The success of Mycobacterium tuberculosis, and other pathogenic mycobacteria species, lies in their ability to modulate their phagosome and prevent it from differentiating into an acidic, hydrolytically-competent compartment. The consensus in the field is that the bacterium arrests the normal maturation process leading to retention of the vacuole within the cell's recycling/sorting endosomal system. Although many studies detail the presence or absence of host molecules implicated in control of membrane fusion, the mechanism by which the bacterium modulates the phagosome remain to be determined. This proposal describes an integrated series of projects that will address this issue, place it in the context of the normal maturation process for phagosomes, and determine the consquences to the bacterium if it is delivered to the lysosome. The proposal addresses the following three aims: 1. Analysis of the lipid constituents of the phagosome membrane during maturation of IgG-bead and M. tuberculosis-containing phagosomes. The phosphorylation status of several membrane lipids changes during phagosome biogenesis and these alterations drive the association with the membrane fusion machinery in the cell. We intend to map these changes and correlate them with phenotypic differences in the pathogen-containing compartment. 2. Isolation and characterization of M. tuberculosis mutants defective in modulation of the phagosome. We have developed a genetic screen for such mutants and are currently characterizing the genetic defects that prevent modulation of the phagosome. These mutants should provide the tools that we need to determine how the bacterium regulates its phagosome. 3. Identification of lysosomal constituents that are bactericidal to M. tuberculosis. We have found that isolated lysosomes kill M. tuberculosis when added to bacterial cultures. Preliminary characterization of this fraction suggests that the activity co-purifies with cathepsins S and B. We intend to identify the active component and characterize the mechanism that leads to bacterial death.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI057086-03
Application #
6849336
Study Section
AIDS and Related Research 8 (AARR)
Program Officer
Sizemore, Christine F
Project Start
2003-07-01
Project End
2007-12-31
Budget Start
2005-01-01
Budget End
2005-12-31
Support Year
3
Fiscal Year
2005
Total Cost
$478,231
Indirect Cost
Name
Cornell University
Department
Microbiology/Immun/Virology
Type
Schools of Veterinary Medicine
DUNS #
872612445
City
Ithaca
State
NY
Country
United States
Zip Code
14850
Podinovskaia, Maria; Lee, Wonsik; Caldwell, Shannon et al. (2013) Infection of macrophages with Mycobacterium tuberculosis induces global modifications to phagosomal function. Cell Microbiol 15:843-59
Russell, David G (2011) Mycobacterium tuberculosis and the intimate discourse of a chronic infection. Immunol Rev 240:252-68
Abramovitch, Robert B; Rohde, Kyle H; Hsu, Fong-Fu et al. (2011) aprABC: a Mycobacterium tuberculosis complex-specific locus that modulates pH-driven adaptation to the macrophage phagosome. Mol Microbiol 80:678-94
Mann, Francis M; VanderVen, Brian C; Peters, Reuben J (2011) Magnesium depletion triggers production of an immune modulating diterpenoid in Mycobacterium tuberculosis. Mol Microbiol 79:1594-601
Russell, David G (2011) The galvanizing of Mycobacterium tuberculosis: an antimicrobial mechanism. Cell Host Microbe 10:181-3
Russell, David G; Barry 3rd, Clifton E; Flynn, JoAnne L (2010) Tuberculosis: what we don't know can, and does, hurt us. Science 328:852-6
VanderVen, Brian C; Hermetter, Albin; Huang, Amy et al. (2010) Development of a novel, cell-based chemical screen to identify inhibitors of intraphagosomal lipolysis in macrophages. Cytometry A 77:751-60
Homolka, Susanne; Niemann, Stefan; Russell, David G et al. (2010) Functional genetic diversity among Mycobacterium tuberculosis complex clinical isolates: delineation of conserved core and lineage-specific transcriptomes during intracellular survival. PLoS Pathog 6:e1000988
Russell, David G; VanderVen, Brian C; Lee, Wonsik et al. (2010) Mycobacterium tuberculosis wears what it eats. Cell Host Microbe 8:68-76
Purdy, Georgiana E; Niederweis, Michael; Russell, David G (2009) Decreased outer membrane permeability protects mycobacteria from killing by ubiquitin-derived peptides. Mol Microbiol 73:844-57

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