Impact of environmental factors on bacterial bile acid transforming activity

Abstract Bacterial bile acid (BA) transformations have been studied for decades due to their relevance in several hu- man metabolic disorders and cancers. While the genes responsible for BA transformations have been the fo- cus area for many studies, their identification in bacterial genomes has proved a poor predictor of activity. Without a fundamental understanding of the factors that regulate BA transforming activity, the development of therapeutic interventions that leverage bacterial BA transforming activity to diagnose and treat human meta- bolic disorders remains elusive. The overall objective of this proposal is to determine how environmental fac- tors influence the regulation of genes responsible for BA transformations in communities of gut bacteria. The central hypothesis is that BA transforming activity is context dependent, impacted by the composition of the BA pool and host diet. This hypothesis was formulated based on preliminary data demonstrating that cocultured bacteria compete for available BA substrates, sometimes excluding expected BA transformations from occur- ring. In addition, bacterial BA transforming activity varies as a function of BA concentration and nutrient availa- bility. The rationale for this proposal is that bacterial BA transforming activity is regulated by physiologically rel- evant environmental factors that dictate the network of transformations that occur. The central hypothesis will be tested by two specific aims: 1) determine how BA composition shapes bacterial community composition and transforming activity under physiologically relevant conditions and 2) define the mechanistic contribution of die- tary fat on bacterial BA transforming activity in mice. For Aim 1, small synthetic communities of bacteria with varying capacity for BAs transformations will be tested for activity under a range of BA pools and BA concen- trations. Multi-omics analyses, in combination with quantitative BA measurements, will be performed to deter- mine how BA transforming pathways are regulated under these conditions and a multivariate multiple regres- sion analysis will be used to determine which factors are most likely to predict altered growth and BA trans- forming activity. For Aim 2, conventionally raised and gnotobiotic mice will be tested for their BA transforming activity on high-fat and low-fat diets. 16S sequencing will be used to determine bacterial community composi- tion and RT-qPCR will be used to measure host and bacterial gene expression related to BA transformations. The proposed research is significant because it will provide the fundamental knowledge needed to develop ef- fective therapeutic interventions capitalizing on the gut microbiome and BA pools to improve human health. The proposed aims will allow the applicant to acquire training in 1) the collection and analysis of multi-omics datasets and 2) the use of animal models to disentangle complex interactions between diet, the gut microbiota, and BA metabolism, which are essential to a career as an independent researcher in academia.