Cellular glycosylphosphatidylinositol-specific phospholipase D regulates urokinase receptor shedding and cell surface expression

Abstract

The glycosylphosphatidylinositol (GPI)-anchored, multifunctional receptor for the serine proteinase, urokinase plasminogen activator (uPAR, CD87), regulates plasminogen activation and cell migration, adhesion, and proliferation. uPAR occurs in functionally distinct, membrane-anchored and soluble isoforms (s-uPAR) in vitro and in vivo. Recent evidence indicates that s-uPAR present in the circulation of cancer patients correlates with tumor malignancy and represents a valuable prognostic marker in certain types of cancer. We have therefore analyzed the mechanism of uPAR shedding in vitro. We present evidence that uPAR is actively released from ovarian cancer cells since the rate of receptor shedding did not correlate with uPAR expression. While s-uPAR was derived from the cell surface, it lacked the hydrophobic portion of the GPI moiety indicating anchor cleavage. We show that uPAR release is catalyzed by cellular GPI-specific phospholipase D (GPI-PLD), an enzyme cleaving the GPI anchor of the receptor. Thus, recombinant GPI-PLD expression increased receptor release up to fourfold. Conversely, a 40% reduction in GPI-PLD activity by GPI-PLD antisense mRNA expression inhibited uPAR release by more than 60%. We found that GPI-PLD also regulated uPAR expression, possibly by releasing a GPI-anchored growth factor. Our data suggest that cellular GPI-PLD might be involved in the generation of circulating prognostic markers in cancer and possibly regulate the function of GPI-anchored proteins by generating functionally distinct, soluble counterparts.

Soluble urokinase plasminogen activator receptor levels are associated with severity of fibrosis in nonalcoholic fatty liver disease

“…10 The soluble urokinase plasminogen activator receptor (suPAR) constitutes the circulating form of the glycosylphosphatidylinositol-linked membrane protein urokinase-type plasminogen activator receptor (uPAR; CD87), and is involved in inflammation, tissue remodeling and cancer metastasis. 11,12 uPAR is expressed by a wide range of immune cells whereas suPAR can be detected in different body fluids, including serum and plasma. [11][12][13][14][15][16][17] Cell-surface uPAR expression is up-regulated upon stimulation with growth factors and cytokines such as interleukin-1β and tumor necrosis factor.…”

“…11,12 uPAR is expressed by a wide range of immune cells whereas suPAR can be detected in different body fluids, including serum and plasma. [11][12][13][14][15][16][17] Cell-surface uPAR expression is up-regulated upon stimulation with growth factors and cytokines such as interleukin-1β and tumor necrosis factor. 18,19 The full length suPAR shed from the cell surface contains three domains (DI-III), and suPAR can occur in different cleaved forms consisting of only DI or DII-III, with partly divergent biological functions.…”

Soluble urokinase plasminogen activator receptor—A valuable biomarker in systemic lupus erythematosus?

“…Proteolytic cleavage sites are found between the DI and DII units of uPAR and at the GPI-anchor, rendering three possible soluble forms (full length DI-DIII, DIIDIII and DI, respectively) as well as two membrane-bound variants (full length and DIIDIII) (Figure 1). Enzymes responsible for cleavage of suPAR are uPA itself [40,41], matrix metalloproteinases (MMPs) 3, 12, 19 and 25 [41], GPI-specific phospholipase D [42], neutrophil elastase [43], plasmin [40,41,44] and cathepsin G [43], of which the two latter enzymes are capable of cleaving at both sites ( Figure 1). Factors that have been shown to induce suPAR shedding in an indirect way are cell-cell contact per se [45,46], presence of pro-inflammatory cytokines [25,47], bacterial lipopolysaccharide [46] and growth factors [47].…”

Identification of carcinoma embryonic antigen release mechanism Carcinoembryonic Antigen (CEA) from the surface of colorectal cancer cells

“…Studies have also shown that CEA is released in a nonproteolytic incision, and possibly by an endogenous phospholipase, in normal colon epithelial cells and in colon cancer cells [13]. GPI-PLD androgen responsible for cutting and releasing GPI proteins from their anchorages at the cell surface [14][15][16]. The cDNA has been cloned with a GPI (GPI-PLD) specific phospholipase [17] and expressed in a number of human tissues, including pancreatic beta cells, macrophages, keratinocytes, and liver cells [18].…”

“…The cDNA has been cloned with a GPI (GPI-PLD) specific phospholipase [17] and expressed in a number of human tissues, including pancreatic beta cells, macrophages, keratinocytes, and liver cells [18]. New studies suggest that GPI-PLD is responsible for cutting and releasing a number of GPI proteins from a variety of cells, such as tumor cells [16][17][18][19], and so we are investigating the role of this enzyme in releasing carcinoma-embryonic antigens from the surface Colorectal carcinoma cells.…”