Conserved and diverged functions of the calcineurin-activated Prz1 transcription factor in fission yeast
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ABSTRACT:
ORGANISM(S): Schizosaccharomyces pombe
SUBMITTER: Gordon Chua
PROVIDER: E-GEOD-77761 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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Project description:Gene regulation in response to intracellular calcium is mediated by the calcineurin-activated transcription factor Prz1 in the fission yeast Schizosaccharomyces pombe. Genome-wide studies of the Crz1 and CrzA fungal orthologs have uncovered numerous target genes involved in conserved and species-specific cellular processes. In contrast, very few target genes of Prz1 have been published. This paper identified an extensive list of genes using transcriptome and ChIP-chip analyses under inducing conditions of Prz1, including CaCl2, and tunicamycin treatment, as well as a âpmr1 genetic background. We identified 165 upregulated putative target genes of Prz1 in which the majority contained a calcium-dependent response element in their promoters, similar to that of the Saccharomyces cerevisiae ortholog Crz1. These genes were functionally enriched for Crz1-conserved processes such as cell wall biosynthesis. Overexpression of prz1+ increased resistance to the cell wall degradation enzyme zymolyase, likely from upregulation of the O-mannosyltransferase encoding gene omh1+. Loss of omh1+ abrogates this phenotype. We uncovered a novel inhibitory role in flocculation for Prz1. Loss of prz1+ resulted in constitutive flocculation and upregulation of genes encoding the flocculins Gsf2 and Pfl3, as well as the transcription factor Cbf12. The constitutive flocculation of the âprz1 strain was abrogated by the loss of gsf2+ or cbf12+. This study reveals that Prz1 functions as a positive and negative transcriptional regulator of genes involved in cell wall biosynthesis and flocculation, respectively. Moreover, comparison of target genes between Crz1/CrzA and Prz1 indicate some conservation in DNA-binding specificity, but also substantial rewiring of the calcineurin-mediated transcriptional-regulatory network. We generated 3 overexpression microarrays and 2 deletion microarrays with dye swaps, and 2 deletion microarrays with drug treatments with dye swaps, each with a biological replicate performed as a dye swap. The effect of the mutant strains were all compared to wild type or empty vector strains, except for one experiment where the prz1 deletion mutant was compared to the pmr1 deletion mutant.
2016-02-10 | E-GEOD-77759 | biostudies-arrayexpress
Project description:Gene regulation in response to intracellular calcium is mediated by the calcineurin-activated transcription factor Prz1 in the fission yeast Schizosaccharomyces pombe. Genome-wide studies of the Crz1 and CrzA fungal orthologs have uncovered numerous target genes involved in conserved and species-specific cellular processes. In contrast, very few target genes of Prz1 have been published. This paper identified an extensive list of genes using transcriptome and ChIP-chip analyses under inducing conditions of Prz1, including CaCl2, and tunicamycin treatment, as well as a â??pmr1 genetic background. We identified 165 upregulated putative target genes of Prz1 in which the majority contained a calcium-dependent response element in their promoters, similar to that of the Saccharomyces cerevisiae ortholog Crz1. These genes were functionally enriched for Crz1-conserved processes such as cell wall biosynthesis. Overexpression of prz1+ increased resistance to the cell wall degradation enzyme zymolyase, likely from upregulation of the O-mannosyltransferase encoding gene omh1+. Loss of omh1+ abrogates this phenotype. We uncovered a novel inhibitory role in flocculation for Prz1. Loss of prz1+ resulted in constitutive flocculation and upregulation of genes encoding the flocculins Gsf2 and Pfl3, as well as the transcription factor Cbf12. The constitutive flocculation of the â??prz1 strain was abrogated by the loss of gsf2+ or cbf12+. This study reveals that Prz1 functions as a positive and negative transcriptional regulator of genes involved in cell wall biosynthesis and flocculation, respectively. Moreover, comparison of target genes between Crz1/CrzA and Prz1 indicate some conservation in DNA-binding specificity, but also substantial rewiring of the calcineurin-mediated transcriptional-regulatory network. We generated 2 ChIP-chip experiments, each with a biological replicate performed as a dye swap.
