Localization of urokinase to focal adhesions by human fibrosarcoma cells synthesizing recombinant vitronectin.
Cell surface plasminogen activators have been proposed to participate in cell migration and invasion by activating both intracellular signaling pathways and extracellular proteolysis. Urokinase-type plasminogen activator (uPA) is secreted from many cell types and localizes to focal contact areas when cells are seeded onto the plasma protein vitronectin. Induction of vitronectin synthesis during migration of neural crest cells and growth of certain tumors suggests that the de novo synthesis and deposition of vitronectin into the tissue matrix may remodel the matrix to provide an environment suitable for cell migration and (or) tumor invasion. To investigate the effects of vitronectin secretion and matrix deposition on the localization and activity of cell-associated uPA, HT-1080 fibrosarcoma cells were transfected with the Rc/CMV expression vector containing a vitronectin cDNA insert and stable cell lines expressing vitronectin were selected. Vitronectin-secreting cells were allowed to attach and spread on collagen- and fibronectin-coated substrates. Within 6 h, vitronectin was detected on the substrate; vitronectin synthesis was accompanied by the clustering of both the alpha v beta 5 vitronectin receptor and uPA into vinculin-containing focal adhesions. Although mock transfected cells formed small focal adhesions on both collagen and fibronectin, no co-localization of uPA or alpha v beta 5 to focal adhesions was evident in these cells. Vitronectin-secreting cells also exhibited decreased levels of plasminogen activation and increased levels of cell adhesion as compared with the mock transfected cells. These data demonstrate that the synthesis of vitronectin and its matrix association by transfected HT-1080 fibrosarcoma cells results in localization of uPA to alpha v beta 5 containing focal adhesions, decreased cell surface uPA activity, and an increase in cell adhesion.
Wilcox
S A
SA
Department of Physiology and Cell Biology, Albany Medical College of Union University, NY 12208, USA.
Reho
T
T
Higgins
P J
PJ
Tominna-Sebald
E
E
McKeown-Longo
P J
PJ
D002448:Cell Adhesion; D005109:Extracellular Matrix / Q000378:metabolism; D005341:Fibrinolysin / Q000378:metabolism; D005354:Fibrosarcoma / Q000378:metabolism*; D006801:Humans; D008856:Microscopy, Fluorescence; D011487:Protein Conformation; D011956:Receptors, Cell Surface / Q000032:analysis; D055293:Receptors, Urokinase Plasminogen Activator; D011994:Recombinant Proteins / Q000096:biosynthesis; D014162:Transfection; D014407:Tumor Cells, Cultured; D014568:Urokinase-Type Plasminogen Activator / Q000032:analysis*; D019096:Vitronectin / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism
C524898:PLAUR protein, human; D011956:Receptors, Cell Surface; D055293:Receptors, Urokinase Plasminogen Activator; D011994:Recombinant Proteins; D019096:Vitronectin; D005341:Fibrinolysin; D014568:Urokinase-Type Plasminogen Activator