Toxoplasma gondii response to monensin
Ontology highlight
ABSTRACT:
ORGANISM(S): Toxoplasma gondii
SUBMITTER: Mark Lavine
PROVIDER: E-GEOD-25177 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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Project description:PRMT1 is thought to be responsible for the majority of PRMT activity in Toxoplasma gondii, but its exact function is unknown. We generated T. gondii mutants lacking PRMT1 (∆prmt1) by deletion of the PRMT1 gene. ∆prmt1 parasites exhibit morphological defects during cell division and grow slowly, and this phenotype reverses in the complemented strain ∆prmt::PRMT1mRFP. PRMT1 localizes primarily in the cytoplasm with enrichment at the centrosome, and the strain lacking PRMT1 is unable to segregate progeny accurately. Unlike wild-type and complemented parasites, ∆prmt1 parasites have abnormal daughter buds, perturbed centrosome stoichiometry, and loss of synchronous replication. Whole genome expression profiling demonstrated differences in expression of cell cycle regulated genes in ∆prmt1 relative to the complemented ∆prmt1::PRMT1mRFP and parental wild-type strains, but these changes did not correlate with a specific block in cell cycle. Although PRMT1’s primary biological function was previously proposed to be methylation of histones, our genetic studies suggest that the most critical function of PRMT1 is within the centrosome as a regulator of daughter cell counting to assure the proper replication of the parasite. RNA samples were isolated in triplicates from RH-hxgprt parent strain (W), PRMT1 knockout (K) strain and PRMT1 knockout strain complemented with RFP-tagged PRMT1 protein (C). Parasites were grown for 32h at 37C. Samples were hybridized to the Toxoplasma gondii Affymetrix microarray (ToxoGeneChip: http://ancillary.toxodb.org/docs/Array-Tutorial.html). Hybridization data was preprocessed with Robust Multi-array Average (RMA) and normalized using per chip and per gene median polishing and analyzed using the software package GeneSpring GX (Agilent Technologies).
2015-09-18 | E-GEOD-73177 | biostudies-arrayexpress
Project description:Toxoplasma gondii temperature sensitive mutant 12-109C6 conditionally arrests in the G1 phase due to a single point mutation in a novel protein containing a single RNA-recognition-motif (TgRRM1). The resulting tyrosine to asparagine amino acid change in TgRRM1 causes severe temperature instability that generates an effective null phenotype for this protein when the mutant is shifted to the restrictive temperature. Orthologs of TgRRM1 are widely conserved in diverse eukaryote lineages, and the human counterpart (RBM42) can functionally replace the missing Toxoplasma factor. The interaction of TgRRM1 with factors of the tri-SNP complex (U4/U6 & U5 snRNPs) indicate this factor may be required to assemble an active spliceosome. Thus, the TgRRM1 family of proteins is an unrecognized and evolutionarily conserved class of splicing regulators. We describe transcriptome of the temperature sensitive mutant 12-109C6. Transcriptome studies demonstrated that gene expression is largely downregulated in the mutant at the restrictive temperature 40oC and is accompanied by a severe defect in splicing that affects both cell cycle and constitutively expressed mRNAs. RNA samples were isolated in duplicate from mutant 12-109C6 parasites grown for 24h at permissive (34oC) and 6h and 24h at non-permissive temperatures (40oC). Samples were hybridized to the Toxoplasma gondii Affymetrix microarray (ToxoGeneChip: http://ancillary.toxodb.org/docs/Array-Tutorial.html). Hybridization data was preprocessed with Robust Multi-array Average (RMA) and normalized using per chip and per gene median polishing and analyzed using the software package GeneSpring GX (Agilent Technologies, Santa Clara CA).
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Project description:Toxoplasma gondii cell cycle mutant 11-104A4 reversibly arrests in the G1 phase. The defect in this mutant was mapped by genetic complementation to a gene encoding a novel AAA-ATPase/CDC48 family member (TgNoAP1). A change in a tyrosine to a cysteine upstream of an AAA+ domain leads to protein instability and results in cell cycle arrest. This factor is cell cycle regulated and exclusively expressed in the nucleolus during the G1/S phases. At high temperature the mutant quickly arrests with a single nucleus and expresses transcripts enriched in the G1 subtranscriptome. This unique CDC48 ortholog operates in a parasite mechanism responsible for G1 progression and is the first G1-specific checkpoint factor described in Toxoplasma. We describe transcriptome of the temperature sensitive mutant 11-104A4. Transcriptome studies demonstrated that gene expression is reflective of the parasite arrest in G1 phase. mRNAs normally upregulated in S/M phase were largely downregulated in the mutant at the restrictive temperature 40oC. Genetic complementation with extra copy of the wild type TgNoAP1 rescued growth of the mutant 11-104A4 at 40oC and reversed changes in the transcriptome. RNA samples were isolated in duplicate from mutant 11-104A4 parasites grown for 32h at permissive (34oC) or non-permissive temperatures (40oC). In addition, a duplicate sample of RNA from mutant 11-104A4 complemented with cosmid TOXOV53 encoding wild type copy of TgNoAP1 similarly grown at 40oC was collected. Samples were hybridized to the Toxoplasma gondii Affymetrix microarray (ToxoGeneChip: http://ancillary.toxodb.org/docs/Array-Tutorial.html). Hybridization data was preprocessed with Robust Multi-array Average (RMA) and normalized using per chip and per gene median polishing and analyzed using the software package GeneSpring GX (Agilent Technologies).
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2013-02-06 | E-GEOD-38779 | biostudies-arrayexpress
Project description:Parasitic protozoa such as the apicomplexan Toxoplasma gondii progress through their life cycle in response to stimuli in the environment or host organism. Very little is known about how proliferating tachyzoites reprogram their expressed genome in response to stresses that prompt development into latent bradyzoite cysts. We have previously linked histone acetylation with the expression of stage-specific genes, but the factors involved remain to be determined. We sought to determine if GCN5, which operates as a transcriptional co-activator by virtue of its histone acetyltransferase (HAT) activity, contributed to stress-induced changes in gene expression in Toxoplasma. In contrast to other lower eukaryotes, Toxoplasma has duplicated its GCN5 lysine acetyltransferase (KAT). Disruption of the gene encoding for TgGCN5-A did not produce a severe phenotype under normal culture conditions, but here we show that the TgGCN5-A null mutant is deficient in recovering from alkaline pH, a common stress used to induce bradyzoite differentiation in vitro. The TgGCN5-A knockout is incapable of up-regulating key marker genes expressed during development of the latent cyst form. Complementation of the TgGCN5-A knockout restores the expression of these stress-induced genes and reverses the stress recovery defect. We also describe a genome-wide analysis of the Toxoplasma transcriptional response to alkaline pH stress, finding that TgGCN5-A knockout parasites fail to up-regulate 68% of the stress response genes that are induced 2-fold or more in wild-type. Using chromatin immunoprecipitation, we verify an enrichment of TgGCN5-A at the upstream regions of genes activated by alkaline pH exposure, including developmentally regulated genes. Wild-type and GCN5-A- Knockout organisms were subjected to control and stress treatments
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Project description:This SuperSeries is composed of the SubSeries listed below. Refer to individual Series
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