How do hybrid two-component systems in gut bacteria regulate global gene expression?

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The Bacteroides are key members of the healthy human and farmed-animal gut microbiota. Their ability to degrade a huge variety of complex polysaccharides from host, dietary and microbial sources is key to their ubiquitous colonisation of the human gut. The challenge facing these bacteria is sensing and rapidly responding to the constant nutrient fluctuations in the competitive, environment of the gut. They target different glycans via loci known as polysaccharide utilisation loci (PULs). Typically, each PUL is targeted towards a different glycan, and encodes cell surface enzymes, binding proteins and transporters. Upon import of partially degraded oligosaccharides into the periplasm, a specific oligosaccharide cue is bound and recognised by a regulator protein known as a hybrid two-component system (HTCS). HTCS comprise all the domains of a typical bacterial two-component system (sensor domain, histidine kinase, acceptor domain and DNA binding domain), but in a single transmembrane protein. The activation of an HTCS by binding oligosaccharide then leads to rapid upregulation of PUL expression by 2-3 orders of magnitude. This response enables the bacterium to import and degrade the available glycan rapidly. Previous data shows that each regulator senses a different oligosaccharide, allowing the bacterium to distinguish between linkage and monosaccharide content of a target glycan. Intriguingly, the DNA binding domains are not cleaved into the cytosol upon activation, suggesting their DNA targets must be in close proximity to the membrane. This project will investigate how these HTCS are able to directly regulate gene expression at the membrane, using a combination of biochemistry, bacterial genetics, and global transcriptional analysis (ChIP-Seq and RNA-Seq). This project will enable us to understand the mechanism by which these unusual regulators are able to direct the rapid expression of carbohydrate active enzymes, in response to nutrient fluctuations in gut microbiomes. This paves the way for rational engineering of these strains to release specific molecules or enzymes in response to dietary interventions, enhancing their probiotic properties.This project fits within the agriculture and food security strategic aim of the BBSRC, specifically the food safety and nutrition focus, which includes mechanistic interactions between food/feed and the microbiome. Bacteroidetes bacteria are the dominant degraders of complex carbohydrates in human and animals guts, and understanding how they are able to recognise and specifically respond to certain carbohydrates is critical to understanding their success in colonising the gut. The genetic tractability of these organisms means that once we understand how they regulate their response to carbohydrates at the molecular level, we can begin to manipulate them as potential probiotic strains with e.g. enzymatic activity or secretion regulated by specific dietary fibre/supplements.