From prediction to experimental validation: desmoglein 2 is a functionally relevant substrate of matriptase in epithelial cells and their reciprocal relationship is important for cell adhesion

Abstract

Accurate identification of substrates of a protease is critical in defining its physiological functions. We previously predicted that Dsg-2 (desmoglein-2), a desmosomal protein, is a candidate substrate of the transmembrane serine protease matriptase. The present study is an experimental validation of this prediction. As demanded by our published method PNSAS [Prediction of Natural Substrates from Artificial Substrate of Proteases; Venkatraman, Balakrishnan, Rao, Hooda and Pol (2009) PLoS ONE 4, e5700], this enzyme-substrate pair shares a common subcellular distribution and the predicted cleavage site is accessible to the protease. Matriptase knock-down cells showed enhanced immunoreactive Dsg-2 at the cell surface and formed larger cell clusters. When matriptase was mobilized from intracellular storage deposits to the cell surface there was a decrease in the band intensity of Dsg-2 in the plasma membrane fractions with a concomitant accumulation of a cleaved product in the conditioned medium. The exogenous addition of pure active recombinant matriptase decreased the surface levels of immunoreactive Dsg-2, whereas the levels of CD44 and E-cadherin were unaltered. Dsg-2 with a mutation at the predicted cleavage site is resistant to cleavage by matriptase. Thus Dsg-2 seems to be a functionally relevant physiological substrate of matriptase. Since breakdown of cell-cell contact is the first major event in invasion, this reciprocal relationship is likely to have a profound role in cancers of epithelial origin. Our algorithm has the potential to become an integral tool for discovering new protease-substrate pairs.

MCRS1 overexpression, which is specifically inhibited by miR-129*, promotes the epithelial-mesenchymal transition and metastasis in non-small cell lung cancer

“…In our study, E-cadherin was found to be up-regulated at both the mRNA and protein levels after MCRS1 knockdown, suggesting that MCRS1 overexpression can lead to AJ disruption, cellular invasion, and tumor metastasis. 3) DSG2 is a transmembrane protein found in desmosomes and plays an important role in cell adhesion [ 28 ], the loss of which facilitates the EMT program and tumor metastasis [ 29 ]. We found that MCRS1 overexpression suppressed the expression of DSG2 mRNA, indicating that MCRS1 overexpression could impair cell adhesion and increase tumor invasion and metastasis.…”

Cleavage of transmembrane junction proteins and their role in regulating epithelial homeostasis

“…Additionally, shedding of desmosomal cadherin ectodomains by cellular proteases has been reported during cancer progression and metastasis implying a role of these molecules in modulating invasion and metastasis. For example, cleavage of Dsg2 in intestinal epithelial cells by matriptase has been associated with decreased cancer cell-cell adhesion 57 . ADAM17 and 10 have also been implicated in the shedding of Dsg2 during cancer development 58 .…”

Hepatocyte Growth Factor Activator Inhibitor Type 1 Maintains the Assembly of Keratin into Desmosomes in Keratinocytes by Regulating Protease-Activated Receptor 2–Dependent p38 Signaling

“…Previous studies demonstrated that serine proteases are implicated in the cleavage of desmogleins. For example, desmoglein 2 is a functional and physiological substrate of matriptase and KLK-7, 10 and KLK-5 promotes cleavage of desmoglein-1. 11 In the present study, we found that the desmoglein-3 protein level was reduced in HAI-1 KD HaCaT cells without significant alteration of its mRNA level, and treatment with the p38 inhibitor, SB203580, abrogated the reduction of desmoglein-3.…”

“…For example, matriptase, a type II transmembrane serine protease on the epithelial cell surface, degrades desmoglein-2, and kallikrein 1erelated peptidase (KLK)-5 promotes cleavage of desmoglein-1. 10,11 Hepatocyte growth factor activator inhibitor type 1 (HAI-1; official symbol SPINT1), encoded by the SPINT1 gene, is a serine protease inhibitor abundantly expressed in the placenta and in epithelial tissues. 12,13 HAI-1 regulates several trypsinlike serine proteinases, such as hepatocyte growth factor activator, matriptase, prostasin, hepsin, TMPRSS13, human airway trypsin-like protease, and KLKs 4 and 5.…”