Eosinophilic esophagitis–linked calpain 14 is an IL-13–induced protease that mediates esophageal epithelial barrier impairment
Benjamin P. Davis ... Leah C. Kottyan, Marc E. Rothenberg
Benjamin P. Davis ... Leah C. Kottyan, Marc E. Rothenberg
Published April 7, 2016
Citation Information: JCI Insight. 2016;1(4):e86355. doi:10.1172/jci.insight.86355.
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Categories: Research Article Immunology

Eosinophilic esophagitis–linked calpain 14 is an IL-13–induced protease that mediates esophageal epithelial barrier impairment

Abstract

We recently identified a genome-wide genetic association of eosinophilic esophagitis (EoE) at 2p23 spanning the calpain 14 (CAPN14) gene, yet the causal mechanism has not been elucidated. We now show that recombinant CAPN14 cleaves a calpain-specific substrate and is inhibited by 4 classical calpain inhibitors: MDL-28170, acetyl-calpastatin, E-64, and PD151746. CAPN14 is specifically induced (>100-fold) in esophageal epithelium after IL-13 treatment. Epithelial cells overexpressing CAPN14 display impaired epithelial architecture, characterized by acantholysis, epidermal clefting, and epidermolysis. CAPN14 overexpression impairs epithelial barrier function, as demonstrated by decreased transepithelial resistance (2.1-fold) and increased FITC-dextran flux (2.6-fold). Epithelium with gene-silenced CAPN14 demonstrates increased dilated intercellular spaces (5.5-fold) and less organized basal cell layering (1.5-fold) following IL-13 treatment. Finally, CAPN14 overexpression results in loss of desmoglein 1 (DSG1) expression, whereas the IL-13–induced loss of DSG1 is normalized by CAPN14 gene silencing. Importantly, these findings were specific to CAPN14, as they were not observed with modulation of CAPN1 expression. These results, along with the potent induction of CAPN14 by IL-13 and genetic linkage of EoE to the CAPN14 gene locus, demonstrate a molecular and cellular pathway that contributes to T helper type 2 responses in mucosal epithelium.

Authors

Benjamin P. Davis, Emily M. Stucke, M. Eyad Khorki, Vladislav A. Litosh, Jeffrey K. Rymer, Mark Rochman, Jared Travers, Leah C. Kottyan, Marc E. Rothenberg

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Figure 1

Recombinant calpain 14 has protease activity.

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Recombinant calpain 14 has protease activity. Data from calpain activity...
Data from calpain activity assays comparing enzymatic activity of recombinant calpain 1 (rCAPN1), recombinant calpain 14 (rCAPN14), and rCAPN14 protease core are shown. (A) Calpain activity of rCAPN14 and rCAPN14 protease core as a function of protein amount (0.25–250 nM). (B) Calpain activity of rCAPN1 and rCAPN14 in the presence and absence of known reversible calpain inhibitors (n = 3). (C) Calpain activity of rCAPN1 and rCAPN14 in the presence and absence of known irreversible calpain inhibitor E-64. Data are representative of 3 independent experiments. Data are expressed as the mean ± SEM; ****P < 0.0001; statistical significance determined using a 2-tailed t test. (D) Reaction progress curves for cleavage of Suc-LLVY-AMC by rCAPN1 and rCAPN14 in the absence of inhibitors.