Genome-wide Ste12-binding site mapping in MATa segregants of YJM789 x S96 cross. Saccharomyces cerevisiae

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. Overall design: 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.

本研究首次绘制了个体间转录因子结合差异图谱，并阐明了该变异的遗传基础。本研究在两个高度分化的酵母菌株YJM789与S288c杂交所得的43个分离株及其亲本菌株的α因子处理细胞中，通过染色质免疫共沉淀测序（Chromatin Immunoprecipitation Sequencing, ChIP-Seq）测定了全基因组Ste12结合谱。本研究鉴定到个体间存在广泛的Ste12结合变异，并定位了介导该变异的顺式（cis-）和反式（trans-）作用位点。研究表明，绝大多数转录因子（Transcription Factor, TF）结合变异为顺式连锁变异，且诸多变异与Ste12以及数种已知及推测的Ste12辅因子的结合基序内的多态性相关。本研究还鉴定到两个反式作用因子AMN1与FLO8，它们在α因子处理条件下可调控Ste12与10余个基因启动子的结合。此前尚无研究表明这两个基因可调控Ste12，我们推测它们可能是基因活性与表型多样性的关键介导因子。Ste12结合与200余个基因的基因表达呈显著相关，这表明结合变异具有生物学功能。诸多存在结合变异的基因参与细胞壁组织与生物发生过程。综上，本研究鉴定了介导个体间分子多样性的关键调控因子，并为基因调控机制提供了全新的研究视角。实验设计：针对酵母菌株S96、HS959以及43个MATa型分离株，分别开展2次染色质免疫共沉淀测序（Chromatin Immunoprecipitation Sequencing, ChIP-Seq）实验与1次输入DNA测序（Input DNA Sequencing, Input DNA-Seq）实验，所有实验均在α因子处理条件下进行；部分菌株额外设置了生物学重复。针对每个亲本菌株，各开展1次未经过α因子处理的ChIP-Seq实验；针对24株基因缺失菌株，各开展1次α因子处理条件下的ChIP-Seq实验。