Proteomic Identification of Desmoglein 2 and Activated Leukocyte Cell Adhesion Molecule as Substrates of ADAM17 and ADAM10 by Difference Gel Electrophoresis

Abstract

In contrast with the early view of metalloproteases as simple extracellular matrix-degrading entities, recent findings show that they are highly specific modulators of different signaling pathways involved, positively or negatively, in tumor development. Thus, before considering a given metalloprotease a therapeutic target, it seems advisable to characterize its function by identifying its repertoire of substrates. Here, we present a proteomic approach to identify ADAM17 substrates by difference gel electrophoresis. We found that the shedding of the extracellular domain of the transferrin receptor and those of two cell-cell adhesion molecules, activated leukocyte cell adhesion molecule (ALCAM) and desmoglein 2 (Dsg-2), is increased in cells overexpressing ADAM17. Genetic evidence shows that while ADAM17 is responsible for the shedding of ALCAM, both ADAM17 and ADAM10 can act on Dsg-2. Activation of the epidermal growth factor receptor leads to the upregulation of the shedding of Dsg-2 and to the concomitant upregulation of ADAM17, but not ADAM10, supporting the ability of overexpressed ADAM17 to shed Dsg-2. These results unveil a role of ADAM10 and ADAM17 in the shedding of cell-cell adhesion molecules. Since loss of cell adhesion is an early event in tumor development, these results suggest that ADAM17 is a useful target in anticancer therapy.Mammalian extracellular and membrane-anchored metalloproteases are classified in large families; together, the family of the matrix metalloproteases (MMPs) and the related families of ADAMs (a disintegrin and metalloprotease) and ADAM-TS (ADAMs containing thrombospondin motifs) comprise nearly 75 members (21). Metalloproteases have long been considered targets of antitumor therapy because of their frequent upregulation in cancer and their ability to degrade the extracellular matrix, a hypothetical prerequisite for malignant cells to invade surrounding tissue (25). Following this reasoning, several wide-spectrum metalloprotease inhibitors (MPIs) have been clinically tested; unfortunately, no obvious benefit has been observed in treated patients (12).In contrast with the classical view, accumulating evidence shows that during tumor development, metalloproteases play highly specific roles, not directly related to extracellular matrix degradation. For example, MMP1 seems to promote metastasis through the proteolytic activation of protease-activated receptor 1 (7), a G protein-coupled receptor known to convey signals leading to tumor invasion in a variety of experimental models (11). Further, the proteolytic activity of ADAM17 is required to release ligands that activate the epidermal growth factor receptor (EGFR) (14), a tyrosine kinase receptor which contributes to tumor development by regulating cell proliferation, apoptosis, and migration (32). In contrast to those of MMP1 and ADAM17, the activity of MMP8 seems to play an antitumorigenic role: its absence causes increased tumor susceptibility, probably because of its ability to cleave cytokines that mediate the inflammatory...

EGFR and ADAMs Cooperate to Regulate Shedding and Endocytic Trafficking of the Desmosomal Cadherin Desmoglein 2

“…ADAM family members have been shown to cleave classic cadherins, and Dsg2 was also recently identified as a substrate of ADAM10 and 17 (Bech-Serra et al, 2006). To test whether either of these ADAM family members is responsible for Dsg2 cleavage in SCC68 cells, we introduced ADAM10 or 17 small interfering RNA (siRNA) into SCC68 cells.…”

Increased expression of ALCAM/CD166 in pancreatic cancer is an independent prognostic marker for poor survival and early tumour relapse

“…Recently, Bech-Serra et al (2006) have shown that ADAM17 acts as a proteolytic sheddase of ALCAM. Rosso et al (2007) observed that this process is involved in the motility of epithelial ovarian carcinoma cells.…”

Insights into Desmosome Biology from Inherited Human Skin Disease and Cardiocutaneous Syndromes

“…Substrates include a number of ligands for the EGF receptor (EGFR) and TNF-α , giving ADAM17 its alternative name TNF-α -converting enzyme. ADAM17 has additionally been shown to play a key role in ligandindependent Notch signalling, and in the shedding of cell adhesion molecules including DSG2 (Gschwind et al, 2003;Klessner et al, 2009;Pruessmeyer & Ludwig, 2009;Sahin et al, 2004;Bech-Serra et al, 2006).…”