Project description:In this study, we mapped for the first time differences in transcription binding among individuals and elucidated the genetic basis of such variation. Whole-genome Ste12 binding profiles were determined using ChIP-Seq in pheromone-treated cells of 43 segregants of a cross between two highly diverged yeast strains, YJM789 and S288c, as well as the parental lines. We identified extensive Ste12 binding variation among individuals and mapped underlying cis- and trans- acting loci responsible for such variation. We showed that the majority of TF binding variation is cis-linked and that many variations are associated with polymorphisms residing in the binding motifs of Ste12 as well as those of several known and proposed Ste12 cofactors. We also identified two trans factors, AMN1 and FLO8, that modulate Ste12 binding to promoters of more than 10 genes under α-factor treatment. Neither of these two genes was known to regulate Ste12 previously, and we suggest that they may be key mediators of gene activity and phenotypic diversity. Ste12 binding strongly correlates with gene expression for more than 200 genes, indicating that binding variation is functional. Many of the variable bound genes are involved in cell wall organization and biogenesis. Overall, we identified key regulators of molecular diversity among individuals and provide novel insights into mechanisms of gene regulation. We measured gene expression levels after 30 minutes treatment with alpha factor for 43 MATa segregants from a YJM789 X S96 cross, as well as MATa parental lines. We also measured the gene expression levels without alpha factor treatment for parental lines (S96, HS959) and SEG8 as controls.
Project description:In this study, we mapped for the first time differences in transcription binding among individuals and elucidated the genetic basis of such variation. Whole-genome Ste12 binding profiles were determined using ChIP-Seq in pheromone-treated cells of 43 segregants of a cross between two highly diverged yeast strains, YJM789 and S288c as well as the parental lines. We identified extensive Ste12 binding variation among individuals and mapped underlying cis- and trans- acting loci responsible for such variation. We showed that the majority of TF binding variation is cis-linked and that many variations are associated with polymorphisms residing in the binding motifs of Ste12 as well as those of several known and proposed Ste12 cofactors. We also identified two trans factors, AMN1 and FLO8, that modulate Ste12 binding to promoters of more than 10 genes under α-factor treatment. Neither of these two genes was known to regulate Ste12 previously, and we suggest that they may be key mediators of gene activity and phenotypic diversity. Ste12 binding strongly correlates with gene expression for more than 200 genes indicating that binding variation is functional. Many of the variable bound genes are involved in cell wall organization and biogenesis. Overall, we identified key regulators of molecular diversity among individuals and provide novel insights into mechanisms of gene regulation. Two ChIP-Seq experiments and one Input DNA-Seq experiment for the yeast strains S96, HS959 and 43 MATa segregants were performed under alpha factor treatment conditions; an additional replicate was also performed for some of the strains. One ChIP-Seq experiment for each parental strain was performed without alpha factor treatment, and one ChIP-Seq experiment for each of the 24 deletion strains was performed under alpha factor treatment.
Project description:We measured the response of S. cerevisiae to arrest in the presence of alpha factor. These were collected in support of a related DNaseI-sequencing study. Keywords: Alpha-factor arrest S.cerevisiae R276 (MATa ura3Δ0 leu2Δ0 his3Δ1 met15Δ0 bar1Δ::KanMX) (C. Boone, University of Toronto; S288c background derived from BY4741), was cultured overnight in 50 ml rich medium (YPD) at 30°C, diluted into 500 ml fresh YPD to an OD660 of ~0.8, and treated with yeast α-factor (Sigma-Aldrich) at a final concentration of 50 ng / ml. This culture was incubated at 30°C with shaking for 3 hours (final OD660 ~1). After this treatment, approximately 90% of the cells had formed mating projections when checked by light microscopy. Total RNA from these cells was isolated using hot acidic phenol. 50 μg of total RNA was treated with Turbo Dnase (Ambion), and checked for integrity using a Bioanalyzer 2100 (Agilent). Total RNA was labeled according to the manufacturer’s protocol and applied to Affymetrix Yeast 2.0 arrays. Data were analyzed using the “affy” package from Bioconductor.
Project description:Transcription profile in YPD media of 48 segregants spores obtained from a cross of the yeast strains S96 and YJM789. These spores are a subset of those published by Mancera et al, Nature, 2008. Two CEL files were mislabelled: eQTL_080822_spore_38B.CEL and eQTL_080826_spore_21C.CEL, actually spores 24A and 8D respectively. The correct spore IDs are in the sample annotation (under StrainOrLine).
Project description:In this study, we constructed three isogenic strains of S96 yrr1Δ background (its native YRR1 gene was knocked out) carrying three different YRR1 alleles, YRR1_S96, YRR1_YJM789 and YRR1_S96-I775E, respectively. We then conducted RNA deep sequencing (RNA-Seq) on the three strains grown in Yeast Peptone Dextrose medium (YPD), YPD + 4NQO and Yeast Peptone glycerol medium (YPglycerol).
Project description:In this study, we constructed three isogenic strains of S96 yrr1Δ background (its native YRR1 gene was knocked out) carrying three different YRR1 alleles, YRR1_S96, YRR1_YJM789, YRR1_S96-I775E, respectively. We then conducted chromatin immuno-precipitation followed by high-throughput sequencing (ChIP-Seq) for Yrr1 protein on the three strains grown in Yeast Peptone Dextrose medium (YPD) and YPD + 4NQO.