Abstract
BackgroundMeprin (EC 3.4.24.18), an astacin-like metalloprotease, is expressed in the epithelium of the intestine and kidney tubules and has been related to cancer, but the mechanistic links are unknown.Methodology/Principal FindingsWe used MDCK and Caco-2 cells stably transfected with meprinα and or meprinβ to establish models of renal and intestinal epithelial cells expressing this protease at physiological levels. In both models E-cadherin was cleaved, producing a cell-associated 97-kDa E-cadherin fragment, which was enhanced upon activation of the meprin zymogen and reduced in the presence of a meprin inhibitor. The cleavage site was localized in the extracellular domain adjacent to the plasma membrane. In vitro assays with purified components showed that the 97-kDa fragment was specifically generated by meprinβ, but not by ADAM-10 or MMP-7. Concomitantly with E-cadherin cleavage and degradation of the E-cadherin cytoplasmic tail, the plaque proteins β-catenin and plakoglobin were processed by an intracellular protease, whereas α-catenin, which does not bind directly to E-cadherin, remained intact. Using confocal microscopy, we observed a partial colocalization of meprinβ and E-cadherin at lateral membranes of incompletely polarized cells at preconfluent or early confluent stages. Meprinβ-expressing cells displayed a reduced strength of cell-cell contacts and a significantly lower tendency to form multicellular aggregates.Conclusions/SignificanceBy identifying E-cadherin as a substrate for meprinβ in a cellular context, this study reveals a novel biological role of this protease in epithelial cells. Our results suggest a crucial role for meprinβ in the control of adhesiveness via cleavage of E-cadherin with potential implications in a wide range of biological processes including epithelial barrier function and cancer progression.
Highlights
Meprin is a zinc metalloendopeptidase of the astacin family, which was first discovered as an abundantly expressed protease at the brush border membrane of epithelial cells in the kidney [1] and the intestine [2]
Activation of meprin by in situ trypsin treatment of cell cultures increased the cleavage of E-cadherin and resulted in the accumulation of the 97-kDa fragment at the expense of the 120-kDa mature form (Figure 1A)
Western blot analysis of MDCK cells expressing individual subunits revealed that meprinb but not meprina mediated the cleavage of E-cadherin (Figure 1B)
Summary
Meprin is a zinc metalloendopeptidase of the astacin family, which was first discovered as an abundantly expressed protease at the brush border membrane of epithelial cells in the kidney [1] and the intestine [2]. The a and b subunits form homo- or heterodimers covalently linked via disulfide bonds, as well as higher multimeric structures resulting from non-covalent association of dimers [3]. Both subunits are synthesized as zymogens and targeted to the cell surface. The multimers composed of the b subunits are mainly transmembrane complexes, whereas those composed only of meprina are secreted, due to an intracellular processing event that removes the transmembrane anchor [4,5,6] Their activation requires removal of a propeptide by enzymes such as trypsin or plasmin [7,8]. Meprin (EC 3.4.24.18), an astacin-like metalloprotease, is expressed in the epithelium of the intestine and kidney tubules and has been related to cancer, but the mechanistic links are unknown
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