Oxygen offers considerable metabolic opportunities to aerobic organisms, but it also imposes substantial challenges to life. The Vazquez-Torres’ lab uses the model organism Salmonella to better understand the molecular mechanisms by which oxygen and its oxidative and nitrosative congeners exert damage to biomolecules. We actively study the adaptations that not only facilitate life in the midst of oxidative and nitrosative stress, but also promote persistence, pathogenicity and antibiotic resistance in clinically important bacteria. Specifically, we are examining how 1) redox signaling regulates bacterial RNA polymerase, 2) reactive species generated in the innate host response cause cytotoxicity, 3) metabolic reprograming protects against oxidative and nitrosative stress, and 4) metabolic adaptations to reactive species cross-protect against antibiotics.
- Redox signaling regulates bacterial RNA polymerase. Transcription in Gram-negative bacteria is under the allosteric control that DksA protein and guanosine tetraphosphate nucleotide alarmone exert on the secondary channel of RNA polymerase. Our investigations have revealed a previously unknown layer of transcriptional regulation that depends on redox-based, protein-protein interactions between DksA and the molecular chaperone DnaJ. Tripartite connections between DksA, DnaJ and guanosine tetraphosphate afford a dynamic range of transcriptional responses to various concentrations of reactive species associated with redox signaling or cytotoxicity. The adaptive response mediated by the combined actions of DksA, DnaJ and guanosine tetraphosphate is essential for Salmonella virulence. We are currently investigating the consequences that the competitive binding of DksA and Gre proteins to RNA polymerase has on transcriptional fidelity and DNA integrity.
- Reactive species generated in the innate host response cause cytotoxicity. Despite the wide range of biomolecules damaged by reactive species, as yet very few bacterial molecular targets have been identified. Our research has shown that reactive species posttranslationally modify transcriptional factors that are of pivotal importance for the regulation of metabolism and virulence. We have also demonstrated that cytochrome bd of the electron transport chain is a preferred target of nitric oxide. Investigations are in progress to determine how the nitrosylation of terminal cytochromes reprograms metabolism while eliciting antioxidant and antinitrosative defenses, antibiotic resistance, as well as bacterial persistence and pathogenicity.
- Metabolic reprograming protects against oxidative and nitrosative stress. Our investigations have demonstrated critical roles for glutathione, periplasmic superoxide dismutase, flavohemoprotein and a unique secretion system in the antioxidant and antinitrosative arsenal of Salmonella. In addition to these protective mechanisms, our recent work has revealed unsuspected roles for metabolic reprograming in the antioxidant and antinitrosative defenses of intracellular Salmonella. We are testing the hypothesis that glycolysis and fermentation protect against oxidative and nitrosative stress by assisting with ATP synthesis, balancing redox, and enabling disulfide bond formation in periplasmic proteins.
- Metabolic adaptations to reactive species cross-protect against antibiotics. Despite the strong antimicrobial activity that stems from the nitrosylation of terminal cytochromes of the electron transport chain, diverse bacteria, including Salmonella, Burkholderia and Pseudomonas, use the signaling cascade triggered by the nitrosylation of respiratory cytochromes to become tolerant to a variety of antibiotics of clinical relevance. Our laboratory is elucidating the molecular mechanisms by which nitrosative stress alters nucleotide metabolism and posttranslationally modifies ribosomal proteins, thereby protecting bacteria against antibiotics that inhibit cell wall or protein biosynthesis. In addition to identifying molecular mechanisms of antibiotic tolerance, these investigations will reveal important strategies exploited by human bacterial pathogens to persist in their hosts.
Funding that supports the research in the Vazquez-Torres’ lab:
R01 AI5449 NIAID. (PI, A. Vazquez-Torres). Title: “Analysis of intracellular host defenses in Salmonella pathogenesis.” The major goal of this project is to identify the molecular mechanisms underlying the reactive nitrogen species-mediated repression of Salmonella pathogenicity island-2 transcription. Dates approved: 9/30/03 – 05/30/19.
R01 AI136520 NIAID (PI, A. Vazquez-Torres). Title: “Molecular determinants of oxidative stress in Salmonella pathogenesis.” The major goal of this application is to characterize how central metabolism and the electron transport chain influence the antioxidant defenses of Salmonella. Dates approved: 09/01/18-08/30/23.
I01BX002073 VA-Merit Award. (PI, A. Vazquez-Torres). Title: “Molecular Analysis of Bacterial Adaptive Response to Host Reactive Species.” The goal is to elucidate the molecular mechanisms by which the RNA polymerase-binding regulatory protein DksA regulates bacterial adaptive responses to oxidative and nitrosative stress. Dates approved: 10/01/2013-03/30/2021.
T32 AI052066 Predoctoral Training Grant. (PD, A. Vazquez-Torres). Title: “Molecular Pathogenesis of Infectious Diseases.” The major goals of this pre-doctoral training grant are 1) to educate Ph.D. students in the investigation of fundamental mechanisms by which microbes cause infection, and 2) prepare our graduate students for scientific leadership positions. Dates approved: 09/30/2003-06/30/2023.