Stringent Response Governs the Virulence and Oxidative Stress Resistance of Francisella tularensis
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Issue Date
2019-07
Authors
King, Kayla '19
Degree
Advisor
Malik, Meenakshi
Committee Members
Sharifi, H. John
Parent, Michelle
Parent, Michelle
Journal Title
Journal ISSN
Volume Title
Abstract
Francisella tularensis (F. tularensis) is a Gram-negative facultative intracellular human pathogen responsible for causing tularemia. It is also classified as a Category A agent by the Centers for Disease Control (CDC) based on its possible use as a bioterror agent. Although the molecular basis for the high infectivity and virulence of F. tularensis is not well understood, the pathogenicity of F. tularensis is mainly dependent on its ability to persist and replicate in phagocytic cells. To survive and replicate inside the cells, F. tularensis has to resist the attack of host-generated reactive oxygen and nitrogen species. Several antioxidant enzymes that are important virulence factors against oxidative stress have been identified and implicated in the pathogenesis of F. tularensis in various studies.
In the present study, we investigated the roles of (p)ppGpp; the stringent response molecules produced by RelA and SpoT proteins, in nutritional, environmental, oxidative stress responses, and virulence of F. tularensis LVS. We generated a relA gene deletion mutant (?relA), a relA/spoT double gene deletion mutant (?relA?spoT), and their corresponding transcomplemented strains. These mutant strains were characterized for their sensitivity towards nutritionally challenging conditions induced by amino acid starvation, amino acid and carbohydrate limiting growth conditions, exposure to antibiotics and high temperatures, and survival in macrophages and mice. We also investigated how the stringent response alters the oxidative stress response by comparing the growth of wild type F. tularensis LVS, the mutants, and the transcomplemented strains in the presence of oxidants. We studied transcriptional profiling by exposing ?relA?spoT mutant to oxidative stress conditions and compared it with that of the wild type F. tularensis LVS. The results from these studies demonstrate that RelA/SpoT-mediated stringent response in F. tularensis plays an important role in providing resistance against nutritional, environmental, and oxidative stresses, and contributes to intramacrophage survival and virulence in mice. Transcriptome analysis revealed that stringent response plays an essential role in the regulation of a large number of genes that aid in the survival of the Francisella under oxidative stress conditions. This study provides a novel link between the stringent response and the oxidative stress response and demonstrates how stringent response governs the oxidative stress responses of F. tularensis.
In the present study, we investigated the roles of (p)ppGpp; the stringent response molecules produced by RelA and SpoT proteins, in nutritional, environmental, oxidative stress responses, and virulence of F. tularensis LVS. We generated a relA gene deletion mutant (?relA), a relA/spoT double gene deletion mutant (?relA?spoT), and their corresponding transcomplemented strains. These mutant strains were characterized for their sensitivity towards nutritionally challenging conditions induced by amino acid starvation, amino acid and carbohydrate limiting growth conditions, exposure to antibiotics and high temperatures, and survival in macrophages and mice. We also investigated how the stringent response alters the oxidative stress response by comparing the growth of wild type F. tularensis LVS, the mutants, and the transcomplemented strains in the presence of oxidants. We studied transcriptional profiling by exposing ?relA?spoT mutant to oxidative stress conditions and compared it with that of the wild type F. tularensis LVS. The results from these studies demonstrate that RelA/SpoT-mediated stringent response in F. tularensis plays an important role in providing resistance against nutritional, environmental, and oxidative stresses, and contributes to intramacrophage survival and virulence in mice. Transcriptome analysis revealed that stringent response plays an essential role in the regulation of a large number of genes that aid in the survival of the Francisella under oxidative stress conditions. This study provides a novel link between the stringent response and the oxidative stress response and demonstrates how stringent response governs the oxidative stress responses of F. tularensis.
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