Francisella tularensis is a facultative intracellular pathogen of macrophages and the causative agent of tularemia. Although it is clear that inhalation of as few as ten F. tularensis subsp. tularensis can be fatal, and that macrophages are the major reservoir of this organism in vivo, how Ft evades elimination is only beginning to be defined. Within a few hours of uptake, Ft breaches the phagosome membrane and replicates in the cytosol. Our recent data demonstrate for the first time that fully virulent Ft strain SchuS4 disrupts phagosome maturation at an earlier stage of the pathway than has previously been appreciated and escapes from a novel compartment that excludes the late endoosome markers Rab7, mannose-6-phosphate receptor, lysobisphosphatidic acid and cathepsin S, despite local accumulation of lamp-1. Generation of this compartment requires live, MglA-positive Ft and is achieved via the ability of this pathogen to inhibit profoundly the activity of protein kinase C-a (PKCa) throughout infected cells. Thus, our data identify PKCa as the first host factor known to be targeted by Ft to evade intracellular killing. PKCa is also required for other aspects of innate defense, and our preliminary data support the hypothesis that MHC class II antigen presentation and oxidative defense mechanisms may also be impaired. At the same time, how Ft breaches the phagosome membrane is unclear, and we present evidence to support a model in which clathrin adapters and Rab family GTPases play important roles in phagosome dissolution and bacterial escape to the cytosol. From the bacterial perspective, few Francisella virulence factors have been identified and their mechanisms of action remain obscure. We recently developed a Tn5 transposon mutagenesis system for Francisella that we have used to identify mutants in FTL0347 and FTL1542 which encode a hypothetical membrane permease and acyl-CoA synthetase, respectively. Expression of known virulence factors is markedly reduced in both these mutants and preliminary characterization suggests that these genes are required for phagosome escape and inhibition of the phagocyte respiratory burst, respectively. Additionally, we constructed a TraSH (Transposon Site (Hybridization) derivative of our transposon that will allow us to identify novel genes required for Ft entry, growth and survival in human macrophages, Accordingly, our Specific Aims are: 1) to elucidate the mechanisms and functional consequences of Ft-mediated inhibition of PKCa signaling in macrophages;and 2) to identify and characterize novel F. tularensis genes required for evasion of innate macrophage defenses using transposon mutagenesis, including TRASH.

Public Health Relevance

Inhalation of a type of bacteria called Francisella tularensis causes severe, sometime fatal pneumonia, and no vaccine is available to prevent this infection. In this study we will define the mechanisms by which Francisella avoids being killed by macrophages (a type of immune system cell that can ingest and destroy most types of bacteria but not Francisella). The results of this study may lead to new treatments to combat or prevent this severe and often fatal lung infection.

National Institute of Health (NIH)
National Institute of Allergy and Infectious Diseases (NIAID)
Specialized Center--Cooperative Agreements (U54)
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Special Emphasis Panel (ZAI1-DDS-M)
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Washington University
Saint Louis
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Bandyopadhyay, Sarmistha; Long, Matthew E; Allen, Lee-Ann H (2014) Differential expression of microRNAs in Francisella tularensis-infected human macrophages: miR-155-dependent downregulation of MyD88 inhibits the inflammatory response. PLoS One 9:e109525
Virgin, Herbert W (2014) The virome in mammalian physiology and disease. Cell 157:142-50
Bialasiewicz, Seweryn; McVernon, Jodie; Nolan, Terry et al. (2014) Detection of a divergent Parainfluenza 4 virus in an adult patient with influenza like illness using next-generation sequencing. BMC Infect Dis 14:275
Rasmussen, Jed A; Post, Deborah M B; Gibson, Bradford W et al. (2014) Francisella tularensis Schu S4 lipopolysaccharide core sugar and O-antigen mutants are attenuated in a mouse model of tularemia. Infect Immun 82:1523-39
Patel, Dhara A; Patel, Anand C; Nolan, William C et al. (2014) High-throughput screening normalized to biological response: application to antiviral drug discovery. J Biomol Screen 19:119-30
Rohatgi, Anjali; Corbo, Joseph C; Monte, Kristen et al. (2014) Infection of myofibers contributes to increased pathogenicity during infection with an epidemic strain of chikungunya virus. J Virol 88:2414-25
Ermler, Megan E; Traylor, Zachary; Patel, Krupen et al. (2014) Rift Valley fever virus infection induces activation of the NLRP3 inflammasome. Virology 449:174-80
Moorman, Nathaniel J; Murphy, Eain A (2014) Roseomics: a blank slate. Curr Opin Virol 9:188-93
Canny, Susan P; Reese, Tiffany A; Johnson, L Steven et al. (2014) Pervasive transcription of a herpesvirus genome generates functionally important RNAs. MBio 5:e01033-13
Barker, Jason H; Kaufman, Justin W; Zhang, De-Sheng et al. (2014) Metabolic labeling to characterize the overall composition of Francisella lipid A and LPS grown in broth and in human phagocytes. Innate Immun 20:88-103

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