Gene Expression Analysis of the Effect of Microbial Tryptophan Metabolites on T‐cell Differentiation
Authors: Cheng Clint , Steinmeyer Shelby , Jayaraman Arul , Alaniz Robert 
Published: June 2021

The goal of this work is to elucidate the molecular basis underlying the effect of tryptophan‐derived microbiota metabolites on T‐cell differentiation and function. Naive T‐cells differentiate into various subsets with specialized functions, including pro‐tolerogenic regulatory T (Treg) cells and pro‐inflammatory T helper 17 (Th17) cells. An imbalance in the relative proportions of the different T‐cell subtypes is associated with a number of autoimmune and inflammatory diseases. While the role of host cytokines and growth factors in T‐cell differentiation are well established, recent studies have suggested that the gut microbiota metabolites also modulate T‐cell differentiation. Recent work from our laboratory has demonstrated that the microbiota metabolite indole, which has been previously shown to attenuate indicators of inflammation in intestinal epithelial cells, augments TGF‐β mediated Treg differentiation from naïve T‐cells as well as suppresses IL‐6 mediated Th17 differentiation, promoting an anti‐inflammatory phenotype. On the other hand, 5‐hydroxyindole (5HI) shows reciprocal effects, suppressing Treg differentiation and augmenting Th17 differentiation. However, the mechanisms by which indole and 5HI modulate T‐cell differentiation and their effects on function are unknown. Using microarrays, we identified 551 and 219 differentially expressed genes in indole treated Treg cells and 5‐HI treated Th17 cells, respectively, as compared to conventionally differentiated cells. Indole treatment resulted in a slight decrease in overall transcription, with 290 of the differentially expressed genes being upregulated, while 5HI increased overall transcription with 182 differentially expressed genes being upregulated. The differentially expressed genes in Treg cells with indole treatment was largely comprised of enzymes (14%), transcription factors (10%), and cytokines (7%), while 5HI resulted in differential regulation of enzymes (10%), cytokines (9%), and transmembrane receptors (5%) in Th17 cells. The glycine amidinotransferase (Gatm) gene that is involved in creatine biosynthesis and contains an Aryl Hydrocarbon Responsive Element II (AHRE‐II) in its 5’ regulatory region was up‐regulated 12‐fold in indole treated Treg cells, suggesting a role for the aryl hydrocarbon receptor (AhR) in indole signaling. The gene encoding the IL‐13 cytokine was the most strongly induced by indole treatment (82‐fold increase). IL‐13 is produced by Th2 cells and has both pro and anti‐inflammatory properties. Interestingly, other Th2 cytokines genes, Il4 and Il5, were also up‐regulated at 10‐ and 26‐fold, respectively. This unexpected pattern of signaling suggests that indole may induce the differentiation of a unique cell type with phenotypes characteristic of both Treg cells and Th2 cells. In 5HI treated Th17 cells, a 15‐ and 3‐fold increase in the expression of genes encoding the pro‐inflammatory cytokines IL‐22 and IFNg was observed. The gene for IL‐17, which is the characteristic cytokine of Th17 cells was upregulated 1.6 fold. These expression patterns support the induction of pro‐inflammatory signaling by 5HI. Overall, these results suggest that different tryptophan‐derived microbiota metabolites can have different effects on T‐cell differentiation and function. Further characterization of cell function and investigation into the role of the AhR the signaling of these metabolites is warranted.Support or Funding InformationNational Institutes of Health R01AI110642This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.