Protein export or secretion is one means by which pathogenic microbes injure the host cells of infected animals and humans. For example, bacteria export proteins that can kill host cells or promote adherence to tissues and block host defense mechanisms. Gram- negative bacterial pathogens utilize at least six distinct extracellular protein secretion systems to export proteins through their multi-layered cell envelope and in some cases into host cells. We propose to study the function of one such system, the Type VI secretion system (T6SS) that is expressed by numerous bacterial pathogens including Pseudomonas aeruginosa. P. aeruginosa is an opportunistic pathogen that frequently causes chronic lung infections in cystic fibrosis patients. The T6SS of this organism is an important virulence factor within the lung and thus is an exciting new target for vaccine and drug discovery. T6S apparatus is thought to be composed of 15-20 proteins whose biochemical function is not well understood. Our preliminary data suggest that proteins involved in T6SS function are structural analogs of a protein complex called the `phage tail'of the bacteriophage T4. This complex is a special machine that attacks the outer surface of host cells during the phage infection process. We propose to study whether the T6S apparatus of P. aeruginosa is a "nano-machine" similar to the phage tail complex, and introduces exported proteins into target cells via a similar membrane puncturing device. Thus, this proposal seeks funding for a research program to address the following general questions about T6SS: 1) Can we visualize components of the T6S apparatus and confirm its structural relationship to phage tail-like structures? 2) Does the T6S apparatus form channels in membranes that could be involved in protein deliver into target cells? 3) What controls the expression of the two other highly conserved T6SS gene clusters in P. aeruginosa? 4) Do components of the T6S apparatus appear on the surface of cells and do these actually spin? 5) Is it possible to control bacterial infection by developing drugs or vaccines targeting components of the T6SS?

Public Health Relevance

The bacterium Pseudomonas aeruginosa is the cause of severe infections in burn, immunocompromised and cystic fibrosis patients. In order to cause disease, this bacterium requires a tiny machine called the Type VI secretion system (T6SS). The T6SS likely transports toxic proteins into human cells as part of its function. The proposed study seeks to understand how the T6SS `nanomachine'drills holes into host cells and then injects proteins that kill or alter the function of human cells. Because no vaccine exists for this organism, these studies may also provide a new way to prevent disease due to this bacterium.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI026289-25
Application #
8416395
Study Section
Bacterial Pathogenesis Study Section (BACP)
Program Officer
Hall, Robert H
Project Start
1988-05-01
Project End
2014-07-31
Budget Start
2013-02-01
Budget End
2014-07-31
Support Year
25
Fiscal Year
2013
Total Cost
$483,904
Indirect Cost
$198,415
Name
Harvard University
Department
Microbiology/Immun/Virology
Type
Schools of Medicine
DUNS #
047006379
City
Boston
State
MA
Country
United States
Zip Code
02115
Ho, Brian T; Dong, Tao G; Mekalanos, John J (2014) A view to a kill: the bacterial type VI secretion system. Cell Host Microbe 15:9-21
Basler, Marek; Ho, Brian T; Mekalanos, John J (2013) Tit-for-tat: type VI secretion system counterattack during bacterial cell-cell interactions. Cell 152:884-94
Shneider, Mikhail M; Buth, Sergey A; Ho, Brian T et al. (2013) PAAR-repeat proteins sharpen and diversify the type VI secretion system spike. Nature 500:350-3
Fu, Yang; Waldor, Matthew K; Mekalanos, John J (2013) Tn-Seq analysis of Vibrio cholerae intestinal colonization reveals a role for T6SS-mediated antibacterial activity in the host. Cell Host Microbe 14:652-63
Danilchanka, Olga; Mekalanos, John J (2013) Cyclic dinucleotides and the innate immune response. Cell 154:962-70
Ho, Brian T; Basler, Marek; Mekalanos, John J (2013) Type 6 secretion system-mediated immunity to type 4 secretion system-mediated gene transfer. Science 342:250-3
Basler, M; Pilhofer, M; Henderson, G P et al. (2012) Type VI secretion requires a dynamic contractile phage tail-like structure. Nature 483:182-6
Davies, Bryan W; Bogard, Ryan W; Dupes, Nicole M et al. (2011) DNA damage and reactive nitrogen species are barriers to Vibrio cholerae colonization of the infant mouse intestine. PLoS Pathog 7:e1001295
Chin, Chen-Shan; Sorenson, Jon; Harris, Jason B et al. (2011) The origin of the Haitian cholera outbreak strain. N Engl J Med 364:33-42
Ma, Amy T; Mekalanos, John J (2010) In vivo actin cross-linking induced by Vibrio cholerae type VI secretion system is associated with intestinal inflammation. Proc Natl Acad Sci U S A 107:4365-70

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