title: Discovery of a novel Toxoplasma gondii conoid-associated protein important for parasite resistance to reactive nitrogen intermediates.
abstract: Toxoplasma gondii modifies its host cell to suppress its ability to become activated in response to IFN-γ and TNF-α and to develop intracellular antimicrobial effectors, including NO. Mechanisms used by T. gondii to modulate activation of its infected host cell likely underlie its ability to hijack monocytes and dendritic cells during infection to disseminate to the brain and CNS where it converts to bradyzoites contained in tissue cysts to establish persistent infection. To identify T. gondii genes important for resistance to the effects of host cell activation, we developed an in vitro murine macrophage infection and activation model to identify parasite insertional mutants that have a fitness defect in infected macrophages following activation but normal invasion and replication in naive macrophages. We identified 14 independent T. gondii insertional mutants out of >8000 screened that share a defect in their ability to survive macrophage activation due to macrophage production of reactive nitrogen intermediates (RNIs). These mutants have been designated counter-immune mutants. We successfully used one of these mutants to identify a T. gondii cytoplasmic and conoid-associated protein important for parasite resistance to macrophage RNIs. Deletion of the entire gene or just the region encoding the protein in wild-type parasites recapitulated the RNI-resistance defect in the counter-immune mutant, confirming the role of the protein in resistance to macrophage RNIs.
authors.affiliation: Department of Microbiology and Immunology, New York Medical College, Valhalla, NY 10595, USA.
authors.forename: Sini
authors.initials: S
authors.lastname: Skariah
authors.forename: Robert B
authors.initials: RB
authors.lastname: Bednarczyk
authors.forename: Matthew K
authors.initials: MK
authors.lastname: McIntyre
authors.forename: Gregory A
authors.initials: GA
authors.lastname: Taylor
authors.forename: Dana G
authors.initials: DG
authors.lastname: Mordue
mesh_terms: D020121:5' Untranslated Regions / Q000235:genetics; D017398:Alternative Splicing; D000818:Animals; D003600:Cytosol / Q000737:chemistry; D017353:Gene Deletion; D017125:Genes, Protozoan; D008262:Macrophage Activation; D008264:Macrophages / Q000469:parasitology; D051379:Mice; D008810:Mice, Inbred C57BL; D016254:Mutagenesis, Insertional; D020030:Nitric Oxide Donors / Q000494:pharmacology; D015388:Organelles / Q000737:chemistry; D020033:Protein Isoforms / Q000737:chemistry / Q000502:physiology; D015800:Protozoan Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000502:physiology; D026361:Reactive Nitrogen Species / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D014122:Toxoplasma / Q000187:drug effects / Q000235:genetics / Q000502:physiology* / Q000648:ultrastructure
chemical_list: D020121:5' Untranslated Regions; D020030:Nitric Oxide Donors; D020033:Protein Isoforms; D015800:Protozoan Proteins; D026361:Reactive Nitrogen Species