High-resolution mapping and quantitative modeling of cooperative RNA binding in mRNPs

我们之前已经开发和优化了将mRNP组分和其他RNbinding因子映射到体内转录组上的方法(Schulz等人，Cell 2013；Baejen等人，Mol.单元格2014)。这些方法使识别前mRNA和短寿命RNA的蛋白质的全球图谱成为可能，并证明了RNA识别不能用标准的方法来描述，这些方法用于描述DNA识别，如位置权重矩阵(PWMS)。在这里，我们提出了一种综合的实验-理论方法，以促进新的Schwerpenkt计划(SPP)的两个主要目标。首先，我们将利用PAR-CLIP(光活化核糖核苷增强的交联和免疫沉淀)系统地鉴定mRNA结合蛋白的细胞结合部位，以研究mRNPs在萌芽酵母中的形成和功能。其次，我们将开发新的技术，通过多个因素的协同RNA结合来定量描述RNA识别，这将使我们能够预测RNA序列结合的偏好。本工作的目的是了解mRNP在体内的形成，并根据不同RNA类别与细胞因子的关系了解它们的生物发生和命运。衍生技术和生物信息学工具将提供给SPP团队。

We have previously developed and optimized methods to map mRNP components and other RNAbinding factors onto the transcriptome in vivo (Schulz et al., Cell 2013; Baejen et al., Mol. Cell 2014). These methods enabled the first global mapping of proteins recognizing pre-mRNA and shortlived RNAs, and demonstrated that RNA recognition cannot be described by standardmethods that are used to describe DNA recognition such as position weight matrices (PWMs). Here we propose an integrated experimental-theoretical approach to contribute to two major goals of the new Schwerpunktprogramm (SPP). First, we will systematically identify cellular binding sitesfor mRNA-binding proteins with the use of PAR-CLIP (photoactivatable ribonucleoside-enhanced crosslinking and immunoprecipitation), to study formation and function of mRNPs in the budding yeast S. cerevisiae. Second, we will develop new techniques enabling a quantitative description of RNA recognition by cooperative RNA binding of multiple factors that will allow us to predict RNAsequence-binding preferences. The aim of this work is to understand mRNP formation in vivo and to understand the biogenesis and fate of different RNA classes based on their association with cellular factors. The derived technology and bioinformatic tools will be provided to SPP teams.