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Quantitative proteomic analysis of Toxoplasma gondii mutants of the mitochondrial and plastidic Iron sulfur cluster synthesis pathways.

ABSTRACT: Toxoplasma gondii is a parasitic protist that is the agent of toxoplasmosis. It is capable of infecting all mammals, including humans. The infection is mainly asymptomatic in immunocompetent patients, but in case of immunosuppression or for the congenital form of toxoplasmosis it can lead to severe pathologies with a possible fatal outcome. T. gondii contains two organelles of endosymbiotic origin: the mitochondrion and the apicoplast, which is a non-photosynthetic plastid. These organelles contain important biochemical pathways which might be interesting targets for future therapeutic strategies. Iron-sulfur clusters are one of the most ancient and ubiquitous prosthetic groups, and they are required by a variety of proteins involved in important metabolic processes. As for plants, T. gondii has several pathways for biosynthesis of iron-sulfur proteins, located in three different cellular compartments: the cytoplasm, the mitochondrion and the apicoplast. We have investigated the relative contributions of the mitochondrion and the apicoplast to the iron-sulfur proteome of the parasite by generating specific mutants for key proteins of the mitochondrial (TgIscU) and plastidic (TgSufS) pathways, on which we performed a quantitative proteomic analysis.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Toxoplasma Gondii Rh

DISEASE(S): Toxoplasmosis

SUBMITTER: sonia hem

LAB HEAD: Sébastien Besteiro

PROVIDER: PXD023854 | Pride | 2021-11-09

REPOSITORIES: Pride

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			Action	DRS
	PF_200618_ToxoFeS-I1-1.raw	Raw
	PF_200618_ToxoFeS-I1-2.raw	Raw
	PF_200618_ToxoFeS-I1-3.raw	Raw
	PF_200618_ToxoFeS-I3-1.raw	Raw
	PF_200618_ToxoFeS-I3-2.raw	Raw

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Project description:The population structure of Toxoplasma gondii includes three highly prevalent clonal lineages, types I, II, and III, which differ greatly in virulence in the mouse model. Previous studies have implicated a family of serine threonine protein kinases found in rhoptries (ROPs) as important in mediating virulence differences between types I vs. III and II vs. III. Here, we explored the genetic basis of differences in virulence between the highly virulent type I lineage and moderately virulent type II based on a new genetic cross and linkage mapping. Genome-wide association revealed a single quantitative trait locus controls the > 4 log difference in lethality between these strains. Neither ROP16 nor ROP18, previously implicated in virulence differences in T. gondii, were found to contribute to differences between types I and II. Instead, the major virulence locus contained a cluster of pseudokinases denoted as rhoptry protein 5 (ROP5); this locus contains a tandem cluster of polymorphic alleles that differed in expression levels between strains. ROP5 alleles contained only part of the catalytic triad of canonical S/T kinases, and consistent with this they lack demonstrable kinase activity in vitro. Genetic disruption of the rop5 locus in the type I lineage lead to a > 5 log increase in the lethal dose, and surviving mice developed lasting immunity and were protected from an otherwise lethal challenge. These findings reveal that amplification of a polymorphic cluster of pseudokinases plays an important role in pathogenesis of toxoplasmosis in the mouse model. We used a new genotyping protocol based on hybridization of parental and recombinant progeny genomic DNA (gDNA) to Affymetrix gene arrays constructed for T. gondii (http://ancillary.toxodb.org/docs/Array-Tutorial.html). We identified 1,603 single feature polymorphisms (SFP) based on probes that successfully discriminated perfect match (type II) from mismatch (type I) hybridization across all clones and provided reliable SFP markers for this cross.

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Quantitative proteomic analysis of Toxoplasma gondii mutants of the mitochondrial and plastidic Iron sulfur cluster synthesis pathways.

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