Global iTRAQ-based proteomic profiling of Toxoplasma gondii oocysts during sporulation

Zhou, Chun-Xue; Zhu, Xing‐Quan; Elsheikha, Hany M.; He, Shuai; Li, Qian; Zhou, Dong‐Hui; Suo, Xun

doi:10.1016/j.jprot.2016.07.010

Cited by 27 publications

(22 citation statements)

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“…Type II Prugniuad (PRU) strain of T. gondii was maintained in our laboratory by passaging the cysts in mice as described previously (Zhou et al, 2016). …”

Section: Methodsmentioning

confidence: 99%

“…Also, it allows parallel biological or technical replicates to be multiplexed (4-plex or 8-plex iTRAQ labeling) in one LC-MS/MS experiment, thus overcomes the inter-assay variations that occur in a single MS-based shotgun profiling experiment (Wu et al, 2006; Pierce et al, 2008; Craft et al, 2013). Also, this method has been successfully used to determine the differentially expressed proteins of T. gondii oocysts during sporulation (Zhou et al, 2016). The present study aimed to unravel the set of proteins that are unique to three life-cycle stages of T. gondii , namely oocyst, tachyzoite, and bradyzoites-containing cyst using 8-plex iTRAQ labeling and LC-MS/MS.…”

Section: Introductionmentioning

confidence: 99%

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Proteomic Differences between Developmental Stages of Toxoplasma gondii Revealed by iTRAQ-Based Quantitative Proteomics

et al. 2017

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Toxoplasma gondii has a complex two-host life-cycle between intermediate host and definitive host. Understanding proteomic variations across the life-cycle stages of T. gondii may improve the understanding of molecular adaption mechanism of T. gondii across life-cycle stages, and should have implications for the development of new treatment and prevention interventions against T. gondii infection. Here, we utilized LC–MS/MS coupled with iTRAQ labeling technology to identify differentially expressed proteins (DEPs) specific to tachyzoite (T), bradyzoites-containing cyst (C) and sporulated oocyst (O) stages of the cyst-forming T. gondii Prugniuad (Pru) strain. A total of 6285 proteins were identified in the three developmental stages of T. gondii. Our analysis also revealed 875, 656, and 538 DEPs in O vs. T, T vs. C, and C vs. O, respectively. The up- and down-regulated proteins were analyzed by Gene Ontology enrichment, KEGG pathway and STRING analyses. Some virulence-related factors and ribosomal proteins exhibited distinct expression patterns across the life-cycle stages. The virulence factors expressed in sporulated oocysts and the number of up-regulated virulence factors in the cyst stage were about twice as many as in tachyzoites. Of the 79 ribosomal proteins identified in T. gondii, the number of up-regulated ribosomal proteins was 33 and 46 in sporulated oocysts and cysts, respectively, compared with tachyzoites. These results support the hypothesis that oocyst and cystic stages are able to adapt to adverse environmental conditions and selection pressures induced by the host's immune response, respectively. These findings have important implications for understanding of the developmental biology of T. gondii, which may facilitate the discovery of novel therapeutic targets to better control toxoplasmosis.

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“…Type II Prugniuad (PRU) strain of T. gondii was maintained in our laboratory by passaging the cysts in mice as described previously (Zhou et al, 2016). …”

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Proteomic Differences between Developmental Stages of Toxoplasma gondii Revealed by iTRAQ-Based Quantitative Proteomics

et al. 2017

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“…Quite a few transcriptomic and proteomic studies examined the gene expression and protein abundance changes of the hosts upon parasite infection. For examples, RNA-Seq was used to look at the gene expression changes in cat liver, mouse liver and pig peripheral blood mononuclear cells (PBMCs) after infection (He et al, 2016a ; Zhou et al, 2016a , b ). Similarly, iTRAQ-based proteomic analysis on mice liver suggested that the relative abundances of hundreds of proteins were changed upon parasite infection, many of which were involved in key metabolic pathways (He et al, 2016b ).…”

Section: Molecular Basis For Pathogenesismentioning

confidence: 99%

Sixty Years (1957–2017) of Research on Toxoplasmosis in China—An Overview

et al. 2017

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Toxoplasma gondii is a ubiquitous zoonotic pathogen belonging to apicomplexan parasites. Infection in humans and animals may cause abortion and other severe symptoms under certain circumstances, leading to great economical losses and public health problems. T. gondii was first discovered in China in 1955 and the corresponding work was published in 1957. Since then, a lot of work has been done on this parasite and the diseases it causes. This review summarizes the major progress made by Chinese scientists over the last 60 years, and gives our perspectives on what should be done in the near future. A wide variety of diagnostic approaches were designed, including the ones to detect T. gondii specific antibodies in host sera, and T. gondii specific antigens or DNA in tissue and environmental samples. Further work will be needed to translate some of the laboratory assays into reliable products for clinic uses. Epidemiological studies were extensively done in China and the sero-prevalence in humans increased over the years, but is still below the world average, likely due to the unique eating and cooking habits. Infection rates were shown to be fairly high in meat producing animals such as, pigs, sheep, and chickens, as well as in the definitive host cats. Numerous subunit vaccines in the form of recombinant proteins or DNA vaccines were developed, but none of them is satisfactory in the current form. Live attenuated parasites using genetically modified strains may be a better option for vaccine design. The strains isolated from China are dominated by the ToxoDB #9 genotype, but it likely contains multiple subtypes since different ToxoDB #9 strains exhibited phenotypic differences. Further studies are needed to understand the general biology, as well as the unique features of strains prevalent in China.