2016-02-10 | E-GEOD-77760 | biostudies-arrayexpress
Project description:In Schizosaccharomyces pombe, over 90% of transcription factor genes are nonessential. Moreover, the majority do not exhibit significant growth defects under optimal conditions when deleted, complicating their functional characterization and target gene identification. Here, we systematically overexpressed 99 transcription factor genes with the nmt1 promoter. Screening the overexpression array revealed that 64 transcription factor genes exhibited reduced fitness when ectopically expressed. Cell cycle defects were also often observed. We further investigated three uncharacterized transcription factor genes (toe1+-toe3+) which displayed cell elongation when overexpressed. Ectopic expression of toe1+ resulted in a G1 delay while toe2+ and toe3+ overexpression produced an accumulation of septated cells with abnormalities in septum formation and nuclear segregation, respectively. Transcriptome profiling and ChIP-chip analysis of the transcription factor overexpression strains indicated that Toe1 activates target genes of the pyrimidine-salvage pathway, while Toe3 regulates target genes involved in polyamine synthesis. We also found that ectopic expression of the putative target genes SPBC3H7.05c, and dad5+ and SPAC11D3.06 could recapitulate the cell cycle phenotypes of toe2+ and toe3+ overexpression, respectively. Furthermore, the phenotypes of toe1+and toe2+ overexpression could be suppressed by deletion of the putative target genes urg2+ and SPAC1399.04c, and SPBC3H7.05c, SPACUNK4.15 and rds1+, respectively. This study implicates new transcription factors and metabolism genes in cell cycle regulation and demonstrates the potential of systematic overexpression analysis to elucidate the function and target genes of transcription factors in S. pombe. We generated 3 overexpression microarrays and 2 deletion microarray with dye swaps, 3 deletion microarrays with drug treatments with dye swaps, 1 deletion single replicate microarray experiment, and 3 single replicate chIP-chip experiments. The effect of the mutant strains were all compared to wild type or empyty vector strains except for two experiments one that looked at the toe1 mutant with and without chlorpromazine and another that looked at wild type with and without chlorpromazine.
2013-05-11 | E-GEOD-46811 | biostudies-arrayexpress
Project description:The regulation of flocculation, surface adhesion and invasive growth in the fission yeast Schizosaccharomyces pombe has focused primarily at the transcriptional level, but little is known with regards to posttranscriptional control. Here, we identified the Pumilio protein Pfr1 as a novel posttranscriptional regulator of these processes. Deletion of pfr1+ prevented flocculation, surface adhesion and invasive growth under inducing conditions, while overexpression of pfr1+ was sufficient to trigger flocculation. The flocculent phenotype of pfr1+ overexpression was dependent on the presence of the Gsf2 flocculin, but not on the Mbx2, Cbf12 and Adn3 transcription factors. In addition, we used RNA immunoprecipitation and expression microarrays to identify pvg1+ and SPBPB7E8.01, which encode a galactose pyruvyltransferase and glycophosphatidylinositol membrane protein, respectively, as putative mRNA targets potentially degraded by Pfr1. The mRNAs of these genes were upregulated and downregulated in the pfr1 deletion and overexpression strains, respectively, and contained putative binding sites in the 3â-untranslated region. We also discovered that ccr4+ and ste13+, which encode components of the mRNA decay machinery, were required for these processes, but did not suppress the pfr1+ overexpression flocculent phenotype when deleted. This data suggest that these processes in S. pombe involve multiple posttranscriptional-regulatory pathways of which one requires Pfr1. We generated 2 overexpression microarrays with dye swap that were biological replicates, 2 deletion microarrys with dye swap. Mutants samples were compared to empty vector control or wild type. 1 RIP-chip array was generated with IP rna compared to total RNA from the sample.
2016-03-24 | E-GEOD-79537 | biostudies-arrayexpress
Project description:The choice of growth media is a very important consideration of any cell-based proteomics experiment. Alterations thereof may result in differences in basal proteomes simply due to disparities in the metabolite composition of the media. We investigate the effect of growth media on the proteomes of three microorganisms, specifically E. coli, S. cerevisiae, and S. pombe, using tandem mass tag (TMT)-based quantitative proteomics. We compared the protein abundance profiles of these microorganisms propagated in two distinct growth media that are commonly used for the respective organism. Our sample preparation strategy included SP3 bead-assisted protein isolation and digestion. In addition, we assembled a replicate set of samples in which we altered the proteolytic digestion from sequential treatment with LysC and trypsin to only LysC. Despite differences in peptides identified and a drop in quantified proteins, the results were similar between the two datasets for all three microorganisms. Approximately 10% of the respective microbial proteomes were significantly altered in each dataset. As expected, gene ontology analysis revealed that the majority of differentially expressed proteins are implicated in metabolism. We emphasize further the importance of growth media selection and the potential consequences thereof.
