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Distinct strains of Toxoplasma gondii feature divergent transcriptomes regardless of developmental stage.

Croken MM, Ma Y, Markillie LM, Taylor RC, Orr G, Weiss LM, Kim K - PLoS ONE (2014)

Bottom Line: These functions are likely associated with cell-cycle arrest.Using GSEA, we identified that ribosomal proteins were expressed at significantly higher levels in the CTG strain than in either the RH or PLK strains.This association holds true regardless of life cycle stage.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, Albert Einstein College of Medicine, Bronx, New York, United States of America.

ABSTRACT
Using high through-put RNA sequencing, we assayed the transcriptomes of three different strains of Toxoplasma gondii representing three common genotypes under both in vitro tachyzoite and in vitro bradyzoite-inducing alkaline stress culture conditions. Strikingly, the differences in transcriptional profiles between the strains, RH, PLK, and CTG, is much greater than differences between tachyzoites and alkaline stressed in vitro bradyzoites. With an FDR of 10%, we identified 241 genes differentially expressed between CTG tachyzoites and in vitro bradyzoites, including 5 putative AP2 transcription factors. We also observed a close association between cell cycle regulated genes and differentiation. By Gene Set Enrichment Analysis (GSEA), there are a number of KEGG pathways associated with the in vitro bradyzoite transcriptomes of PLK and CTG, including pyrimidine metabolism and DNA replication. These functions are likely associated with cell-cycle arrest. When comparing mRNA levels between strains, we identified 1,526 genes that were differentially expressed regardless of culture-condition as well as 846 differentially expressed only in bradyzoites and 542 differentially expressed only in tachyzoites between at least two strains. Using GSEA, we identified that ribosomal proteins were expressed at significantly higher levels in the CTG strain than in either the RH or PLK strains. This association holds true regardless of life cycle stage.

No MeSH data available.


Related in: MedlinePlus

Expression of cell-cycle genes altered during differentiation.A) Boxplot represents differences in expression levels of cell cycle-dependent genes following differentiation conditions. Green boxes represent changes in S/M gene expression, blue boxes represent changes in G1 gene expression. A positive difference indicates up regulation of the gene in tachyzoite conditions, while a negative difference in expression values indicates greater expression in the alkaline stress induced bradyzoites. The black bar indicates median value; the red cross indicates mean value. Significance tested by one-way ANOVA. A star (*) indicates: p<0.001. B) Cell cycle genes are annotated as either G1 or S/M [8]. We then sorted genes into groups based on time of peak expression. Each of these gene sets was tested by GSEA [26]. Gene sets with significant (FWER-p value <0.05) normalized enrichment scores (NES) are plotted. Positive scores indicate association with the unstressed (tachyzoite) condition, negative scores indicate association with the stress (bradyzoite) condition. Blue and green bar across middle of plots represent an eight hour RH tachyzoite cell cycle. Counter-clockwise from the top, the plots show cell cycle gene sets influenced following RH differentiation, cell cycle gene sets influenced following PLK differentiation, cell cycle gene sets influenced following CTG differentiation. Note that time of expression is based on the RH cell cycle as defined, which is shorter than that of either PLK or CTG.
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pone-0111297-g004: Expression of cell-cycle genes altered during differentiation.A) Boxplot represents differences in expression levels of cell cycle-dependent genes following differentiation conditions. Green boxes represent changes in S/M gene expression, blue boxes represent changes in G1 gene expression. A positive difference indicates up regulation of the gene in tachyzoite conditions, while a negative difference in expression values indicates greater expression in the alkaline stress induced bradyzoites. The black bar indicates median value; the red cross indicates mean value. Significance tested by one-way ANOVA. A star (*) indicates: p<0.001. B) Cell cycle genes are annotated as either G1 or S/M [8]. We then sorted genes into groups based on time of peak expression. Each of these gene sets was tested by GSEA [26]. Gene sets with significant (FWER-p value <0.05) normalized enrichment scores (NES) are plotted. Positive scores indicate association with the unstressed (tachyzoite) condition, negative scores indicate association with the stress (bradyzoite) condition. Blue and green bar across middle of plots represent an eight hour RH tachyzoite cell cycle. Counter-clockwise from the top, the plots show cell cycle gene sets influenced following RH differentiation, cell cycle gene sets influenced following PLK differentiation, cell cycle gene sets influenced following CTG differentiation. Note that time of expression is based on the RH cell cycle as defined, which is shorter than that of either PLK or CTG.

Mentions: In keeping with the existing model, Figure 4A shows that S/M genes are more highly expressed in bradyzoite populations while G1 genes are more closely associated with tachyzoites. In bradyzoite differentiated parasites, there is an up regulation of S/M associated genes, while tachyzoites have higher steady state expression levels of G1-linked genes. The mean difference of each group (red cross) also illustrates these relationships. Further, there is again a clear difference in how each strain is affected by stress (significant difference of means by ANOVA). The cell cycle genes of RH are relatively unaffected by differentiation stress, while PLK experiences significant changes in expression of these genes and CTG more so. These data underscore the fundamental link between cell cycle regulation and life cycle advancement.


