Unknown,Transcriptomics,Genomics,Proteomics

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Gene expression analysis of Hammondia hammondi and Toxoplasma gondii sporulated oocysts

ABSTRACT: To date little is known about the transcriptome of Hammondia hammondi, the nearest extant relative of Toxoplasma gondii. In this study we used an existing microarray to query Toxoplasma gondii and Hammondia hammondi transcript abundance in sporulated oocysts. Oocysts of the VEG strain of Toxoplasma gondii, and the HhCatGer041 strain of Hammondia hammondi, were isolated from cat feces by sucrose flotation, and sporulated for ~3-6 months in 2% sulfuric acid. RNA was isolated from Bleach-treated oocyst preparations using the Trizol reagent. RNA was biotinylated for hybridization to Toxo 169 Affymetrix chips using the Illumina Total Prep RNA labeling kit (Ambion). For each species 3 separate RNA isolations were performed on the same batch of oocysts and hybridized to individual microarrays.

ORGANISM(S): Toxoplasma gondii

SUBMITTER: Jon Boyle

PROVIDER: E-GEOD-61963 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Phosphorylation is a regulatory mechanism by which intracellular apicomplexan parasites control the process of invading and modifying host cells. However, the sporulated oocysts’ phosphoprotein repertoires between virulent and avirulent strains of Toxoplasma gondii and the underlying mechanisms contributing to virulence is still a mystery. A quantitative analysis of the sporulated oocysts’ phosphoproteome profile of T. gondii virulent and avirulent strains belonging to different genotypes was conducted to reveal phosphoprotein expression patterns that contribute to the virulence of a particular phenotype. Phosphopeptides from the sporulated oocysts of the virulent strain PYS (Chinese ToxoDB#9) and the avirulent strain type II (PRU strain) were enriched by titanium dioxide (TiO2) affinity chromatography and quantified using iBT technology. A total of 10,645 unique phosphopeptides, 8,181 nonredundant phosphorylation sites and 2,792 phosphoproteins were identified. The quantitative analysis identified 4,129 differentially expressed phosphopeptides (DEPs) between sporulated oocysts of the PYS strain and PRU strain (|log1.5 fold change| > 1 and P < 0.05), including 2,485 upregulated and 1,644 downregulated phosphopeptides. Motif analysis identified 24 motifs from the upregulated phosphorylated peptides including 22 serine motifs and two threonine motifs (TPE and TP), and 15 motifs from the downregulated phosphorylated peptides including 12 serine motifs and three threonine motifs (TP, RxxT and KxxT) in PYS strain when comparing PYS/PRU. Several kinases were consistent with motifs of overrepresented phosphopeptides, such as PKA, PKG, CKII, IKK, MAPK, EGFR, INSR, Jak, Syk, Src, Ab1. GO analysis, KEGG pathway analysis and STRING analysis revealed differentially expressed phosphoproteins (DEPs) exhibiting significant functional properties. Kinase associated network analysis showed that AGC kinase had the greatest connected peptides. Our findings reveal significant difference in phosphopeptides of sporulated oocysts between virulent and avirulent T. gondii strains, which provides solid foundation for further exploring the mechanisms contributing to the virulence of T. gondii.

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Gene expression analysis of Hammondia hammondi and Toxoplasma gondii sporulated oocysts

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