| Despite being designated a Category A potential biothreat by the Centers for Disease Control and Prevention, there is relatively little known about the molecular genetics of Francisella species pathogenesis. Recent in vivo genetic screens have identified hundreds of Francisella genes predicted to be involved in virulence, though in what way many of these genes may contribute to disease is unknown. Therefore, I began my thesis work with an intracellular replication screen to identify which virulence factors are specifically required for intracellular replication, an important component of Francisella infection. From the results of this screen, two novel proteins were chosen for further study: FTN_1133 and NaxD. Characterization of FTN_1133 revealed the role of this novel Ohr-like organic hydroperoxide resistance protein in withstanding reactive oxygen species both in macrophages and in mice. This is the first report of an Ohr-like protein involved in virulence. Next, NaxD, a member of the previously functionally uncharacterized YdjC superfamily of proteins, was shown to function as a deacetylase required for a lipid A modification important for pathogenesis, intramacrophage proliferation, and resistance to polymyxin B. Importantly, the role of NaxD was conserved in the human pathogenic F. tularensis as well as the mammalian pathogenic Bordetella bronchiseptica. Further studies indicate that this lipid A modification is required for resistance to specific intracellular host CAMPs and evasion of innate immune recognition. Taken together, the work presented in this dissertation elucidated the role of Francisella proteins in intramacrophage proliferation and resistance to host innate antimicrobial defenses. These studies have contributed not only to the understanding of Francisella 's genetic requirements for intracellular proliferation, but have revealed the role of Ohr-like proteins in pathogenesis and elucidated the function and role in virulence of members of the previously uncharacterized YdjC superfamily of proteins. Given that both Ohr and YdjC proteins are encoded by many virulent bacteria, this work has implications for our understanding of the pathogenesis of both intracellular and extracellular organisms and may represent attractive targets for drug and vaccine development. |
