The Cleavage of the Urokinase Receptor Regulates Its Multiple Functions

Abstract

The urokinase-type plasminogen activator (uPA) is able to cleave its cell surface receptor (uPAR) anchored to the cell membrane through a glycophosphatidylinositol tail. The cleavage leads to the formation of cell surface truncated forms, devoid of the N-terminal domain 1 (D1) and unmasks or disrupts, depending on the cleavage site, a sequence in the D1-D2 linker region (residues 88 -92), which in the soluble form is a potent chemoattractant for monocyte-like cells. To investigate the possible role(s) of the cleaved forms of cell surface glycophosphatidylinositol-anchored uPAR, uPAR-negative human embrional kidney 293 cells were transfected with the cDNA of intact uPAR (uPAR-293) or with cDNAs corresponding to the truncated forms of uPAR exposing (D2D3-293) or lacking (D2D3wc-293) the peptide 88 -92 (P88 -92). Cell adhesion assays and co-immunoprecipitation experiments indicated that the removal of D1, independently of the presence of P88 -92, abolished the lateral interaction of uPAR with integrins and its capability to regulate integrin adhesive functions. The expression of intact uPAR induced also a moderate increase in 293 cell proliferation, which was accompanied by the activation of ERK. Also this effect was abolished by D1 removal, independently of the presence of P88 -92. The expression of intact and truncated uPARs regulated cell directional migration toward uPA, the specific uPAR ligand, and toward fMLP, a bacterial chemotactic peptide. In fact, the uPA-dependent cell migration required the expression of intact uPAR, including D1, whereas the fMLP-dependent cell migration required the expression of a P88 -92 containing uPAR and was independent of the presence of D1. Together these observations indicate that uPA-mediated uPAR cleavage and D1 removal, occurring on the cell surface of several cell types, can play a fundamental role in the regulation of multiple uPAR functions.

An urokinase receptor antagonist that inhibits cell migration by blocking the formyl peptide receptor

“…2A). A likely possibility is that pER-ERY-NH 2 recognizes FPR which is constitutively expressed by HEK-293 cells [14]. To study the role of FPR, we took advantage of RBL-2H3 cells which are devoid of FPR and RBL-2H3/ETFR which stably express FPR [16].…”

Gelatinase-mediated migration and invasion of cancer cells

“…Furthermore, uPAR can be cleaved by uPA (Hoyer-Hansen et al, 1992) or MMPs in vitro (Andolfo et al, 2002;Koolwijk et al, 2001) and the cleaved uPAR form is also found in invasive tumor xenografts (Solberg et al, 1994). The cleavage occurs between domains 1 and 2 impairs uPA and integrin binding (Montuori et al, 2002) and exposes a chemotactic epitope (Fazioli et al, 1997) suggesting that this cleavage is one of the cell migration regulating events on the cell surface.…”

“…Only a few other proteins have been shown to directly participate in cell detachment, namely tenascin-C, thrombospondin-1 and -2 and SPARC (secreted protein, acidic and rich in cysteine) (Murphy-Ullrich, 2001). The proteases can also indirectly modulate the affinity and hence the detachment of the cells by processing the extracellular matrix (Giannelli et al, 1997) or by cleaving integrin associated molecules (Andolfo et al, 2002;Montuori et al, 2002).…”

Urokinase Induces Basophil Chemotaxis through a Urokinase Receptor Epitope That Is an Endogenous Ligand for Formyl Peptide Receptor-Like 1 and -Like 2

“…The membrane was blocked with 5% nonfat dry milk and probed with 1 g/ml of an anti-uPAR polyclonal Ab. Finally, washed filters were incubated with HRP-conjugated anti-rabbit Ab and detected by ECL (10).…”

“…Therefore, uPAR can exist on the cell surface in either a three-domain form (D1D2D3), which is capable of binding uPA, or a two-domain form (D2D3), which does not bind uPA (5). Despite the lack of a transducing cytoplasmic tail, the uPA receptor is able to activate cell signaling pathways, probably by interacting with other cell surface proteins, such as integrins (7,8) and FMLP receptors (9,10), that interact with the cell interior (11).…”