Project description:Combinatorial promoter expression level estimation via cell sorting The purpose of this experiment was to determine the expression level of a library of synthetic promoters. The promoters were cloned in front of a GFP reporter and the resulting library transformed into yeast, sorted by FACS into six fluorescence bins, and the contents of the bins sequenced to determine the distribution of each promoter among each fluorescence bin. This was then used to calculate an expression level for each promoter with enough data.
Project description:We analyzed genome-wide transcriptional profiles of Saccharomyces cerevisiae BY4742 strain in response to BPA, focusing on two exposure scenarios: (i) low-observed-effect concentration (<10% inhibition) to examine chronic effect of BPA on yeast population, and (ii) high-inhibitory concentration (>70% inhibition) to study acute effect. Initially, yeast cells were exposed to various concentrations of BPA. 50 mg/L and 300 mg/L BPA were determined as low-observed-effect concentration and the high-inhibitory concentration, respectively. Transcriptional profiles indicated that 81 genes were repressed and 104 genes were induced in response to 50 mg/L BPA. On the other hand, in 300 mg/L BPA exposure, 378 genes were down-regulated, while 606 genes were significantly up-regulated. Our data showed that there were similar processes affected by both concentrations such as mitochondria, nucleobase-containing small molecule metabolic process, transcription from RNA polymerase II promoter, and mitotic cell cycle and associated processes. However, different modes of actions of the BPA were found between two concentrations. 300 mg/L BPA exposure showed severe effects on the processes by repressing or inducing several genes or total mechanisms with high level of expression changes, while 50 mg/L BPA exposure changed the expression of some important genes with low level of expression changes in the processes. These results suggest that yeast cells respond via different ways to the different concentrations of BPA at transcriptomic level.
Project description:This study focus a comparative toxicogenomic analysis of the effects of four herbicides (alachlor, ALA, S-metolachlor, S-MET, diuron, DIU, and MCPA-methyl ester, MCPA-ME), one insecticide (carbofuran, CAB), and one fungicide (pyrimethanil, PYR), in the model yeast Saccharomyces cerevisiae, to predict potential cytotoxic effects of these xenobiotics while providing mechanistic clues possibly relevant for experimentally less accessible non-target eukaryotes. The six model pesticides selected have been used worldwide in agricultural activities, at the present time or in the past, and have different modes of action on their target-organisms. Moreover, some of them are currently in Annex 1 of EC Directive 1107/2009 (repealing 91/414), that is they are in use in the EU, but having some ecotoxicological concerns (e.g. S-MET, PYR, MCPA-ME), others have their use restricted and/or are priority substances under the Water Framework Directive (e.g. ALA, DIU), and one was banned (e.g. CAB). We used genome microarrays to obtain gene expression profiles in S. cerevisiae strain BY4741 upon 2 hours exposure to pesticide concentrations exerting a moderate inhibition of yeast growth rate. To obtain comparable gene expression profiles representing yeast response to the six different pesticides, all exposure treatments were performed with equitoxic pesticide concentrations that generated equivalent responses at the phenotypic level (namely, 20% growth-inhibitory concentrations, hereafter designated as the pesticide IC20).
Project description:Yeast cells were treated with different doses of pheromone in order to obtain dose-response profiles for induced genes. Specifically, dose-dependencies of transcriptional feedback regulators of the pheromone-response pathway were quantified.