α4β1Integrin-dependent Cell Adhesion Is Regulated by a Low Affinity Receptor Pool That Is Conformationally Responsive to Ligand

Ted A Yednock,Catherine Cannon,Christopher Vandevert,Erich G Goldbach,Gray Shaw,Debra K Ellis,Chen Liaw,Lawrence C Fritz,Laura I Tanner

doi:10.1074/jbc.270.48.28740

Ted A Yednock, Catherine Cannon + Show 7 more

Open Access

https://doi.org/10.1074/jbc.270.48.28740

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Abstract

alpha 4 beta 1 integrin (VLA-4) appears to be unique among the leukocyte integrins in that it can initiate the adhesion of circulating lymphocytes without cellular activation. It is not known how lymphocytes or other cell types maintain constitutive levels of alpha 4 beta 1 integrin activity. The current report describes a monoclonal antibody, 15/7, that recognizes a high affinity or ligand-occupied conformation of beta 1 integrin. Studies with 15/7 revealed that alpha 4 beta 1 integrin-dependent adhesion of leukocytic cell lines is mediated by a population of low affinity receptors that is conformationally responsive to ligand; the 15/7 epitope could be induced by nanomolar concentrations of soluble VCAM-1 or by micromolar concentrations of a peptide derived from the type III connecting segment domain of fibronectin (as ligands for alpha 4 beta 1 integrin). The same receptors were also responsive to adhesion activating reagents, such as Mn2+, activating anti-beta 1 integrin antibodies, and phorbol myristate acetate, which induced the 15/7 epitope directly and/or decreased the concentration of ligand required for epitope induction. In addition to the responsive receptor pool, cells expressed a second population of alpha 4 beta 1 integrin that was conformationally restrained, failing to respond to ligand or to any of the activating reagents. The relative size of the responsive and inactive receptor pools, as well as the affinity of the responsive receptors, represented a stable phenotype of different cell types and played important roles in defining the cells' adhesive capacity and ligand specificity. Similar receptor populations were measured on lymphocyte subsets in whole blood. These studies provide insight into how cells maintain different constitutive levels of alpha 4 beta 1 integrin activity, and how the activity of beta 1 integrin can be modulated by activators of cell adhesion.

Highlights

The regulation of ␣4 integrin activity on circulating immune cells appears to be different from that of the other leukocyte integrins described above. ␣4␤1 and ␣4␤7 integrin can mediate leukocyte adhesion to their endothelial ligands VCAM-1 [5] and MAdCAM-1 [6], respectively, without cellular activation, and can do so even in the presence of the shear forces encountered in normal blood flow [7,8,9,10]
These results suggest that circulating leukocytes maintain a constitutive level of ␣4 integrin activity. ␣4␤1 and ␣4␤7 integrin bind to the alternatively spliced connecting segment III (CS1)1 domain of fibronectin (FN; 11–13), but freshly isolated blood lymphocytes require activation in order to adhere to this ligand [7]
Cell lines maintained in culture exhibit a range of ␣4␤1 integrin activity; some cells bind to both VCAM-1 and the CS1 domain of fibronectin, others bind VCAM-1 but require activation to bind FN CS1, while others cannot bind FN CS1 even in the presence of activating reagents [14]

Summary

Introduction

The regulation of ␣4 integrin activity on circulating immune cells appears to be different from that of the other leukocyte integrins described above. ␣4␤1 and ␣4␤7 integrin can mediate leukocyte adhesion to their endothelial ligands VCAM-1 [5] and MAdCAM-1 [6], respectively, without cellular activation, and can do so even in the presence of the shear forces encountered in normal blood flow [7,8,9,10]. ␣4␤1 and ␣4␤7 integrin can mediate leukocyte adhesion to their endothelial ligands VCAM-1 [5] and MAdCAM-1 [6], respectively, without cellular activation, and can do so even in the presence of the shear forces encountered in normal blood flow [7,8,9,10]. These results suggest that circulating leukocytes maintain a constitutive level of ␣4 integrin activity. Another class of antibodies recognize ligand-induced binding sites associated with ␣IIb␤3 integrin [33], and document changes that occur in receptor conformation upon ligand occupation

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