The Human Integrin α8β1 Functions as a Receptor for Tenascin, Fibronectin, and Vitronectin

Lynn M Schnapp,Nan Hatch,Daniel M Ramos,Irina V Klimanskaya,Dean Sheppard,Robert Pytela

doi:10.1074/jbc.270.39.23196

Lynn M Schnapp, Nan Hatch + Show 4 more

Open Access

https://doi.org/10.1074/jbc.270.39.23196

Copy DOI

Abstract

The integrin family of adhesion receptors consists of at least 21 heterodimeric transmembrane proteins that differ in their tissue distribution and ligand specificity. The recently identified alpha 8 integrin subunit associates with beta 1 and is predominantly expressed in smooth muscle and other contractile cells in adult tissues, and in mesenchymal and neural cells during development. We now show that alpha 8 beta 1 specifically localizes to focal contacts in cells plated on the extracellular matrix proteins fibronectin or vitronectin. In addition we show that human embryonic kidney cells (293), transfected with alpha 8 cDNA, express alpha 8 beta 1 on their surface and use this receptor for adhesion to fibronectin and vitronectin. Furthermore, alpha 8 beta 1 binds to both fibronectin- and vitronectin-Sepharose and can be specifically eluted from either matrix protein by the arginine-glycine-aspartic acid (RGD)-containing peptide, GRGDSP. Because fibronectin and vitronectin adhesion appeared to be mediated by RGD, we examined additional RGD-containing proteins, including tenascin, fibrinogen, thrombospondin, osteopontin, and denatured collagen type I. We found that only tenascin was able to mediate adhesion of alpha 8-transfected 293 cells. By using recombinant fragments of tenascin in adhesion assays, we were able to localize the alpha 8 beta 1 binding domain of tenascin to the RGD-containing third fibronectin type III repeat. These data strongly suggest that tenascin, fibronectin, and vitronectin are ligands for alpha 8 beta 1 and that this integrin binds to the RGD site in each of these ligands through mechanisms that are distinct and separate from alpha 5- and alpha v-containing integrins.

Highlights

Integrins are a class of cell adhesion glycoproteins composed of two noncovalently associated subunits, ␣ and ␤
Focal contact localization of integrins is liganddependent; i.e. a particular integrin will accumulate at focal contacts only when its ligand is present in the substrate [18, 19]
In our experiments we used a human smooth muscle cell line (HISM) and a rat embryo fibroblast (REF) cell line, which we found to express ␣8␤1 by immunoprecipitation (Fig. 1, lanes 1 and 2)

Summary

Introduction

Integrins are a class of cell adhesion glycoproteins composed of two noncovalently associated subunits, ␣ and ␤. Cell Adhesion Assays—Non-tissue culture-treated plates were coated with increasing concentrations (0.3, 1, 3, 10, 20 ␮g/ml) of fibronectin, vitronectin, thrombospondin, osteopontin, denatured collagen type I, fibrinogen, and intact tenascin. ␣8␤1: Tenascin, Fibronectin, and Vitronectin Receptor cells plated on various extracellular matrix proteins as a first step toward identifying ligands for ␣8␤1. In cells plated on fibronectin or vitronectin, ␣8 co-localized to vinculin-containing focal contacts (Fig. 2, A–D).

Results

Conclusion