Autophagy-mediated reentry of Francisella tularensis into the endocytic compartment after cytoplasmic replication

Tularemia Pathogenesis Section, Laboratory of Intracellular Parasites, Rocky Mountain Laboratories, National Institute of Allergy and Infectious Diseases, National Institutes of Health, 903 South Fourth Street, Hamilton, MT 59840, USA.
Proceedings of the National Academy of Sciences (Impact Factor: 9.67). 10/2006; 103(39):14578-83. DOI: 10.1073/pnas.0601838103
Source: PubMed


Intracellular bacterial pathogens evade the bactericidal functions of mammalian cells by physical escape from their phagosome and replication into the cytoplasm or through the modulation of phagosome maturation and biogenesis of a membrane-bound replicative organelle. Here, we detail in murine primary macrophages the intracellular life cycle of Francisella tularensis, a highly infectious bacterium that survives and replicates within mammalian cells. After transient interactions with the endocytic pathway, bacteria escaped from their phagosome by 1 h after infection and underwent replication in the cytoplasm from 4 to 20 h after infection. Unexpectedly, the majority of bacteria were subsequently found to be enclosed within large, juxtanuclear, LAMP-1-positive vacuoles called Francisella-containing vacuoles (FCVs). FCV formation required intracytoplasmic replication of bacteria. Using electron and fluorescence microscopy, we observed that the FCVs contained morphologically intact bacteria, despite fusing with lysosomes. FCVs are multimembranous structures that accumulate monodansylcadaverine and display the autophagy-specific protein LC3 on their membrane. Formation of FCVs was significantly inhibited by 3-methyladenine, confirming a role for the autophagic pathway in the biogenesis of these organelles. Taken together, our results demonstrate that, via autophagy, F. tularensis reenters the endocytic pathway after cytoplasmic replication, a process thus far undescribed for intracellular pathogens.

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    • "During later intervals of infection, some F. tularensis have been observed in the multi-membrane vacuolar compartment of the endocytic pathway that has the characteristics of an autophagosome [41] [42]. Still unclear, however, are the reason why F. tularensis reenters the membranous compartment and the consequences for the further dissemination of infection and induction of immune response (the scheme of whole process see Fig. 1). "
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    ABSTRACT: Tularemia is a debilitating febrile and potentially fatal zoonotic disease of humans and other vertebrates caused by the Gram-negative bacterium Francisella tularensis. The natural reservoirs are small rodents, hares, and possibly amoebas in water. The etiological agent, Francisella tularensis, is a non-spore forming, encapsulated, facultative intracellular bacterium, a member of the γ-Proteobacteria class of Gram-negative bacteria. Francisella tularensis is capable of invading and replicating within phagocytic as well as non-phagocytic cells and modulate inflammatory response. Infection by the pulmonary, dermal, or oral routes, respectively, results in pneumonic, ulceroglandular, or oropharyngeal tularemia. The highest mortality rates are associated with the pneumonic form of this disease. All members of Francisella tularensis species cause more or less severe disease Due to their abilities to be transmitted to humans via multiple routes and to be disseminated via biological aerosol that can cause the disease after inhalation of even an extremely low infectious dose, Francisella tularensis has been classified as a Category A bioterrorism agent. The current standard of care for tularemia is treatment with antibiotics, as this therapy is highly effective if used soon after infection, although it is not, however, absolutely effective in all cases.
    Central European Journal of Biology 03/2015; 10(1):117-138. DOI:10.1515/biol-2015-0013 · 0.71 Impact Factor
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    • "In addition, invasive intracellular bacteria can be targeted and engulfed into autophagosomes for their eventual degradation in autolysosomes through a subtype of autophagy, known as xenophagy (Deretic, 2011; Knodler and Celli, 2011; Levine, 2005; Yuk et al., 2012). Eukaryotic cells can deploy xenophagy to restrict the rapid multiplication of internalized extracellular pathogens such as group A Streptococcus (GAS) (Nakagawa et al., 2004; Sakurai et al., 2010; Yamaguchi et al., 2009) or typical intracellular pathogen such as Francisella tularensis (Checroun et al., 2006) and Listeria monocytogenes (Py et al., 2007; Yano et al., 2008). "
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    ABSTRACT: Macroautophagy (hereinafter called autophagy) is a highly regulated process used by eukaryotic cells to digest portions of the cytoplasm that remodels and recycles nutrients and disposes of unwanted cytoplasmic constituents. Currently 36 autophagy-related genes (ATG) and their homologs have been characterized in yeast and higher eukaryotes, including insects. In the present study, we identified and functionally characterized the immune function of an ATG8 homolog in a coleopteran insect, Tenebrio molitor (TmATG8). The cDNA of TmATG8 comprises of an ORF of 363 bp that encodes a protein of 120 amino acid residues. TmATG8 transcripts are detected in all the developmental stages analyzed. TmAtg8 protein contains a highly conserved C-terminal glycine residue (Gly116) and shows high amino acid sequence identity (98%) to its Tribolium castaneum homolog, TcAtg8. Loss of function of TmATG8 by RNAi led to a significant increase in the mortality rates of T. molitor larvae against Listeria monocytogenes. Unlike dsEGFP-treated control larvae, TmATG8-silenced larvae failed to turn-on autophagy in hemocytes after injection with L. monocytogenes. These data suggest that TmATG8 play a role in mediating autophagy-based clearance of Listeria in T. molitor.
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    • "This process is summarized in Figure 1. There is evidence that following escape of F. tularensis LVS into the cytoplasm, a proportion of cytoplasmic bacteria re-enters the endocytic pathway through autophagy.44 It is not certain whether this process is an adaptive strategy by the bacterium or a part of the cellular defense against infection; however, evidence suggests that autophagy provides intracellular bacteria with nutrients required for replication.45 "
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    ABSTRACT: Francisella tularensis is an intracellular Gram-negative bacterium that causes life-threatening tularemia. Although the prevalence of natural infection is low, F. tularensis remains a tier I priority pathogen due to its extreme virulence and ease of aerosol dissemination. F. tularensis can infect a host through multiple routes, including the intradermal and respiratory routes. Respiratory infection can result from a very small inoculum (ten organisms or fewer) and is the most lethal form of infection. Following infection, F. tularensis employs strategies for immune evasion that delay the immune response, permitting systemic distribution and induction of sepsis. In this review we summarize the current knowledge of F. tularensis in an immunological context, with emphasis on the host response and bacterial evasion of that response.
    Infection and Drug Resistance 09/2014; 7:239-51. DOI:10.2147/IDR.S53700
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