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“…Following the nal wash, the supernatant was clari ed, the pellet was suspended in 5 volumes of sucrose solution (1.15 speci c gravity) and centrifugated for 10 mins at 350 g in order to concentrate oocysts. The oocysts were collected from the supernatant, blended in TE buffer (10 mM Tris-HCl, 1 mM EDTA, pH = 8.0) and puri ed applying a discontinuous CsCl density gradient method as previously described [15][16]. Finally, oocysts within the opaque-to-white layer were harvested and washed twice with 0.85% saline followed by resuspension in PBS and storage at 4 °C.…”

Section: Methods Production Puri Cation and Sporulation Of Oocystsmentioning

confidence: 99%

“…Phosphosite motifs are crucial amino acid sequences, which are involved in the recognition of substrate by the corresponding kinase [16][17]. To investigate the potential motifs of upregulated or downregulated phosphorylated peptides of sporulated oocysts between PYS strain and PRU strain, substantial preference of amino-acid residue in sequence from − 7 to + 7 surrounding the phosphorylation sites was analyzed by the motif-X software tool.…”

Section: Motifs Analysis Of Phosphorylation Sites Of Pys Strain Versumentioning

confidence: 99%

Global Phosphoproteome Analysis Reveals Significant Differences Between Sporulated Oocysts of Virulent and Avirulent Strains of Toxoplasma Gondii

Wang

Zhu

et al. 2020

Preprint

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Background: The apicomplexan protozoan parasite Toxoplasma gondii is one of the most successful intracellular parasites capable of infecting warm-blooded animals. Several strains of T. gondii have been described and typed as virulent or avirulent based on their pathogenicity in mice, as well as sporulated oocysts of strains belonging to distinct T. gondii genotypes. Phosphorylation is a reversable form of protein post-translational modification (PTM) that influences many biological processes and is used by some apicomplexan parasites to facilitate manipulation of the host cells. Phosphoproteomic analysis of oocysts of T. gondii strains belonging to distinct genotypes may reveal mechanisms contributing to the differences in the virulence of this parasite at the posttranslational level.Methods: In this study, the differences in the phosphoproteomic landscape of sporulated oocysts between virulent and avirulent strains of Toxoplasma gondii were examined using a global phosphoproteomics approach. Phosphopeptides from sporulated oocysts of the virulent PYS strain (Chinese ToxoDB#9) and the avirulent PRU strain (type II) were enriched by titanium dioxide (TiO2) affinity chromatography and quantified using isobaric tag (iBT) approach. Motif analysis, GO enrichment, KEGG pathway analysis, STRING analysis and kinase related network analysis of phosphopeptides were conducted to discover distinct difference in sporulated oocysts between virulent and avirulent T. gondii strains. Results: A total of 10,645 unique phosphopeptides, 8,181 nonredundant phosphorylation sites and 2,792 phosphoproteins were identified. We also detected 4,129 differentially expressed phosphopeptides (DEPs) between sporulated oocysts of 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 strain to PRU strain. Several kinases were consistent with motifs of overrepresented phosphopeptides, such as PKA, PKG, CKII, IKK, MAPK, EGFR, INSR, Jak, Syk, Src, Ab1. GO enrichment, KEGG pathway analysis and STRING analysis revealed DEPs significantly enriched in many biological processes and pathways. Kinase related network analysis showed that AGC kinase had the greatest connected peptides.Conclusions: The present study revealed the global phosphoproteomic differences in sporulated oocysts between virulent and avirulent T. gondii strains of different genotypes. Abundant phosphopeptides in sporulated oocysts between virulent and avirulent T. gondii strains exhibited distinct difference in the conserved motifs, GO terms, enriched pathways and PPI networks. The kinase associated network analysis indicated the role of phosphorylation in the transformation of T. gondii kinases. AGC kinase was the most connected kinases peptides. These data provide new insight into the phenotypic differences in sporulated oocysts of virulent and avirulent T. gondii strains.

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Global iTRAQ-based proteomic profiling of Toxoplasma gondii oocysts during sporulation

Cited by 27 publications

References 52 publications

Proteomic Differences between Developmental Stages of Toxoplasma gondii Revealed by iTRAQ-Based Quantitative Proteomics

Proteomic Differences between Developmental Stages of Toxoplasma gondii Revealed by iTRAQ-Based Quantitative Proteomics

Sixty Years (1957–2017) of Research on Toxoplasmosis in China—An Overview

Global Phosphoproteome Analysis Reveals Significant Differences Between Sporulated Oocysts of Virulent and Avirulent Strains of Toxoplasma Gondii

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