Synaptopodin is upregulated by IL-13 in eosinophilic esophagitis and regulates esophageal epithelial cell motility and barrier integrity
Mark Rochman, Jared Travers, J. Pablo Abonia, Julie M. Caldwell, Marc E. Rothenberg
Mark Rochman, Jared Travers, J. Pablo Abonia, Julie M. Caldwell, Marc E. Rothenberg
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Research Article Gastroenterology Inflammation

Synaptopodin is upregulated by IL-13 in eosinophilic esophagitis and regulates esophageal epithelial cell motility and barrier integrity

Abstract

Eosinophilic esophagitis (EoE) is an allergic inflammatory disease of the esophagus mediated by an IL-13–driven epithelial cell transcriptional program. Herein, we show that the cytoskeletal protein synaptopodin (SYNPO), previously associated with podocytes, is constitutively expressed in esophageal epithelium and induced during allergic inflammation. In addition, we show that the SYNPO gene is transcriptionally and epigenetically regulated by IL-13 in esophageal epithelial cells. SYNPO was expressed in the basal layer of homeostatic esophageal epithelium, colocalized with actin filaments, and expanded into the suprabasal epithelium in EoE patients, where expression was elevated 25-fold compared with control individuals. The expression level of SYNPO in esophageal biopsies correlated with esophageal eosinophil density and was improved following anti–IL-13 treatment in EoE patients. In esophageal epithelial cells, SYNPO gene silencing reduced epithelial motility in a wound healing model, whereas SYNPO overexpression impaired epithelial barrier integrity and reduced esophageal differentiation. Taken together, we demonstrate that SYNPO is induced by IL-13 in vitro and in vivo, is a nonredundant regulator of epithelial cell barrier function and motility, and is likely involved in EoE pathogenesis.

Authors

Mark Rochman, Jared Travers, J. Pablo Abonia, Julie M. Caldwell, Marc E. Rothenberg

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Figure 2

SYNPO expression in esophageal epithelial cells.

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SYNPO expression in esophageal epithelial cells. (A) Schematic structur...
(A) Schematic structure of the SYNPO isoforms; black and white rectangles represent exons and introns, respectively. SYNPO-long and SYNPO-short represent the common protein isoforms 1 and 2, respectively. The SYNPO regions detected by RT-PCR are indicated by dotted lines; SYNPO 1–2 and SYNPO 3–4 represent common PCR products for indicated isoforms. (B) An image capture from the USCS browser shows aligned RNA-sequencing reads on the SYNPO gene in TE-7 cells either untreated or treated with IL-13 for 24 hours. Black rectangles represent exons. The indicated P value comparing expression under these conditions was calculated by DESeq analysis (59). Note that reads are detected only on the exons corresponding to SYNPO-long and SYNPO-short isoforms. (C) Expression of SYNPO isoforms by TE-7 cells induced by IL-13 was assessed by RT-PCR. *P < 0.05, t test. (D) Expression of SYNPO was assessed by RNA sequencing in TE-7 cells subjected to gene silencing control (scrambled) and STAT6 knockdown (STAT6KD) pool. SYNPO expression in RPKM is shown. ****P < 0.0001 for 24-hour time point, DESeq analysis. (E) An image capture from the USCS browser shows aligned RNA-sequencing reads on the SYNPO gene in control TE-7 cells and STAT6KD pool from D. Note that both isoforms are affected by the knockdown. The indicated P value comparing expression under these conditions was calculated by DESeq analysis (59). (F) Expression of SYNPO isoforms (mean ± SD) was assessed by RT-PCR. ****P < 0.001, Holm-Sidak method. (G) Expression of SYNPO was analyzed in primary esophageal epithelial cells stimulated with IL-13. Combined data from cells isolated from 3 independent biopsies are shown. *P < 0.05, t test. For C, F, and G, expression was normalized to expression of GAPDH. For box-and-whisker plots, the box represents 50th percentile of the data, whiskers show minimum and maximum values, and the line in the box represents the median. For B and E, the read depth is of the same scale within each panel.