PROJECT SUMMARY The microbiota of the mammalian gut is a complex community of individual strains shaped in part by microbial competition over diet components. Despite computational analyses predicting enzymatic capacity of diverse bacteria, this knowledge is not sufficient to determine how diet influences bacterial abundance in the gut. In particular, little is known about regulation of carbon utilization enzymes in gut bacteria. Do they have mechanisms similar to E. coli carbon catabolite repression to consume preferred nutrients sequentially? Or do they consume all available nutrients simultaneously? How do these different strategies contribute to microbial abundance in the gut? We have identified a mechanism resembling carbon catabolite repression in Collinsella aerofaciens that may be a disadvantage when there is an abundance of secondary carbon source in the gut. Our laboratory seeks to characterize regulatory mechanisms governing carbon consumption in Collinsella species, in culture and in the mouse gut. Collinsella species are poorly studied Actinobacteria that are linked to chronic human diseases including type 2 diabetes and atherosclerosis. We have studied a group of closely related species and strains that vary in their regulation of carbon consumption. We will use this existing variability and experimental evolution to identify a common pathway of carbon catabolite repression in these bacteria. We will measure the heterogeneity of this pathway and related metabolic functions using single-cell RNA-seq. Finally, we will characterize the impact of this regulation on bacterial growth and competition in the mouse gut. Together, this research will define regulatory pathways that contribute to advantageous strategies in the complex nutrient environment of the mammalian intestine. Despite the vast number of correlative studies implicating a role for the gut microbiome in human disease, there remains much to explore in identifying bacterial metabolic pathways governing bacterial abundance and function in the gut. This gap limits both our understanding of the basic biology of these community interactions as well as the ability to design effective microbial therapeutics for human diseases characterized by complex microbial dysbiosis.