Project description:The apicomplexan parasite Toxoplasma gondii has a divergent mitochondrial electron transport chain (mETC). Complex III and IV associate together to form supercomplexes. We identified and deleted a Complex IV subunit (ApiCox10) which mediates this interaction. We perfomred immunoprecipitation of complex IV, via an HA epitope tagged Cox2a subunit in a parental and ApiCox10 knockout lines extracted in the mild detergent digitonin and analysed the elution via LFQ mass spectrometry. In the parental line, by immunoprecipitating Complex IV, complex III is also recovered, whereas in the ApiCox10 mutant only Complex IV is recovered due to impaired supercomplex formation.
Project description:Purpose: To shed light on the parasiticidal mechanisms of Altiratinib, we have adapted a workflow which combines a forward genetic approach based on transcriptome sequencing, computational mutation discovery and CRISPR/Cas9 genome editing in Toxoplasma gondii. Drug-resistant parasites were generated by chemical random mutagenesis. Multiple independent resistant lines were isolated. Single nucleotide variations (SNVs) were identified based on NGS transcriptomic analysis. By focusing on mutations present in coding sequences, we identified a single gene, TgPRP4K, that harbored SNVs leading to amino acid substitutions in 5 out of the 6 drug-resistant lines that were not present in the parental strain. Finally, using CRISPR/Cas9 genome editing we confirmed that the mutations identified confer resistance against Altiratinib.
Project description:Mutant T. gondii parasites lacking the hexokinase gene were viable. We carried genome wide expression analysis to identify genes that are differentially expressed in the mutant parasite with reference to wild type parental line. For this microarray analysis, we used the Affymetrix Toxoplasma gondii Custom GeneChip (CDF: ToxoDB version 5); Further details for this array can be found in the GEO platform ID GPL10000
Project description:TgFbxL2 interactome from Toxoplasma gondii tachyzoites were performed in 3 Biological Replicates, with 3 Technical Replicates each, by comparing FbxL2-HA tagged strain against parental, non-tagged strain.
Project description:TgFbxO13 interactome from Toxoplasma gondii tachyzoites were performed in 2 Biological Replicates, with 3 Technical Replicates each, by comparing FbxO13-HA tagged strain against parental, non-tagged strain.
Project description:TgFbxO1 interactome from Toxoplasma gondii tachyzoites were performed in 5 Biological Replicates, with 2 Technical Replicates each, by comparing FbxO1-HA tagged strain against parental, non-tagged strain.
Project description:TgFbxO14 interactome from Toxoplasma gondii tachyzoites were performed in 2 Biological Replicates, with 3 Technical Replicates each, by comparing FbxO14-HA tagged strain against parental, non-tagged strain.
Project description:Toxoplasma gondii is a zoonotic pathogen for which felids serve as definitive hosts. In cats, the parasite undergoes several rounds of asexual replication before entering the sexual cycle which gives rise to oocysts that are shed into the environment. These then sporulate and become infective to humans and live stock. To understand the genes involved in the parasite development in the felid host and identify potential intervention targets, we designed a transcriptomic approach to compare the cat intestinal stages with the well characterised tachyzoites that mediate acute infection and tissue cysts that are responsible for chronic infection. Cats were infected with T. gondii tissue cysts from mouse brain and sampled the intestinal stages at day 3, 5 and 7 post infection. As an input sample, we also collected tissue cysts from mouse brain as well as in vitro cultivated tachyzoites. Total RNA was extracted, enriched for mRNA and used for cDNA synthesis. RNA-Seq was then performed to describe the transcriptomic repertoire of each time point/life cycle stage.
