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Background and aimsThe circadian clock orchestrates ~24-hour oscillations of gastrointestinal (GI) epithelial structure and function that drive diurnal rhythms in the composition, localization, and metabolism of gut microbiota. Here, we use experimental and computational approaches in enteroids to reveal reciprocal effects of microbial metabolites on intestinal epithelial timekeeping by an epigenetic mechanism.MethodsWe cultured 3D PER2∷LUCIFERASE andBmal1-ELuciferasejejunal enteroids in media supplemented with sterile supernatants from the altered Schaedler Flora (ASF), a defined murine microbiota. Circadian oscillations of bioluminescent PER2 andBmal1were measured in enteroids cultured in the presence or absence of individual ASF supernatants. Separately, we applied machine learning to ASF metabolic profiles to identify phase-shifting metabolites.ResultsFiltrates from 3 of 7 ASF species (ASF360Lactobacillus intestinalis, ASF361Ligilactobacillus murinus, ASF502Clostridiumspp.) induced minimal alterations in circadian rhythms, whereas 4 ASF species (ASF356Clostridiumspp., ASF492Eubacterium plexicaudatum, ASF500Pseudoflavonifactorspp., ASF519Parabacteroides goldsteinii) induced profound, concentration-dependent phase delays. Random forest classification identified short chain fatty acids (SCFA: butyrate, propionate, acetate, and isovalerate) production as a discriminating feature of “shifters”, i.e., ASF taxa whose metabolites induce phase delay. Experiments with SCFAs confirmed machine learning predictions, with a median phase delay of 6.2 hours. Pharmacological or botanical HDAC inhibitors generated similar phase delays. Further, mithramycin A, an inhibitor of HDAC inhibition, abrogated SCFA-induced phase delays by 20% (PBmal1-luciferaseenteroids.ConclusionsGut microbe-generated SCFAs entrain intestinal epithelial circadian rhythms, in part, by an HDACi-dependent mechanism, with critical implications for understanding microbial and circadian network regulation of intestinal epithelial homeostasis. |
