The receptor for urokinase-type plasminogen activator regulates fibronectin matrix assembly in human skin fibroblasts.
Previous studies have indicated that the receptor for urokinase-type plasminogen activator, uPAR, can form functional complexes with integrin receptors thereby modulating integrin activity. In the present study, the role of uPAR in the regulation of alpha5beta1-dependent polymerization of the fibronectin matrix was investigated. Incubation of fibroblast monolayers with the P-25 peptide, a uPAR ligand, resulted in a 12-15-fold increase in the accumulation of exogenous fibronectin in the cell layer. The exogenous fibronectin co-localized in the extracellular matrix with endogenous cell-derived fibronectin, and its deposition into the matrix was inhibited by blocking antibodies against the beta1 integrin receptor. The P-25-dependent increase in fibronectin assembly was associated with a 7-8-fold increase in the expression of matrix assembly sites as well as a 37-fold increase in the rate of transfer of cell surface-bound fibronectin into a detergent-insoluble matrix. The effects of P-25 on the matrix assembly were attenuated by incubating cells with either phospholipase C or with antibodies against uPAR, confirming a role for uPAR in the P-25-dependent increase in matrix assembly. P-25-treated cells exhibited a 10-fold increase in the binding of the 120-kDa cell-binding fragment of fibronectin suggesting an increase in alpha5beta1 affinity for fibronectin. Consistent with this, treatment of cells with P-25 also resulted in a 6-10-fold increase in the binding of two different monoclonal antibodies that recognize the active conformation of the beta1 integrin. These results indicate that P-25 increases matrix assembly by altering the activation state of the alpha5beta1 integrin receptor and suggest that changes in integrin activation affect both the number of matrix assembly sites as well as the rate of transfer of cell-bound fibronectin into a detergent-insoluble matrix. These data provide direct evidence that uPAR and integrin receptors synergistically regulate the levels of fibronectin in the extracellular matrix.
Monaghan
Elizabeth
E
Center for Cell Biology and Cancer Research, Albany Medical College, Albany, New York 12208, USA.
Gueorguiev
Volodia
V
Wilkins-Port
Cynthia
C
McKeown-Longo
Paula J
PJ
D002448:Cell Adhesion; D002460:Cell Line; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D005109:Extracellular Matrix / Q000378:metabolism; D005347:Fibroblasts / Q000378:metabolism*; D005353:Fibronectins / Q000378:metabolism*; D006801:Humans; D039081:Integrin alpha5beta1 / Q000378:metabolism; D016023:Integrins / Q000378:metabolism; D008024:Ligands; D010455:Peptides / Q000737:chemistry; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D011956:Receptors, Cell Surface / Q000378:metabolism / Q000502:physiology*; D017469:Receptors, Fibronectin / Q000378:metabolism; D055293:Receptors, Urokinase Plasminogen Activator; D012867:Skin / Q000378:metabolism*; D013997:Time Factors; D014568:Urokinase-Type Plasminogen Activator / Q000378:metabolism
D005353:Fibronectins; D039081:Integrin alpha5beta1; D016023:Integrins; D008024:Ligands; C524898:PLAUR protein, human; D010455:Peptides; D011956:Receptors, Cell Surface; D017469:Receptors, Fibronectin; D055293:Receptors, Urokinase Plasminogen Activator; D014568:Urokinase-Type Plasminogen Activator