Yeast heterozygous FHL1 deletion mutant
Ontology highlight
ABSTRACT:
ORGANISM(S): Saccharomyces cerevisiae
PROVIDER: GSE95591 | GEO | 2017/03/02
SECONDARY ACCESSION(S): PRJNA377576
REPOSITORIES: GEO
Similar Datasets
Project description:Affymetrix experiment performed on RNA isolated from Wild type, FHL1 deleted and FHL1,IFH1 double deleted strains. Data aquired in duplicate. In S. cerevisiae the mRNAs from the 138 ribosomal protein (RP) genes are amongst the most abundant in the cell, and their transcription is regulated tightly so that they are the most prominent cluster in most transcriptome experiments. It has recently been observed that the proteins Fhl1p and Ifh1p are found almost exclusively at RP genes (Lee et al., Science, 298, 799-804, 2002;Jorgensen et al., Genes Dev, 18, 2491-2505 2004; Schawalder et al Nature in press; Rudra et al , EMBO J. submitted), and data suggests they are the true transcription factors. This experiment utilizes Affymetrix arrays to measure the level of all mRNAs in cells with a deletion of FHL1 or of both FHL1 and IFH1, compared to the wild type strain, W303. Such mutant cells are viable but grow very slowly (Hermann-Le Denmat S. et al., Mol Cell Biol, 14, 2905-2913, 1994; Cherel and Thuriaux, Yeast, 11, 261-270, 1995). Total RNA was made from log phase cells, amplified according to conventional methods, and hybridized to the array. (Since the total RNA/cell of the mutants is only a fifth of the wt, each mutant required five times the number of cells to provide the same amount of RNA.) The experiment was carried out in duplicate and the six tables provide the raw data from the wt, W303, delFHL1, and delFHL1, delIFH1 strains. Although we expected that the RP genes would be substantially reduced in these mutant cells, they are only slightly less abundant than normal in comparison to all other mRNAs! The interesting result is that the cells respond to a severe deficiency of ribosomes by reducing the level of nearly all mRNAs to match the capacity of the translational apparatus. Keywords = Ribosome Keywords = FHL1 Keywords = IFH1. Keywords: parallel sample
2004-12-22 | GSE2096 | GEO
Project description:Fhl1-9myc ChIP-chip, YPD, OD600=0.8, 2 arrays with duplicate spotting of yeast intergenic regions. AND Ifh1-9myc ChIP-chip, cells grown in YPD, OD600=0.8, 2 arrays with duplicate spotting of yeast intergenic regions. Keywords = Fhl1 Keywords: other
2004-11-09 | GSE1930 | GEO
Project description:Fhl1-9myc ChIP-chip, YPD, OD600=0.8, 2 arrays with duplicate spotting of yeast intergenic regions. AND Ifh1-9myc ChIP-chip, cells grown in YPD, OD600=0.8, 2 arrays with duplicate spotting of yeast intergenic regions. Keywords = Fhl1
2010-06-05 | E-GEOD-1930 | biostudies-arrayexpress
Project description:Affymetrix experiment performed on RNA isolated from Wild type, FHL1 deleted and FHL1,IFH1 double deleted strains. Data aquired in duplicate.,In S. cerevisiae the mRNAs from the 138 ribosomal protein (RP) genes are amongst the most abundant in the cell, and their transcription is regulated tightly so that they are the most prominent cluster in most transcriptome experiments. It has recently been observed that the proteins Fhl1p and Ifh1p are found almost exclusively at RP genes (Lee et al., Science, 298, 799-804, 2002;Jorgensen et al., Genes Dev, 18, 2491-2505 2004; Schawalder et al Nature in press; Rudra et al , EMBO J. submitted), and data suggests they are the true transcription factors.,This experiment utilizes Affymetrix arrays to measure the level of all mRNAs in cells with a deletion of FHL1 or of both FHL1 and IFH1, compared to the wild type strain, W303. Such mutant cells are viable but grow very slowly (Hermann-Le Denmat S. et al., Mol Cell Biol, 14, 2905-2913, 1994; Cherel and Thuriaux, Yeast, 11, 261-270, 1995). Total RNA was made from log phase cells, amplified according to conventional methods, and hybridized to the array. (Since the total RNA/cell of the mutants is only a fifth of the wt, each mutant required five times the number of cells to provide the same amount of RNA.) The experiment was carried out in duplicate and the six tables provide the raw data from the wt, W303, delFHL1, and delFHL1, delIFH1 strains.,Although we expected that the RP genes would be substantially reduced in these mutant cells, they are only slightly less abundant than normal in comparison to all other mRNAs! The interesting result is that the cells respond to a severe deficiency of ribosomes by reducing the level of nearly all mRNAs to match the capacity of the translational apparatus.
