectin or longer incubation periods decreased the inhibition. A peptide inhibitory for fibronectin-mediated cell spreading also inhibited fibronectin-mediated attachment of cells to type I Collagen, but it did not affect concanavalin A-mediated spreading .These results demonstrate the potential of a cell adhesion molecule and its biologically active peptide fragments to act as competitive inhibitors, and they suggest that fibronectin may act by binding to a saturable cell surface receptor.Cell adhesion to other cells or to substrates is a complex process which is still poorly understood at the molecular level (e.g., references 1 and 2). Adhesive events mediated by fibronectin and other extracellular attachment proteins provide experimental systems for analyzing polypeptide domains that mediate binding and adhesive functions (3-9). For example, a region offibronectintermed the "cell-binding" region, which interacts with the cell surface to mediate cell attachment and spreading on substrates, has been identified and localized to polypeptide fragments (10, 11). Even a synthetic peptide from this region of fibronectin can mediate cell attachment to a plastic substrate (12).Theoretically, a cell adhesion protein might become a specific inhibitor of its own function if it were bound in excess to a cellular receptor. Saturation of receptors by soluble adhesive molecules might prevent receptor interactions with substrate-or cell-attached adhesive molecules . A rough analogy might be the prozone effect in immunoprecipitation, in which an excess ofantibody can saturate sites on antigens and prevent the formation ofaggregates.THE JOURNAL OF CELL BIOLOGY " VOLUME 99 JULY 1984 [29][30][31][32][33][34][35][36] We tested this theoretical model with soluble plasma fibronectin and certain peptide fragments of this glycoprotein . These molecules were found to be competitive, reversible inhibitors of fibronectin function . Our results suggest that adhesive proteins can have a dualistic nature, mediating positive or negative effects depending on their concentrations and whether they are in solution or attached to a substrate . MATERIALS AND METHODSFibronectin and Fibronectin Fragments : Plasma fibronectin was isolated from human plasma (National Institu...

Influence of substratum surface chemistry/energy and topography on the human fetal osteoblastic cell line hFOB 1.19: Phenotypic and genotypic responses observed in vitro☆

“…Adherent cells then slowly (typically within hours) spread over the surface, depending on compatibility with the surface, expressing a strong 'biological component of adhesion' [38] that includes secretion of extracellular matrix (ECM) and results in the flattening of cells on the substratum [2] (spreadcell length is about 3-10 times height [9]). Needless to say, (protein) composition of the fluid phase can greatly affect the entire cell adhesion process [31,33,[55][56][57][58][59]. Thus, it is apparent that the short-term (<3 days) cell adhesion/proliferation process spans a broad range of time and length scales.…”

An RGD Motif Present in Cadherin 17 Induces Integrin Activation and Tumor Growth

“…The tripeptide RGD motif is a major recognition site for integrins within some ligands, such as fibronectin or fibrinogen (11,12). However, although ␣2␤1 integrin contains an RGD-binding site, their natural ligands, collagens type I and type IV, used a different binding sequence (GFOGER) (13) and are classified as RGD-independent ligands (14,15).…”

Antimetastatic effects of synthetic polypeptides containing repeated structures of the cell adhesive Arg-Gly-Asp (RGD) and Tyr-Ile-Gly-Ser-Arg (YIGSR) sequences

“…Common or characteristic core sequences in cell adhesion molecules such as fibronectin (Kornblihtt et al, 1985), vitronectin (Suzuki et al, 1985) and laminin Sasaki have been found to contribute to cell adhesion, and to the spread or migration of cells (Yamada & Kennedy, 1984;McCarthy & Furcht, 1984;Humphries et al, 1986). It has been shown that the domain of fibronectin in cellular recognition is carried by an ArgGly-Asp-Ser (RGDS) sequence.…”

“…The RGD sequence exists commonly in many adhesion molecules (Pierschbacher & Ruoslahti, 1982. RGD-containing peptides have been shown to promote cell adhesive capability after their surface immobilisation, and to inhibit cell adhesion to fibronectin when added freely in solution (Yamada & Kennedy, 1984Akiyama & Yamada, 1985).…”