Unknown,Transcriptomics,Genomics,Proteomics

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Transcription profiling of S. cerevisiae response to heme deficiency and hypoxia

ABSTRACT: We addressed changes in gene expression profile in response to ; reduced oxygen tension in wild type or heme depletion caused by ; deletion of heme biosynthesis gene hem1 Experiment Overall Design: I.Heme depletion.The hem1 delta strain was grown for 6 hrs in YPD media supplemented with 250 uM ALA. Cells were washed three times with water to remove excess ALA. Cells were then inoculated in duplicates into 15 ml cultures of Experiment Overall Design: YPD media that was supplemented either with or without ALA. Cultures were induced for 16 h before 5 ml of cells were harvested and washed Experiment Overall Design: three times in DEPC water. The culture density at the time of harvest Experiment Overall Design: was approximately OD600=0.5. Experiment Overall Design: II.Hypoxia. BY45742 strain was grown for 6 hrs in Experiment Overall Design: YPD medium and then inoculated in 15 ml the same medium and grown Experiment Overall Design: under normoxic or hypoxic conditions for 12 hrs. To create hypoxic Experiment Overall Design: conditions, cultures were incubated in an anaerobic chamber using a Experiment Overall Design: BBL-GasPak (BD Biosciences).

ORGANISM(S): Saccharomyces cerevisiae

SUBMITTER: Olga Protchenko

PROVIDER: E-GEOD-9514 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Sampangine, a plant-derived alkaloid found in the Annonaceae family, exhibits strong inhibitory activity against the opportunistic fungal pathogens Candida albicans, Cryptococcus neoformans and Aspergillus fumigatus. In the present study, transcriptional profiling experiments coupled with the analysis of mutants were performed in an effort to elucidate its mechanism of action. Using Saccharomyces cerevisiae as a model organism, we show that sampangine produces a transcriptional response indicative of hypoxia, altering the expression of genes known to respond to low oxygen conditions. Several additional lines of evidence obtained suggest that these responses could involve effects on heme. First, the hem1 deletion mutant lacking the first enzyme in the heme biosynthetic pathway showed increased sensitivity to sampangine, and exogenously supplied hemin partially rescued the inhibitory activity of sampangine in wild-type cells. In addition, heterozygous mutants with deletions in genes involved in five out of eight steps in the heme biosynthetic pathway showed increased susceptibility to sampangine. Furthermore, spectral analysis of pyridine extracts indicated significant accumulation of free porphyrins in sampangine-treated cells. Transcriptional profiling experiments were also performed in C. albicans to investigate the response of a pathogenic fungal species to sampangine. Taking into account the known differences in the physiological responses of C. albicans and S. cerevisiae to low oxygen, significant correlations were observed between the two transcription profiles suggestive of heme-related defects. Our results indicate that the antifungal activity of the plant alkaloid sampangine is due, at least in part, to perturbations in the biosynthesis or metabolism of heme. Experiment Overall Design: S. cerevisiae S288C cells at OD 0.2 were treated with either sampangine (SMP) at IC50 concentration (1.17 ug/ml), or solvent (0.25% DMSO), allowed to grow to OD 0.5, then harvested and frozen. Three biological replicate samples were analyzed for each treatment.

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Transcription profiling of S. cerevisiae response to heme deficiency and hypoxia

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