Infectious pathogens have devised strategies to evade host defenses. Intracellular respiratory pathogens such as Mycobacterium tuberculosis, Chlamydia pneumoniae, and Legionella pneumophila are all able to block phagosome-lysosome fusion, an important virulence determinant that allows these bacteria to replicate inside of mammalian cells. L. pneumophila is able to grow within alveolar macrophages in the lung. The dotA gene of L. pneumophila is required for intracellular growth and inhibition of phagosome-lysosome fusion. DotA is one of the only bacterial virulence factors identified that has a demonstrated role in blocking phagosome maturation in macrophages. DotA membrane topology suggests that this protein works in association with other L. pneumophila proteins to regulate phagosome trafficking in macrophages. The goal of the proposed research is to understand at a molecular level how L. pneumophila are able to regulate phagosome trafficking. Toward this end a mutational analysis will be conducted to characterize functional domains in the protein. Proteins that interact with DotA will be identified using molecular and genetic assays. The in vivo role of dotA will be determined by comparing the molecular biogenesis of L. pneumophila phagosomes that are formed by both wild type and dotA mutant bacteria. Studying the structure and function of the DotA protein will provide a foundation for our understanding of how intracellular bacterial pathogens that grow within the phagolysosome are able to exploit host cellular processes to establish infection.

National Institute of Health (NIH)
National Institute of Allergy and Infectious Diseases (NIAID)
First Independent Research Support & Transition (FIRST) Awards (R29)
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Bacteriology and Mycology Subcommittee 2 (BM)
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Yale University
Other Basic Sciences
Schools of Medicine
New Haven
United States
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Kohler, Lara J; Roy, Craig R (2017) Autophagic targeting and avoidance in intracellular bacterial infections. Curr Opin Microbiol 35:36-41
Shames, Stephanie R; Liu, Luying; Havey, James C et al. (2017) Multiple Legionella pneumophila effector virulence phenotypes revealed through high-throughput analysis of targeted mutant libraries. Proc Natl Acad Sci U S A 114:E10446-E10454
Tørring, Thomas; Shames, Stephanie R; Cho, Wooyoung et al. (2017) Acyl Histidines: New N-Acyl Amides from Legionella pneumophila. Chembiochem 18:638-646
Horenkamp, Florian A; Mukherjee, Shaeri; Alix, Eric et al. (2014) Legionella pneumophila subversion of host vesicular transport by SidC effector proteins. Traffic 15:488-99
Choy, Augustine; Roy, Craig R (2013) Autophagy and bacterial infection: an evolving arms race. Trends Microbiol 21:451-6
Mishra, Ashwini K; Del Campo, Claudia M; Collins, Robert E et al. (2013) The Legionella pneumophila GTPase activating protein LepB accelerates Rab1 deactivation by a non-canonical hydrolytic mechanism. J Biol Chem 288:24000-11
Campanacci, Valerie; Mukherjee, Shaeri; Roy, Craig R et al. (2013) Structure of the Legionella effector AnkX reveals the mechanism of phosphocholine transfer by the FIC domain. EMBO J 32:1469-77
Hori, Juliana I; Pereira, Marcelo S F; Roy, Craig R et al. (2013) Identification and functional characterization of K(+) transporters encoded by Legionella pneumophila kup genes. Cell Microbiol 15:2006-19
Choy, Augustine; Dancourt, Julia; Mugo, Brian et al. (2012) The Legionella effector RavZ inhibits host autophagy through irreversible Atg8 deconjugation. Science 338:1072-6
Alix, Eric; Mukherjee, Shaeri; Roy, Craig R (2011) Subversion of membrane transport pathways by vacuolar pathogens. J Cell Biol 195:943-52

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