Large-scale identification and translocation of type IV secretion substrates by Coxiella burnetii

Chen Chen; Simran Banga; Katja Mertens; Mary M Weber; Ivana Gorbaslieva; Yunhao Tan; Zhao-Qing Luo; James E Samuel

doi:10.1073/pnas.1010485107

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Large-scale identification and translocation of type IV secretion substrates by Coxiella burnetii

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Large-scale identification and translocation of type IV secretion substrates by Coxiella burnetii

Chen Chen, Simran Banga, Katja Mertens, Mary M Weber, Ivana Gorbaslieva, Yunhao Tan, Zhao-Qing Luo and James E Samuel

Proceedings of the National Academy of Sciences - PNAS, Vol.107(50), pp.21755-21760

12/14/2010

DOI: 10.1073/pnas.1010485107

PMCID: PMC3003115

PMID: 21098666

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Abstract

Coxiella burnetii is an obligate intracellular bacterial pathogen responsible for acute and chronic Q fever. This bacterium harbors a type IV secretion system (T4SS) highly similar to the Dot/Icm of Legionella pneumophila that is believed to be essential for its infectivity. Protein substrates of the Coxiella T4SS are predicted to facilitate the biogenesis of a phagosome permissive for its intracellular growth. However, due to the lack of genetic systems, protein transfer by the C. burnetii Dot/Icm has not been demonstrated. In this study, we report the identification of 32 substrates of the C. burnetii Dot/Icm system using a fluorescence-based β-lactamase (TEM1) translocation assay as well as the calmodulin-dependent adenylate cyclase (CyaA) assay in the surrogate host L. pneumophila . Notably, 26 identified T4SS substrates are hypothetical proteins without predicted function. Candidate secretion substrates were obtained by using ( i ) a genetic screen to identify C. burnetii proteins interacting with DotF, a component of the T4SS, and ( ii ) bioinformatic approaches to retrieve candidate genes that harbor characteristics associated with previously reported substrates of the Dot/Icm system from both C. burnetii and L. pneumophila . Moreover, we have developed a shuttle plasmid that allows the expression of recombinant proteins in C. burnetii as TEM fusion products. Using this system, we demonstrated that a Dot/Icm substrate identified with L. pneumophila was also translocated by C. burnetii in a process that requires its C terminus, providing direct genetic evidence of a functional T4SS in C. burnetii .

Biological Sciences

effectors

transporter

protein translocation

Details

Title: Subtitle: Large-scale identification and translocation of type IV secretion substrates by Coxiella burnetii
Creators: Chen Chen - Department of Microbial and Molecular Pathogenesis
Simran Banga - Department of Biological Sciences
Katja Mertens - Department of Microbial and Molecular Pathogenesis
Mary M Weber - Department of Microbial and Molecular Pathogenesis
Ivana Gorbaslieva - Department of Microbial and Molecular Pathogenesis
Yunhao Tan - Department of Biological Sciences
Zhao-Qing Luo - Department of Biological Sciences
James E Samuel - Department of Microbial and Molecular Pathogenesis
Resource Type: Journal article
Publication Details: Proceedings of the National Academy of Sciences - PNAS, Vol.107(50), pp.21755-21760
DOI: 10.1073/pnas.1010485107
PMID: 21098666
PMCID: PMC3003115
NLM abbreviation: Proc Natl Acad Sci U S A
ISSN: 0027-8424
eISSN: 1091-6490
Publisher: National Academy of Sciences
Language: English
Date published: 12/14/2010
Academic Unit: Microbiology and Immunology
Record Identifier: 9984083268402771

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