title: Indispensable role for the eukaryotic-like ankyrin domains of the ankyrin B effector of Legionella pneumophila within macrophages and amoebae.
abstract: The Dot/Icm-translocated ankyrin B (AnkB) effector of Legionella pneumophila exhibits molecular mimicry of eukaryotic F-box proteins and is essential for intracellular replication in macrophages and protozoa. In addition to two eukaryotic-like ankyrin (ANK) domains, AnkB harbors a conserved eukaryotic F-box domain, which is involved in polyubiquitination of proteins throughout the eukaryotic kingdom. We have recently shown that the F-box domain of the AnkB effector is essential for decoration of the Legionella-containing vacuole (LCV) with polyubiquitinated proteins within macrophages and protozoan hosts. To decipher the role of the two ANK domains in the function of AnkB, we have constructed in-frame deletion of either or both of the ANK domain-encoding regions (ankB Delta A1, ankB Delta A2, and ankB Delta A1A2) to trans-complement the ankB null mutant. Deletion of the ANK domains results in defects in intracellular proliferation and decoration of the LCV with polyubiquitinated proteins. Export of the truncated variants of AnkB was reduced, and this may account for the observed defects. However, while full-length AnkB ectopically expressed in mammalian cells trans-rescues the ankB null mutant for intracellular proliferation, ectopic expression of AnkB Delta A1, AnkB Delta A2, and AnkB Delta A1A2 fails to trans-rescue the ankB null mutant. Importantly, ectopically expressed full-length AnkB is targeted to the host cell plasma membrane, where it recruits polyubiquitinated proteins. In contrast, AnkB Delta A1, AnkB Delta A2, and AnkB Delta A1A2 are diffusely distributed throughout the cytosol and fail to recruit polyubiquitinated proteins. We conclude that the two eukaryotic-like ANK domains of AnkB are essential for intracellular proliferation, for targeting AnkB to the host membranes, and for decoration of the LCV with polyubiquitinated proteins.
authors.affiliation: Department of Microbiology and Immunology, College of Medicine, University of Louisville, Louisville, KY 40202, USA.
authors.forename: Christopher T D
authors.initials: CT
authors.lastname: Price
authors.forename: Souhaila
authors.initials: S
authors.lastname: Al-Khodor
authors.forename: Tasneem
authors.initials: T
authors.lastname: Al-Quadan
authors.forename: Yousef
authors.initials: Y
authors.lastname: Abu Kwaik
mesh_terms: D000656:Amoeba / Q000382:microbiology*; D017487:Ankyrins / Q000502:physiology*; D001426:Bacterial Proteins / Q000502:physiology*; D002462:Cell Membrane / Q000737:chemistry; D017353:Gene Deletion; D005816:Genetic Complementation Test; D054884:Host-Pathogen Interactions; D006801:Humans; D016952:Legionella pneumophila / Q000472:pathogenicity*; D008264:Macrophages / Q000382:microbiology*; D017384:Sequence Deletion; D037521:Virulence Factors / Q000502:physiology*
chemical_list: D017487:Ankyrins; D001426:Bacterial Proteins; D037521:Virulence Factors