Georgetown University
Georgetown University, Brinsmade Lab
Georgetown University, Dept of Biology

About Our Lab

Introduction

Our lab is interdisciplinary in nature. We use cutting-edge and genome-wide methods to answer biological questions at the interface of metabolism and pathogenesis in Gram-positive bacteria. Currently, we are seeking to understand how a bacterium couples changes in intracellular abundance of key metabolites to changes in physiology and global gene expression, including genes that direct the synthesis of virulence factors. We are using the CodY regulon of Staphylococcus aureus as a model system.

Mapping intersecting metabolic and virulence gene expression patterns in S. aureus.

crystal structure of CodYIn S. aureus, CodY is activated as a DNA-binding protein by two classes of small molecule signals – the branched-chain amino acids isoleucine, leucine and valine (ILV), and GTP (Figure 1). We know the targets of CodY in S. aureus, but the threshold concentrations of active CodY required to regulate each gene remain unknown. We are systematically modulating CodY activity by using the unique cultivation conditions of a chemostat to titrate the abundance of CodY’s signals and by exploiting key genetic variants to block the reception of the ILV signal. Determining these threshold activities will allow us to construct a hierarchy (essentially a firing sequence) of gene expression mediated by CodY. Of particular interest to us is the location of the CodY-dependent virulence genes in this hierarchy. Are they turned on when CodY activity is moderately altered? Are they turned on when CodY activity drops substantially? Or, are the virulence genes turned on at different thresholds according to their role during infection of the host?

Staphylococcal pathogenesis is rooted in bacterial physiology.

CodY is central to metabolism

CodY is a global regulator of metabolism and pathogenesis gene expression in S. aureus. Genes under CodY control include those that control the rate of synthesis of its ILV signal, establishing a potential negative feedback loop. By sensing ILV availability, S. aureus can use CodY to monitor the status (or in fact adjust the activity) of multiple pathways that depend on ILV or its precursors. This can impact virulence factor production (Figure 2). We are collaborating with Professors Kyu Rhee and Elizabeth Alexander at Weill Cornell Medical College to understand how S. aureus uses CodY to reorient carbon and nitrogen metabolism to prioritize specific pathways during changes in nutrient availability. Adding labeled nutrients allows us to trace their fate. Understanding metabolic pathways favored under different environmental conditions may reveal new antimicrobial targets.


 

Quantifying changes in the CodY regulon upon induction of physiological stress response systems.

All cellular metabolites are connected to each other through their common origins in central metabolic pathways. Therefore, an environmental stress might indirectly alter the levels of metabolites used to measure nutrient sufficiency, including those monitored by CodY. We are testing the hypothesis that CodY is a central integrator of both nutritional limitation and environmental stress responses that are relevant to infection.