2007-07-07 | E-GEOD-2096 | biostudies-arrayexpress
Project description:Misincorporation of ribonucleotides into DNA during genome replication has recently become recognized as a significant source of genomic instability. The frequency of ribonucleotides in the genome is determined by dNTP/rNTP ratios, the ability of the DNA polymerases to discriminate against ribonucleotides, and by the capacity of repair mechanisms to remove misincorporated ribonucleotides. To simultaneously compare how the nuclear and mitochondrial genomes incorporate and remove ribonucleotides, we challenged these processes by imbalancing cellular dNTP pools. Using a collection of yeast strains with altered dNTP pools, we discovered an inverse relationship between the concentration of individual dNTPs and the amount of the corresponding ribonucleotides incorporated in mitochondrial DNA, while in nuclear DNA the ribonucleotide pattern was only altered in the absence of ribonucleotide excision repair. Our analysis uncovers major differences in ribonucleotide repair between the two genomes and provides concrete evidence that yeast mitochondria lack mechanisms for repair of misincorporated ribonucleotides.
2017-11-07 | GSE100352 | GEO
Project description:Replication stress is a common feature of cancer cells, and thus a potentially important therapeutic target. Here we show that CDK-induced replication stress is synthetic lethal with mutations disrupting dNTP homeostasis in fission yeast. Wee1 inactivation leads to increased dNTP demand and replication stress through CDK-induced firing of dormant replication origins. Subsequent dNTP depletion leads to inefficient DNA replication, Mus81- dependent DNA damage, and to genome instability. Cells respond to this replication stress by increasing dNTP supply through Set2-dependent MBFinduced expression of Cdc22, the catalytic subunit of ribonucleotide reductase (RNR). Disrupting dNTP synthesis following Wee1 inactivation, through loss of Set2-dependent H3K36 tri-methylation, results in further dNTP depletion due to reduced RNR expression, leading to replication catastrophe and cell death. This lethality can be rescued by increasing dNTP levels. Similarly, we find loss of the DNA integrity checkpoint results in synthetic lethality with Wee1 inactivation, which is also associated with replication collapse through critically low dNTP levels. Together, these findings support a ‘dNTP supply and demand’ model in which maintaining dNTP homeostasis is essential in preventing replication catastrophe in response to CDK-induced replication stress.
2018-05-26 | GSE113747 | GEO
Project description:Intracellular levels of deoxyribonucleoside triphosphate (dNTP) must be tightly regulated to preserve genome integrity. Indeed, alterations in dNTP pools have recently been associated with increased mutagenesis, genomic instability and tumorigenesis. However, the mechanisms by which low or imbalanced dNTP pools affect DNA replication remain poorly understood. Here, we have modulated the activity of ribonucleotide reductase (RNR), a key enzyme catalyzing a rate-limiting step of dNTP production, to monitor the effect of altered dNTP levels on replication dynamics in budding yeast. We show that dNTP pools are limiting for normal DNA synthesis as upregulation of RNR activity increases replication fork speed. In contrast, inhibition of RNR activity with hydroxyurea (HU) induces a sharp transition from a regular- to a slow-replication mode within minutes after S-phase entry. Interestingly, we found that upregulation of RNR activity delays this transition and that dNTP levels modulate both fork speed and origin usage under replication stress. Moreover, we report that chromosomal instability (CIN) mutants show increased dNTP pools and enhanced DNA synthesis in the presence of HU. Since upregulation of RNR allows forks to progress faster in the presence of DNA lesions, we propose that CIN mutants adapt to chronic replication stress by upregulating dNTP pools.