Distinct strains of Toxoplasma gondii feature divergent transcriptomes regardless of developmental stage.

Croken MM, Ma Y, Markillie LM, Taylor RC, Orr G, Weiss LM, Kim K - PLoS ONE (2014)

Expression of cell-cycle genes altered during differentiation.A) Boxplot represents differences in expression levels of cell cycle-dependent genes following differentiation conditions. Green boxes represent changes in S/M gene expression, blue boxes represent changes in G1 gene expression. A positive difference indicates up regulation of the gene in tachyzoite conditions, while a negative difference in expression values indicates greater expression in the alkaline stress induced bradyzoites. The black bar indicates median value; the red cross indicates mean value. Significance tested by one-way ANOVA. A star (*) indicates: p<0.001. B) Cell cycle genes are annotated as either G1 or S/M [8]. We then sorted genes into groups based on time of peak expression. Each of these gene sets was tested by GSEA [26]. Gene sets with significant (FWER-p value <0.05) normalized enrichment scores (NES) are plotted. Positive scores indicate association with the unstressed (tachyzoite) condition, negative scores indicate association with the stress (bradyzoite) condition. Blue and green bar across middle of plots represent an eight hour RH tachyzoite cell cycle. Counter-clockwise from the top, the plots show cell cycle gene sets influenced following RH differentiation, cell cycle gene sets influenced following PLK differentiation, cell cycle gene sets influenced following CTG differentiation. Note that time of expression is based on the RH cell cycle as defined, which is shorter than that of either PLK or CTG.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4230917&req=5

pone-0111297-g004: Expression of cell-cycle genes altered during differentiation.A) Boxplot represents differences in expression levels of cell cycle-dependent genes following differentiation conditions. Green boxes represent changes in S/M gene expression, blue boxes represent changes in G1 gene expression. A positive difference indicates up regulation of the gene in tachyzoite conditions, while a negative difference in expression values indicates greater expression in the alkaline stress induced bradyzoites. The black bar indicates median value; the red cross indicates mean value. Significance tested by one-way ANOVA. A star (*) indicates: p<0.001. B) Cell cycle genes are annotated as either G1 or S/M [8]. We then sorted genes into groups based on time of peak expression. Each of these gene sets was tested by GSEA [26]. Gene sets with significant (FWER-p value <0.05) normalized enrichment scores (NES) are plotted. Positive scores indicate association with the unstressed (tachyzoite) condition, negative scores indicate association with the stress (bradyzoite) condition. Blue and green bar across middle of plots represent an eight hour RH tachyzoite cell cycle. Counter-clockwise from the top, the plots show cell cycle gene sets influenced following RH differentiation, cell cycle gene sets influenced following PLK differentiation, cell cycle gene sets influenced following CTG differentiation. Note that time of expression is based on the RH cell cycle as defined, which is shorter than that of either PLK or CTG.
Mentions: In keeping with the existing model, Figure 4A shows that S/M genes are more highly expressed in bradyzoite populations while G1 genes are more closely associated with tachyzoites. In bradyzoite differentiated parasites, there is an up regulation of S/M associated genes, while tachyzoites have higher steady state expression levels of G1-linked genes. The mean difference of each group (red cross) also illustrates these relationships. Further, there is again a clear difference in how each strain is affected by stress (significant difference of means by ANOVA). The cell cycle genes of RH are relatively unaffected by differentiation stress, while PLK experiences significant changes in expression of these genes and CTG more so. These data underscore the fundamental link between cell cycle regulation and life cycle advancement.

Bottom Line: These functions are likely associated with cell-cycle arrest.Using GSEA, we identified that ribosomal proteins were expressed at significantly higher levels in the CTG strain than in either the RH or PLK strains.This association holds true regardless of life cycle stage.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, Albert Einstein College of Medicine, Bronx, New York, United States of America.

ABSTRACT
Using high through-put RNA sequencing, we assayed the transcriptomes of three different strains of Toxoplasma gondii representing three common genotypes under both in vitro tachyzoite and in vitro bradyzoite-inducing alkaline stress culture conditions. Strikingly, the differences in transcriptional profiles between the strains, RH, PLK, and CTG, is much greater than differences between tachyzoites and alkaline stressed in vitro bradyzoites. With an FDR of 10%, we identified 241 genes differentially expressed between CTG tachyzoites and in vitro bradyzoites, including 5 putative AP2 transcription factors. We also observed a close association between cell cycle regulated genes and differentiation. By Gene Set Enrichment Analysis (GSEA), there are a number of KEGG pathways associated with the in vitro bradyzoite transcriptomes of PLK and CTG, including pyrimidine metabolism and DNA replication. These functions are likely associated with cell-cycle arrest. When comparing mRNA levels between strains, we identified 1,526 genes that were differentially expressed regardless of culture-condition as well as 846 differentially expressed only in bradyzoites and 542 differentially expressed only in tachyzoites between at least two strains. Using GSEA, we identified that ribosomal proteins were expressed at significantly higher levels in the CTG strain than in either the RH or PLK strains. This association holds true regardless of life cycle stage.

No MeSH data available.


Related in: MedlinePlus