Induction of cell migration by pro-urokinase binding to its receptor: possible mechanism for signal transduction in human epithelial cells
Détails
ID Serval
serval:BIB_C82408F10C09
Type
Article: article d'un périodique ou d'un magazine.
Collection
Publications
Institution
Titre
Induction of cell migration by pro-urokinase binding to its receptor: possible mechanism for signal transduction in human epithelial cells
Périodique
Journal of Cell Biology
ISSN
0021-9525 (Print)
Statut éditorial
Publié
Date de publication
07/1994
Volume
126
Numéro
1
Pages
259-70
Notes
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S. --- Old month value: Jul
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S. --- Old month value: Jul
Résumé
A human epithelial cell line, WISH, and a mouse cell line, LB6-uPAR, transfected with the human urokinase receptor (uPAR), both expressed high affinity uPAR but undetectable levels of urokinase (uPA). In two independent assays, binding of exogenous pro-uPA produced an up to threefold enhancement of migration. The migration was time and concentration dependent and did not involve extracellular proteolysis. This biologic response suggested that uPAR can trigger an intracellular signal. Since this receptor is a glycosyl-phosphatidylinositol-linked protein, we postulated that it must do so by interacting with other proteins, among which, by analogy to other systems, would be a kinase. To test this hypothesis, we carried out a solid phase capture of uPAR from WISH cell lysates using either antibodies against uPAR or pro-uPA adsorbed to plastic wells, followed by in vitro phosphorylation of the immobilized proteins. SDS-PAGE and autoradiography revealed two phosphorylated protein bands of 47 and 55 kD. Both proteins were phosphorylated on serine residues. Partial sequence of the two proteins showed a 100% homology to cytokeratin 18 (CK18) and 8 (CK8), respectively. A similar pattern of phosphorylation was obtained with lysates from A459 cells, a lung carcinoma, but not HL60, LB6-uPAR or HEp3 cell lysates, suggesting that the identified multiprotein uPAR-complex may be specific for simple epithelia. Moreover, immunocapture with antibody to another glycosyl-phosphatidylinositol-linked protein, CD55, which is highly expressed in WISH cells, was ineffective. The kinase was tentatively identified as protein kinase C, because it was inhibited by an analogue of staurosporine more specific for PKC and not by a PKA or tyrosine kinase inhibitors. The kinase was tentatively identified as PKC epsilon because of its resistance to PMA down-modulation, independence of Ca2+ for activity, and reaction with a specific anti-PKC epsilon antibody in Western blots. Cell fractionation into cytosolic and particulate fractions revealed that all four proteins, the kinase, uPAR, CK18, and CK8, were present in the particulate fraction. In vivo, CK8, and to a lesser degree CK18, were found to be phosphorylated on serine residues. Occupation of uPAR elicited a time-dependent increase in the phosphorylation intensity of CK8, a cell shape change and a redistribution of the cytokeratin filaments. These results strongly suggest that uPAR serves not only as an anchor for uPA but participates in a signal transduction pathway resulting in a pronounced biological response.
Mots-clé
Amino Acid Sequence
Cell Line
Cell Movement/*physiology
Cell Size/physiology
Enzyme Precursors/metabolism
Epithelial Cells
Epithelium/enzymology
Humans
Keratins/metabolism
Macromolecular Substances
Molecular Sequence Data
Multiprotein Complexes
Phosphorylation
Plasminogen Activators/*metabolism
Protein-Tyrosine Kinases/metabolism
Receptors, Cell Surface/*metabolism
Recombinant Proteins/metabolism
Signal Transduction/*physiology
Urinary Plasminogen Activator/*metabolism
Pubmed
Web of science
Création de la notice
25/01/2008 9:29
Dernière modification de la notice
20/08/2019 16:43