2012-01-01 | GSE33686 | GEO
Project description:Array design -Platform: amino-silane coated glass slides (GAPS II, Corning) -S. cerevisiae intergenic regions amplified from S288C genomic DNA (ResGen) using the intergenic region primer oligonucleotides (ResGen) (Harismendy et al. EMBO J. 22(18): 4738-4747, 2003). The primers allow the amplification of the sequence located on either side of elements such as open reading frames, tRNAs, small nuclear RNAs, Ty elements, solo δ, etc. A complete description of the primers can be obtained on the web site (ftp://ftp.resgen.com/pub/genepairs/yeast_intergenic). -The PCR products were purified and spotted using an automated arrayer (MicroGrid II, BioRobotics) Experiment design -Type of experiment: genome wide binding analysis -Experimental factor: genotype (Wild-Type, Fhl1-myc, Ifh1-myc); drug (rapamycin) -Number of hybridization performed in the experiment: 15 (four conditions with three to five biological replicates each) -Reference used for each condition: ChIP from the wild-type strain -Quality control: one dye-swap per condition Samples used, extract preparation and labeling -The origin of the biological sample: o Species: S. cerevisiae o Cell type: (1) W303; (2) W303+IFH1-13myc-TRP1MX6;(3) W303+FHL1-13myc-HIS3MX6. -Growth conditions: log phase (2x107/mL) at 30ºC in YPD medium; when indicated rapamycin was added at the final concentration of 200 ng/mL for 90 min. -Cross-linking for 30 min at 30ºC -Chromatin immunoprecipitation: Dynabeads M280 coupled to sheep anti-mouse IgG or sheep anti-rabbit IgG and mouse monoclonal antibodies against the myc epitope (culture supernatant of 9E10) were used for immunoprecipitation according to standard techniques. -Labeling protocol(s): each IP was blunt-ended, ligated to linker DNA, then subjected to Ligation Mediated (LM)-PCR in the presence of amino-allyl dUTP. Labeling was performed for 1 hour at room temperature in the presence of NHS-ester Cy3 or Cy5 in 0.1 M Sodium Carbonate. The labeling reaction was stopped with 2 M Hydroxylamine and the labeled DNA purified. Hybridization procedures and parameters -Standard protocols were used Measurement data and specifications -Software for scanning: GenePix Pro 4.0 (Axon Instruments, Inc.) -Scanner: GenePix 4000A (Axon Instruments, Inc.) Keywords: dose response
2004-11-10 | GSE1944 | GEO
Project description:Yeast heterozygous FHL1 deletion mutant
| PRJNA377576 | ENA
Project description:Array design -Platform: amino-silane coated glass slides (GAPS II, Corning) -S. cerevisiae intergenic regions amplified from S288C genomic DNA (ResGen) using the intergenic region primer oligonucleotides (ResGen) (Harismendy et al. EMBO J. 22(18): 4738-4747, 2003). The primers allow the amplification of the sequence located on either side of elements such as open reading frames, tRNAs, small nuclear RNAs, Ty elements, solo δ, etc. A complete description of the primers can be obtained on the web site (ftp://ftp.resgen.com/pub/genepairs/yeast_intergenic). -The PCR products were purified and spotted using an automated arrayer (MicroGrid II, BioRobotics) Experiment design -Type of experiment: genome wide binding analysis -Experimental factor: genotype (Wild-Type, Fhl1-myc, Ifh1-myc); drug (rapamycin) -Number of hybridization performed in the experiment: 15 (four conditions with three to five biological replicates each) -Reference used for each condition: ChIP from the wild-type strain -Quality control: one dye-swap per condition Samples used, extract preparation and labeling -The origin of the biological sample: o Species: S. cerevisiae o Cell type: (1) W303; (2) W303+IFH1-13myc-TRP1MX6;(3) W303+FHL1-13myc-HIS3MX6. -Growth conditions: log phase (2x107/mL) at 30ºC in YPD medium; when indicated rapamycin was added at the final concentration of 200 ng/mL for 90 min. -Cross-linking for 30 min at 30ºC -Chromatin immunoprecipitation: Dynabeads M280 coupled to sheep anti-mouse IgG or sheep anti-rabbit IgG and mouse monoclonal antibodies against the myc epitope (culture supernatant of 9E10) were used for immunoprecipitation according to standard techniques. -Labeling protocol(s): each IP was blunt-ended, ligated to linker DNA, then subjected to Ligation Mediated (LM)-PCR in the presence of amino-allyl dUTP. Labeling was performed for 1 hour at room temperature in the presence of NHS-ester Cy3 or Cy5 in 0.1 M Sodium Carbonate. The labeling reaction was stopped with 2 M Hydroxylamine and the labeled DNA purified. Hybridization procedures and parameters -Standard protocols were used Measurement data and specifications -Software for scanning: GenePix Pro 4.0 (Axon Instruments, Inc.) -Scanner: GenePix 4000A (Axon Instruments, Inc.) Keywords: dose response
2010-06-05 | E-GEOD-1944 | biostudies-arrayexpress