Epithelial-derived gasdermin D mediates nonlytic IL-1β release during experimental colitis
Katarzyna Bulek, … , Theresa T. Pizarro, Xiaoxia Li
Katarzyna Bulek, … , Theresa T. Pizarro, Xiaoxia Li
Published June 29, 2020
Citation Information: J Clin Invest. 2020;130(8):4218-4234. https://doi.org/10.1172/JCI138103.
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Research Article Gastroenterology Inflammation

Epithelial-derived gasdermin D mediates nonlytic IL-1β release during experimental colitis

Abstract

Gasdermin D (GSDMD) induces pyroptosis via the pore-forming activity of its N-terminal domain, cleaved by activated caspases associated with the release of IL-1β. Here, we report a nonpyroptotic role of full-length GSDMD in guiding the release of IL-1β–containing small extracellular vesicles (sEVs) from intestinal epithelial cells (IECs). In response to caspase-8 inflammasome activation, GSDMD, chaperoned by Cdc37/Hsp90, recruits the E3 ligase, NEDD4, to catalyze polyubiquitination of pro–IL-1β, serving as a signal for cargo loading into secretory vesicles. GSDMD and IL-1β colocalize with the exosome markers CD63 and ALIX intracellularly, and GSDMD and NEDD4 are required for release of CD63+ sEVs containing IL-1β, GSDMD, NEDD4, and caspase-8. Importantly, increased expression of epithelial-derived GSDMD is observed both in patients with inflammatory bowel disease (IBD) and those with experimental colitis. While GSDMD-dependent release of IL-1β–containing sEVs is detected in cultured colonic explants from colitic mice, GSDMD deficiency substantially attenuates disease severity, implicating GSDMD-mediated release of IL-1β sEVs in the pathogenesis of intestinal inflammation, such as that observed in IBD.

Authors

Katarzyna Bulek, Junjie Zhao, Yun Liao, Nitish Rana, Daniele Corridoni, Agne Antanaviciute, Xing Chen, Han Wang, Wen Qian, William A. Miller-Little, Shadi Swaidani, Fangqiang Tang, Belinda B. Willard, Keith McCrae, Zizhen Kang, George R. Dubyak, Fabio Cominelli, Alison Simmons, Theresa T. Pizarro, Xiaoxia Li

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Figure 1

GSDMD localizes to the colonic epithelium and its deficiency protects from DSS-induced colitis.

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GSDMD localizes to the colonic epithelium and its deficiency protects fr...
Colitis was induced in cohoused, sex-matched Gsdmd–/– (n = 10) and control Gdsmd+/– (n = 11) littermate mice by administration of 3% DSS in drinking water. Severity of colitis, assessed by (A) weight loss, (B) disease activity index (DAI), and (C) colon length (decreased length indicates increased inflammation). (D) Representative images of H&E-stained colons from DSS-treated mice (left), with histologic analysis of colitis performed on day 9 (right). Scale bars: 50 µM. (E) Representative images of immunofluorescence staining for CD4, F4/80, and LyG6 on frozen sections of inflamed colons taken at day 9 (left), with number of infiltrating immune cells enumerated and averaged over 10 HPF/section (right). Original magnification, ×40. (F) RT-PCR showing relative differences of indicated proinflammatory gene transcripts in colon tissues harvested on day 9 from experimental mice. Data are presented as fold induction over the mean of Gdsmd+/– group. (G) Representative IHC images showing immunolocalization of GSDMD to IECs in colons of DSS-challenged Gsdmd+/– control mice, with total absence in Gsdmd–/– littermates. Scale bars: 50 µM. Data are presented as mean ± SEM with *P < 0.05, ***P < 0.0001 by Student’s t test. All experiments were repeated twice and yielded consistent results.