Reconstructing the evolution of transcription factor DNA binding specificity across living species and their common ancestors

转录因子是动植物发育过程和环境适应的主要调节因子。它们通过与一组特定的DNA区域结合来实现其功能。 然而，目前还不清楚它们的DNA结合特异性如何与蛋白质序列进化相关。这些知识对于理解它们的功能进化多样性很重要：它们如何获得新的靶基因和新的生物学功能，以及它们在基因复制事件后如何分化。因此，本提案的目的是开发新的计算方法来表征跨物种TF家族中DNA结合特异性的进化。这就是：1）在不同的进化时间尺度上对研究中TF家族的蛋白质序列进行遗传学重建，以及2）使用高通量体外SELEX-seq实验表征和比较重建的祖先TF的DNA结合特异性。新的计算方法，以确定DNA结合特异性的进化变化的重要性将举例说明的精确表征的AP 1/FUL谱系的植物MADS盒TF基因，其复制在起源的核心真双子叶植物和随后的多样化产生了三个旁系同源TF谱系具有非常不同的生物学功能，在开花时间控制，茎生叶和果实发育。因此，该项目将回答DNA结合特异性的变化如何促进这些重要发育调节因子的功能多样化的问题。此外，实验和计算方法的结合也为其他TF系列和模型系统的可比分析提供了一个起点。

Transcription factors (TFs) are major regulators of developmental processes and environmental adaptation in plants and animals. They achieve their function by binding to a specific set of DNA regions. However, it is not well understood how their DNA-binding specificity evolves associated with protein sequence evolution. This knowledge is important to understand their functional evolutionary diversification: how they acquire new target genes and new biological functions, and how they diverge after gene duplication events. Therefore, the aim of this proposal is to develop new computational methods to characterize the evolution of DNA binding specificities in a TF family across species. This is: 1) to phylogenetically reconstruct the protein sequence of the TF family at study at different evolutionary time scales and 2) characterize and compare the DNA binding specificities of reconstructed ancestral TFs using high-throughput in vitro SELEX-seq experiments. The importance of the novel computational methods to determine evolutionary changes in DNA-binding specificity will be exemplified by the precise characterization of the AP1/FUL lineage of the plant MADS-box TF genes, whose duplication at the origin of the core eudicots and subsequent diversification gave rise to three paralogous TF lineages with very different biological functions in flowering time control, cauline leaf and fruit development. Thus, the project will answer the question of how changes in DNA binding specificity have contributed to the functional diversification of these important developmental regulators. In addition, the combination of experimental and computational methods established here also provides a starting point for comparable analyzes in other TF families and model systems.