Impact of Tfrc deficiency on gut microbiome [16s rRNA-seq]

Iron metabolism is pivotal for cell fitness in the mammalian host. However, its role in group 3 innate lymphoid cells (ILC3s) is unknown. Here we show that transferrin receptor CD71-mediated iron metabolism cell-intrinsically controls ILC3 maintenance, cytokine production and host protection against Citrobacter rodentium infection, and metabolically affects mitochondrial respiration by switching of oxidative phosphorylation toward glycolysis. Iron deprivation or Tfrc deficiency reduces the expression and/or activity of the aryl hydrocarbon receptor (Ahr), a ligand-dependent transcription factor and a key ILC3 regulator. Furthermore, consistent with its role in generation of Ahr ligand, microbiome exerts a negative impact on CD71 expression in an Ahr-dependent manner. Genetic ablation or activation of Ahr in ILC3s leads to CD71 upregulation or downregulation, respectively, suggesting an active suppression of CD71 by Ahr. Iron overload partially restores defective ILC3 compartment in the small intestine of Ahr-deficient mice, representing compensatory action of CD71 upregulation during Ahr deficiency. Mechanistically, Ahr directly binds to the promoter region of the Tfrc locus to inhibit Tfrc transcription. These data collectively demonstrate an under-appreciated role of the Ahr-CD71-iron axis in regulation of ILC3 maintenance and function. Overall design: For microbiome analyses, fecal DNA was isolated the Quick-DNA Fecal/Soil Microbe Miniprep Kit (ZYMO, 11-322), followed by amplification and library generation with the Quick-16S NGS Library Prep Kit (ZYMO, D6410) according to the manufacturer's instructions. Sequencing was performed on the Illumina MiSeq (Illumina, Inc., San Diego, CA). Raw FASTQ sequences were filtered and analyzed using QIIME2 (v2022.2.0) and Dada2. Operational taxonomic units were defined based on sequence similarity threshold of 97%. Taxonomy was assigned based on the Silva 138 reference database.