Identification and biological effects of microbiome-derived sulfated metabolites - Project Summary/Abstract Recent studies have shown that metabolites produced by human gut bacteria causally influence health and disease. At the same time, the development of modern metabolomic techniques has enabled the investigation of underexplored classes of metabolites. Here, we focus on one such class of underappreciated microbiome- dependent molecules, the sulfated metabolome. While sulfated metabolites were long considered waste products, new studies have revealed that they can have a variety of important physiological roles, including effects on immune response, host metabolism, and neurological function. In addition, recent studies have revealed that these compounds are not only host-produced – commensal bacteria have been identified that either chemically modify or synthesize a variety of sulfated compounds. Indeed, work from our groups showed that abundant human gut bacteria sulfonate steroidal compounds found in the gut lumen, including cholesterol. We identified the bacterial gene responsible for this activity, and we showed that the presence of this gene inhibited the migration of T cells in vivo. These results suggest that microbiome-dependent sulfated metabolites affect immune function. In this research, we propose to use analytical chemistry pipelines that we have developed to identify microbiome- dependent sulfated metabolites, and we propose to use in vitro and in vivo studies to elucidate the immunological effects of sulfated compounds. First, we will use bioinformatic approaches to identity putative sulfotransferases (SULTs), enzymes that produce sulfated metabolites, in human gut bacteria. Using a comparative metabolomics workflow that we have developed that tracks the incorporation of labeled sulfur, we will further identify sulfated metabolites produced by SULT-containing gut bacteria, identify the responsible bacteria, and characterize the genes responsible for sulfated metabolite biosynthesis using both biochemical and genetic approaches. Second, we will investigate the effects of sulfated bacterial metabolites on T cell development, function, and trafficking in vitro and in vivo. Third, we will perform feeding studies in mice and ex vivo culturing experiments in human feces using labeled sulfate followed by comparative metabolomics in order to identify microbiome-dependent sulfated metabolites. We will also use microbial culturing workflows to isolate gut bacteria that produce sulfated metabolites. This work will broaden our understanding of the sulfated metabolome. By identifying microbiome-dependent sulfated metabolites, the genes responsible for their production, and the biological activities of these compounds, this research will lay the groundwork for the development of new small molecule and probiotic therapies to treat human disease, including inflammatory and autoimmune diseases.