2021-05-11 | PXD020816 | Pride
Project description:Whole genome microarray was used to compare the transcriptional profile of C. parapsilosis growing in normoxic (21% oxygen) and hypoxic (1% oxygen) conditions. RNA was isolated from cells grown in SD media at 37°C for 3 h in atmosphere oxygen of 1% oxygen, and labeled with Cy5 or Cy3. Six independent biological replicates were compared. 4 out of 6 hypoxic samples were labeled with Cy5, and 2 were labeled with Cy3.
2010-05-06 | E-GEOD-13722 | biostudies-arrayexpress
Project description:Rheb, a ras-like small GTPase conserved from human to yeast, controls Tor kinase and plays a central role in regulation of cell growth depending on extracellular conditions. Fission yeast Rheb regulates amino acid uptake as well as response to nitrogen starvation. In this study we generated two mutants of Rheb, rhb1-DA4 and rhb1-DA8, and characterized them genetically. V17A mutation within the G1 box defined for the ras-like GTPases was responsible for rhb1-DA4, and Q52R I76F within the switch II domain for rhb1-DA8. In fission yeast, two events, induction of a meiosis initiating gene mei2+ and cell division without cell growth, are a typical response to nitrogen starvation. Under nitrogen-rich conditions, Rheb stimulates Tor kinase, which, in turn, suppresses the response to nitrogen starvation. While amino acid uptake was prevented by both rhb1-DA4 and rhb1-DA8 in a dominant fashion, the response to nitrogen starvation was prevented only by rhb1-DA4. rhb1-DA8 thereby allowed genetic dissection of the Rheb-dependent signaling cascade. We postulate that Rheb in fission may have two downstream elements, Tor kinase for regulation of the response to nitrogen starvation and the other element for regulation of amino acid uptake. Gene expression profile under nitrogen starvation in fission yeast. Type of experiment: Comparing between vegetatively-growing control cells and cells 3 hrs after nitrogen starvation. Experimental factor: Gene expression profile after nitrogen starvation in the rhb1-D4 or rhb1-D8 cells. Quality control steps taken: All experiments were repeated more than twice except for the tsc2D which was previously reported. Keywards: Nitrogen starvation, rhb1-D4, rhb1-D8
2009-09-28 | E-GEOD-14626 | biostudies-arrayexpress
Project description:RNA interference (RNAi) is a gene silencing mechanism conserved from fungi to mammals. Small interfering RNAs are products and mediators of the RNAi pathway and act as specificity factors in recruiting effector complexes. The Schizosaccharomyces pombe genome encodes one of each of the core RNAi proteins, Dicer, Argonaute and RNA-dependent RNA polymerase (dcr1, ago1, rdp1). Even though the function of RNAi in heterochromatin assembly in S. pombe is established, its role in controlling gene expression is elusive. Here, we report the identification of small RNAs mapped anti-sense to protein coding genes in fission yeast. We demonstrate that these genes are up-regulated at the protein level in RNAi mutants, while their mRNA levels are not significantly changed. We show that the repression by RNAi is not a result of heterochromatin formation. Thus, we conclude that RNAi is involved in post-transcriptional gene silencing in S. pombe. Total RNA from wild type, dcr1M-NM-^T and rpd1M-NM-^T cells; 3 biological replicates for each strain.
2014-01-23 | E-GEOD-54190 | biostudies-arrayexpress
Project description:This SuperSeries is composed of the following subset Series: GSE13717: Transcriptional profile of Candida parapsilosis in SD media GSE13722: Transcriptional response of Candida parapsilosis in low oxygen (hypoxic) conditions in SD media Refer to individual Series
2010-05-06 | E-GEOD-13832 | biostudies-arrayexpress
Project description:This experiment aims at analyzing crossover distribution genome-wise, in the fission yeast. S. pombe strains PR109 (h- leu1-32 ura4-D18) and PR110 (h+ leu1-32 ura4-D18) were used for three successive rounds of mutagenesis with Ethylmethane Sulfonate Mutagenesis. Five independent clones of the first round of mutagenesis were at the root of two subsequent similar rounds of mutagenesis. Each clone used was checked for its ability to mate and sporulate. Eventually, five mutagenized clones from each of the PR109 and PR110 backgrounds were sequenced to identify de novo mutations and determine the optimal combinations of mutation patterns for recombination analyses. The following h- x h+ crosses were selected and the corresponding tetrads dissected: BLP49 (A) x BLP23 (I), 29 tetrads; BLP59 (C) x BLP19 (H), 30 tetrads and BLP64 (D) x BLP33 (K), 33 tetrads.
2023-03-03 | E-MTAB-12516 | biostudies-arrayexpress