Sprouty2 inhibits expression of the host defense peptide relmβ in the colonic epithelium

Abstract Background Sprouty2 (Spry2) is an inflammation-responsive regulator of intracellular signaling that is suppressed in colitis. Loss of Spry2 levels leads to increased colonic tuft and goblet cell numbers; cell types protective against injury and infection. In particular, goblet cells produce host defense peptides (HDPs) which can enhance barrier effectiveness and limit pathogen colonization. While the activity of these cells is regulated and responsive to environmental cues, the precise mechanisms driving functional changes in goblet cells are largely unknown. Here, we tested the role of Spry2 in mediating colonic goblet cell function and HDP production. Hypothesis Spry2 regulates HDP production in colonic goblet cells and bacterial levels. Methods Distal colons from mice with an intestinal epithelial-specific Spry2 deletion (Spry2flox/flox;Villin-Cre, Spry2KOIE) and littermate controls were collected at 8 w of age, and analyzed by bulk and single-cell RNA sequencing. Fecal pellets from the distal colon were 16S sequenced to assess changes in microbial populations. Changes in HDP expression were assessed at the bulk tissue level and within individual cellular populations. Results Expression of the host defense peptide RELMβ, but not RELMα, RELMγ, or Resistin, was increased 15-fold (p<0.05) in Spry2KOIE mice in bulk RNA sequencing analysis. Single cell RNA sequencing showed RELMβ was highly enriched in Reg4+/Spink4+ putative crypt base goblet cells of both control and Spry2KOIE mice. Quantification of these cells by single-cell RNA sequencing revealed no significant difference in Reg4+/Spink4+ cell numbers in Spry2KOIE mice versus wild-type, suggesting increased RELMβ is due to goblet cell activation and not goblet cell population shifts. Taxonomic analysis of feces from 2 litters of mice showed a decreased abundance of Eggerthella spp. (reduced 61%, p<0.05) and Streptococcus spp. (reduced 50%, p<0.05), genera that have been identified in patients with inflammatory bowel disease. Conclusion Loss of epithelial Spry2 drives HDP production in the colon and alterations in the colonic microbiota. Impairment of this mechanism may impact colitis severity.