Transcription profiling of yeast treated with rapamycin
Ontology highlight
ABSTRACT:
ORGANISM(S): Saccharomyces cerevisiae
SUBMITTER: W Ansorge
PROVIDER: E-MANP-2 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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Project description:Target of rapamycin complex 1 (TORC1) is implicated in growth control and aging from yeast to humans. Fission yeast is emerging as a popular model organism to study TOR signaling, although rapamycin has been thought to not affect cell growth in this organism. Here we analyzed the effects of rapamycin and caffeine, singly and combined, on multiple cellular processes in fission yeast. The two drugs led to diverse and specific phenotypes that depended on TORC1 signaling pathway inhibition, including prolonged chronological lifespan, inhibition of global translation, inhibition of cell growth and division, and reprogramming of global gene expression mimicking nitrogen starvation. Rapamycin and caffeine differentially affected these various TORC1-dependent processes. Combined drug treatment augmented most phenotypes and effectively blocked cell growth. Although rapamycin showed a much more subtle effect on global translation than did caffeine, rapamycin was more effective in prolonging chronological lifespan. Rapamycin prolonged the lifespan of non-growing cells only when applied during the growth phase but not when applied after cells had stopped proliferation. The doses of rapamycin and caffeine strongly correlated with growth inhibition and with lifespan extension. This comprehensive analysis will inform future studies into TORC1 function and cellular aging in fission yeast and beyond.
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Project description:To comparatively analyze the transcriptional responses of yeast cells to the presence of rapamycin or caffeine the homozygous hoΔ/hoΔ strain of S. cerevisiae BY4743 were grown in fully controlled fermenters in the presence of 200 nM rapamycin or 5 mM caffeine. Cells were cultured overnight in YPD medium (2% [w/v] D-glucose, 2% [w/v] peptone, 1% [w/v] yeast extract) at 30°C in an orbital shaker at 180 rpm prior to the fermentations. For the cultivations in the fermenters defined synthetic medium was used. The batch cultivations were carried out in duplicates in 2L B-Braun Biostat B Plus fermenters with 1.5L working volume kept at 30°C with the rate of agitation at 800rpm. pH was controlled at 5.5 with 0.5M NaOH and HCl. Sampling was carried out at the mid-exponential phase of growth at an OD range of 0.6-0.8. Samples harvested for the transcriptome analysis were immediately frozen in liquid nitrogen and were stored at -80oC until RNA isolation.
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Project description:We are interested in the study of the already reported sensitivity of yeast cells lacking the type 1 protein phosphatase Ptc1 to the drug rapamycin. In order to carry out our studies, we analyzed the changes in the transcription profile that a short-term incubation with rapamycin (200 ng/mL) has in both, wild type and ptc1 cells. It has been shown that inhibition of the TOR pathway by treatment with rapamycin has profound effects on the global transcriptional profile. We confirmed the previously described transcription changes induced by the drug in wild type cells. Under our working conditions rapamycin increased, in wild type cells, at least 2-fold the expression of 667 genes, whereas it decreased the mRNA level of 721 genes (13.8% and 14.9% of genes with measurable expression level, respectively). Gene ontology analysis shows that, as previously documented, many induced genes falls into the NCR, Msn2/Msn4 regulated stress response or Retrograde response categories, whereas genes encoding cytoplasmic, but not mitochondrial, ribosomal proteins and the so called RIBI regulon where largely repressed. Deletion of PTC1 decreases the number of genes induced or repressed at least 2-fold by rapamycin treatment. Remarkably, lack of Ptc1 seems to lead to a general attenuation of changes triggered by rapamycin. The wt and the ptc1 mutant strains were analyzed in this series. We compared the expression profile of each strain treated with Rapamycin (200 ng/ml for 1h) with that of the same strain mock-treated (90% ethanol, 10% Tween-20). A Dye-swap was carried out for each RNA sample. Total number of chips analyzed: 4
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Project description:The first aim was to identify genes whose transcription is induced by rapamycin feeding in Drosophila larvae. Secondly, the goal was to find out which contribution the transcription factor REPTOR (=CG13624) has to the observed changes in expression. We thus compared gene epxression between rapamycin fed and control fed larvae in wild type larvae and in REPTOR KO larvae. 3 biological replicates from 4 conditions: control larvae plus/miuns rapamycin, KO larvae plus/minus rapamycin; overall 12 samples
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