The contributions of integrin affinity and integrin-cytoskeletal engagement in endothelial and smooth muscle cell adhesion to vitronectin.
The serine proteinase inhibitor, plasminogen activator inhibitor type-1 (PAI-1), binds to the adhesion protein vitronectin with high affinity at a site that is located directly adjacent to the vitronectin RGD integrin binding sequence. The binding of PAI-1 to vitronectin sterically blocks integrin access to this site and completely inhibits the binding of purified integrins to vitronectin; however, its inhibition of endothelial and smooth muscle cell adhesion to vitronectin is at most 50-75%. Because PAI-1 binds vitronectin with approximately 10-100-fold higher affinity than purified integrins, we have analyzed the mechanism whereby these cells are able to overcome this obstacle. Our studies exclude proteolytic removal of PAI-1 from vitronectin as the mechanism, and show instead that cell adhesion in the presence of PAI-1 is dependent on integrin-cytoskeleton engagement. Disrupting endothelial or smooth muscle cell actin polymerization and/or focal adhesion assembly reduces cell adhesion to vitronectin in the presence of PAI-1 to levels similar to that observed for the binding of purified integrins to vitronectin. Furthermore, endothelial cell, but not smooth muscle cell adhesion to vitronectin in the presence of PAI-1 requires both polymerized microtubules and actin, further demonstrating the importance of the cytoskeleton for integrin-mediated adhesion. Finally, we show that cell adhesion in the presence of PAI-1 leads to colocalization of PAI-1 with the integrins alphavbeta3 and alphavbeta5 at the cell-matrix interface.
Stefansson
Steingrimur
S
Department of Physiology and Center for Vascular and Inflammatory Diseases, University of Maryland School of Medicine, Baltimore, Maryland 21201, USA. stenni@creatvmicrotech.com
Su
Enming J
EJ
Ishigami
Shoji
S
Cale
Jacqueline M
JM
Gao
Yamei
Y
Gorlatova
Natalia
N
Lawrence
Daniel A
DA
D000199:Actins / Q000378:metabolism; D000818:Animals; D002448:Cell Adhesion / Q000502:physiology; D003599:Cytoskeleton / Q000378:metabolism*; D042783:Endothelial Cells / Q000166:cytology / Q000378:metabolism*; D006801:Humans; D039302:Integrin alphaVbeta3 / Q000378:metabolism*; D016023:Integrins / Q000378:metabolism*; D032389:Myocytes, Smooth Muscle / Q000166:cytology / Q000378:metabolism*; D017395:Plasminogen Activator Inhibitor 1 / Q000378:metabolism; D011485:Protein Binding; D019030:Receptors, Vitronectin / Q000378:metabolism*; D019096:Vitronectin / Q000378:metabolism*
D000199:Actins; D039302:Integrin alphaVbeta3; D016023:Integrins; D017395:Plasminogen Activator Inhibitor 1; D019030:Receptors, Vitronectin; D019096:Vitronectin; C064660:integrin alphaVbeta5