Critical role of integrin alpha 5 beta 1 in urokinase (uPA)/urokinase receptor (uPAR, CD87) signaling.
Urokinase-type plasminogen activator (uPA) induces cell adhesion and chemotactic movement. uPA signaling requires its binding to uPA receptor (uPAR/CD87), but how glycosylphosphatidylinositol-anchored uPAR mediates signaling is unclear. uPAR is a ligand for several integrins (e.g. alpha 5 beta 1) and supports cell-cell interaction by binding to integrins on apposing cells (in trans). We studied whether binding of uPAR to alpha 5 beta 1 in cis is involved in adhesion and migration of Chinese hamster ovary cells in response to immobilized uPA. This process was temperature-sensitive and required mitogen-activated protein kinase activation. Anti-uPAR antibody or depletion of uPAR blocked, whereas overexpression of uPAR enhanced, cell adhesion to uPA. Adhesion to uPA was also blocked by deletion of the growth factor domain (GFD) of uPA and by anti-GFD antibody, whereas neither the isolated uPA kringle nor serine protease domain supported adhesion directly. Interestingly, anti-alpha 5 antibody, RGD peptide, and function-blocking mutations in alpha 5 beta 1 blocked adhesion to uPA. uPA-induced cell migration also required GFD, uPAR, and alpha 5 beta 1, but alpha 5 beta 1 alone did not support uPA-induced adhesion and migration. Thus, binding of uPA causes uPAR to act as a ligand for alpha 5 beta 1 to induce cell adhesion, intracellular signaling, and cell migration. We demonstrated that uPA induced RGD-dependent binding of uPAR to alpha 5 beta 1 in solution. These results suggest that uPA-induced adhesion and migration of Chinese hamster ovary cells occurs as a consequence of (a) uPA binding to uPAR through GFD, (b) the subsequent binding of a uPA.uPAR complex to alpha 5 beta 1 via uPAR, and (c) signal transduction through alpha 5 beta 1.
Tarui
Takehiko
T
Department of Cell Biology, The Scripps Research Institute, La Jolla, California 92037, USA.
Andronicos
Nicholas
N
Czekay
Ralf-Peter
RP
Mazar
Andrew P
AP
Bdeir
Khalil
K
Parry
Graham C
GC
Kuo
Alice
A
Loskutoff
David J
DJ
Cines
Douglas B
DB
Takada
Yoshikazu
Y
D000818:Animals; D000911:Antibodies, Monoclonal / Q000378:metabolism; D016466:CHO Cells; D002448:Cell Adhesion; D002465:Cell Movement; D006224:Cricetinae; D018076:DNA, Complementary / Q000378:metabolism; D019281:Dimerization; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005353:Fibronectins / Q000378:metabolism; D017261:Glycosylphosphatidylinositols / Q000378:metabolism; D039081:Integrin alpha5beta1 / Q000502:physiology*; D008024:Ligands; D020935:MAP Kinase Signaling System; D008954:Models, Biological; D009154:Mutation; D009842:Oligopeptides / Q000494:pharmacology; D011233:Precipitin Tests; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011956:Receptors, Cell Surface / Q000378:metabolism*; D055293:Receptors, Urokinase Plasminogen Activator; D015398:Signal Transduction; D013696:Temperature; D014568:Urokinase-Type Plasminogen Activator / Q000378:metabolism*
D000911:Antibodies, Monoclonal; D018076:DNA, Complementary; D004791:Enzyme Inhibitors; D005353:Fibronectins; D017261:Glycosylphosphatidylinositols; D039081:Integrin alpha5beta1; D008024:Ligands; D009842:Oligopeptides; D011956:Receptors, Cell Surface; D055293:Receptors, Urokinase Plasminogen Activator; C047981:arginyl-glycyl-aspartic acid; D014568:Urokinase-Type Plasminogen Activator