Live bacterial ligands universally regulate mTOR activity but are not essential for homeostatic innate immunity

Abstract

Abstract Multiple studies have reported significant impact of gut microbiota on hematopoiesis, including granulocyte development and functional defects. Germ-Free (GF) mice show developmental defects due to lack of live bacteria; underscored by the development of Altered Schaedler Flora (ASF), a community of 8 bacterial species sufficient to rescue developmental immune and gastrointestinal defects. Mimicking gnotobiosis by using antibiotics perturbs the microbiome systemically. Therefore, results from such studies may represent sequelae of the drug perturbation rather than simply modifying microbiota. We studied the impact of gut microbiota signals on murine hematopoiesis without antibiotics or fecal transfer. Namely, by using mice (from Taconic) generated by embryonic transfer of C57/Bl6 fertilized eggs into females with controlled microbiota, totaling 4 groups of mice: 1. GF; 2. ASF (no Gammaproteobacteria); 3. Excluded Flora (EF, includes Gammaproteobacteria); 4. WildR 7(wild mouse microbiome). We found no significant differences in either number or effector function of bone-marrow neutrophil granulocytes, myeloid progenitors, or stem cells. The single-cell genomic landscape of CD117+ bone-marrow progenitors confirmed flow analyses but revealed a profound defect in gnotobiotic hematopoietic signatures. Importantly, standard pathogen free C57/BL6 mice were inappropriate controls. While gnotobiotic mice likely receive inert TLR stimulation via bedding and food, the addition of any group of live bacteria was sufficient to rescue gene expression downstream of mTOR signaling. Such robust signatures could be considered so essential to life that they do not depend on any single bacterial clade. This project is supported by NIH R01HL122661.