Microbes and microbial products that induce ERN2 expression in colonic goblet cells
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Citation
Abstract
OBJECTIVE: The colonic goblet cell is responsible for the production of MUC2, the primary glycoprotein of the mucus layer in the gastrointestinal tract. The ERN2 gene, which is enriched in goblet cells, is responsible for goblet cell differentiation and maintenance of the mucus barrier. The metabolites produced by the microbial community in the colon affect ERN2 induction, which shapes MUC2 secretion, an energy source of the microbes, creating a positive feedback loop. The objective of this study was to determine what microbes and microbial products associated with a normal mucus barrier can induce ERN2 expression.
METHODS: Relative mRNA expression of ERN2 and goblet cell signature genes were measured by qPCR in LS174T cells treated with butyrate for 24 hours. ERN2 promoter activity was assayed using a luciferase reporter plasmid under control of the human ERN2 promoter in HEK293T cells treated with a media control, sodium butyrate, bacteria culture conditioned media, a PPARƴ inhibitor, trichtostatin A, or mouse stool homogenates from ERN2 wild type and ERN2 knockout mice for 4 hours.
RESULTS: The relative mRNA expression of the goblet cell signature genes ERN2, MUC2, and SPDEF increased in LS174T cells when treated with 1mM sodium butyrate. Induction of the ERN2 promoter plasmid activity increased in HEK293T cells after treatment with 1mM sodium butyrate, 1:10 dilutions of stationary phase Clostridium sardiniense culture conditioned media, and 1:10 dilutions of stool homogenates from wild type mice. HEK293T cells treated with stool homogenates from ERN2 knockout mice did not significantly induce ERN2 promoter activity compared to cells treated with media control. Addition of a PPARƴ inhibitor to media control and butyrate treated cells did not reduce ERN2 activity in either treatment group in transfection experiments. However, treatment with the histone deacetylase inhibitor trichostatin A induced ERN2 promoter activity to a similar extent as butyrate treatment.
CONCLUSIONS: Butyrate had the greatest effect on ERN2 mRNA expression and ERN2 promoter activity in both qPCR and transfection experiments. The low molecular weight fraction of stationary phase Clostridium sardiniense culture conditioned media induced ERN2 promoter activity significantly more than media control treated cells. Clostridium sardiniense is a butyrate-producing bacteria of the Firmicutes phylum. Firmicutes bacteria are depleted in the microbiomes of ERN2 knockout mice. The HDAC inhibitor, trichtostatin A, induced ERN2 promoter activity to the same extent as butyrate, whereas the PPARƴ did not, leading us to believe butyrate is acting on the ERN2 promoter as an HDAC inhibitor.
Description
2025