title: A Farnesylated Coxiella burnetii Effector Forms a Multimeric Complex at the Mitochondrial Outer Membrane during Infection.
abstract: Coxiella burnetii, the causative agent of Q fever, establishes a unique lysosome-derived intracellular niche termed theCoxiella-containing vacuole (CCV). The Dot/Icm-type IVB secretion system is essential for the biogenesis of the CCV and the intracellular replication ofCoxiellaEffector proteins, translocated into the host cell through this apparatus, act to modulate host trafficking and signaling processes to facilitate CCV development. Here we investigated the role of CBU0077, a conservedCoxiellaeffector that had previously been observed to localize to lysosomal membranes. CBU0077 was dispensable for the intracellular replication ofCoxiellain HeLa and THP-1 cells and did not appear to participate in CCV biogenesis. Intriguingly, native and epitope-tagged CBU0077 produced byCoxielladisplayed specific punctate localization at host cell mitochondria. As such, we designated CBU0077 MceA (mitochondrialCoxiellaeffector proteinA). Analysis of ectopically expressed MceA truncations revealed that the capacity to traffic to mitochondria is encoded within the first 84 amino acids of this protein. MceA is farnesylated by the host cell; however, this does not impact mitochondrial localization. Examination of mitochondria isolated from infected cells revealed that MceA is specifically integrated into the mitochondrial outer membrane and forms a complex of approximately 120 kDa. EngineeringCoxiellato express either MceA tagged with 3×FLAG or MceA tagged with 2×hemagglutinin allowed us to perform immunoprecipitation experiments that showed that MceA forms a homo-oligomeric species at the mitochondrial outer membrane during infection. This research reveals that mitochondria are a bona fide target ofCoxiellaeffectors and MceA is a complex-forming effector at the mitochondrial outer membrane duringCoxiellainfection.
authors.affiliation: Department of Biochemistry and Molecular Biology and Bio21 Molecular Science and Biotechnology Institute, The University of Melbourne, Melbourne, VIC, Australia.
authors.forename: Laura F
authors.initials: LF
authors.lastname: Fielden
authors.affiliation: Department of Microbiology and Immunology, University of Melbourne at the Peter Doherty Institute for Infection and Immunity, Melbourne, VIC, Australia.
authors.forename: Jennifer H
authors.initials: JH
authors.lastname: Moffatt
authors.affiliation: Department of Biochemistry and Molecular Biology and Bio21 Molecular Science and Biotechnology Institute, The University of Melbourne, Melbourne, VIC, Australia.
authors.forename: Yilin
authors.initials: Y
authors.lastname: Kang
authors.affiliation: Department of Biochemistry and Molecular Biology and Bio21 Molecular Science and Biotechnology Institute, The University of Melbourne, Melbourne, VIC, Australia.
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Baker
authors.affiliation: Department of Microbiology and Immunology, University of Melbourne at the Peter Doherty Institute for Infection and Immunity, Melbourne, VIC, Australia.
authors.forename: Chen Ai
authors.initials: CA
authors.lastname: Khoo
authors.affiliation: Department of Microbial Pathogenesis, Yale University School of Medicine, New Haven, Connecticut, USA.
authors.forename: Craig R
authors.initials: CR
authors.lastname: Roy
authors.affiliation: Department of Biochemistry and Molecular Biology and Bio21 Molecular Science and Biotechnology Institute, The University of Melbourne, Melbourne, VIC, Australia d.stojanovski@unimelb.edu.au hnewton@unimelb.edu.au.
authors.forename: Diana
authors.identifier: http://orcid.org/0000-0002-0199-3222
authors.initials: D
authors.lastname: Stojanovski
authors.affiliation: Department of Microbiology and Immunology, University of Melbourne at the Peter Doherty Institute for Infection and Immunity, Melbourne, VIC, Australia d.stojanovski@unimelb.edu.au hnewton@unimelb.edu.au.
authors.forename: Hayley J
authors.identifier: http://orcid.org/0000-0002-9240-2001
authors.initials: HJ
authors.lastname: Newton
mesh_terms: D002460:Cell Line; D016997:Coxiella burnetii / Q000254:growth & development* / Q000378:metabolism*; D004847:Epithelial Cells / Q000382:microbiology; D054884:Host-Pathogen Interactions; D006801:Humans; D051336:Mitochondrial Membranes / Q000378:metabolism*; D008970:Molecular Weight; D009000:Monocytes / Q000382:microbiology; D055503:Protein Multimerization; D011778:Q Fever / Q000382:microbiology*; D037521:Virulence Factors / Q000737:chemistry / Q000378:metabolism*
chemical_list: D037521:Virulence Factors
keywords: Coxiella burnetii; bacterial effector; host-pathogen interactions; mitochondria; prenylation