Integrative analysis of tissue-specific alternative splicing regulation under adaptive selection

适应性选择条件下组织特异性选择性剪接调控的综合分析 项目摘要 组织特异性选择性剪接（AS）从单个基因产生多个转录本， 哺乳动物细胞和表型的复杂性。这一过程受到RNA结合蛋白的严格调控 （RBP），其识别其靶转录物中的特定调控元件。一个长期存在的假设， 进化观点是顺式调节序列突变导致AS调节改变 为哺乳动物的物种形成提供了主要的驱动力，包括人类和 黑猩猩虽然最近的研究令人信服地证明了AS在多个物种中的普遍物种差异， 组织，两个基本问题仍然存在：1）哪些进化剪接变化是适应性选择 在特定的血统中，因为获得的健身优势？2)是什么潜在的突变导致了 改变选择性的制度？这些挑战与剪接密码的内在简并性有关 缺乏有效的计算和实验方法进行直接比较分析。主 本项目的目标是克服这些局限性，以便我们能够在自适应条件下识别不同的AS事件， 选择和影响重要顺式调节元件的因果突变。为了实现这一总体目标， 为此，我们制定了三个具体目标。在目标1中，我们将使用系统发育遗传学方法来检测谱系- AS的选择强度和适应性进化的特定变化，以及AS调控区的地图， 谱系特异性适应性进化在目标2中，为了阐明导致 剪接分歧，我们将系统地映射不同的蛋白质-RNA相互作用，并重建 通过CLIP足迹，RBP扰动和 比较基因组分析。在目标3中，我们将重点关注MAPT选择性外显子10，它涉及 额颞叶痴呆（FTD），并进行详细分析以揭示其分歧拼接的机制 以及相关的细胞功能。如果成功，这项研究将推动我们的 理解AS进化对不同哺乳动物之间潜在表型差异的贡献 物种及其潜在机制。本研究中确立的战略还将提供一个新的 可推广的范例，以更好地理解拼接代码。

Integrative analysis of tissue-specific alternative splicing regulation under adaptive selection PROJECT SUMMARY Tissue-specific alternative splicing (AS) generates multiple transcripts from single genes and contributes critically to the cellular and phenotypic complexity of mammals. This process is tightly regulated by RNA-binding proteins (RBPs) which recognize specific regulatory elements in their target transcripts. A long-standing hypothesis from the evolutionary perspective is that changes of AS regulation due to mutations in cis-regulatory sequences provide a major driving force of speciation in mammals, including closely related species such as human and Chimpanzee. While recent studies convincingly demonstrated the pervasive species difference of AS in multiple tissues, two fundamental questions remain: 1) which evolutionary splicing changes are under adaptive selection in specific lineages because of acquired fitness advantages? 2) what are the underlying mutations that led to the shifted selective regimes? These challenges are associated with the intrinsic degeneracy of the splicing code and lack of effective computational and experimental approaches for direct comparative analysis. The primary goal of this project is to overcome these limitations, so that we can identify divergent AS events under adaptive selection and the causal mutations affecting important cis-regulatory elements. To achieve this overarching goal, we formulated three specific aims. In Aim 1, we will use phylogenetic genetic methods to detect lineage- specific changes in selection intensity and adaptive evolution of AS and map AS regulatory regions experiencing lineage-specific adaptive evolution. In Aim 2, to elucidate the underlying regulatory mechanisms that led to splicing divergence, we will systematically map divergent protein-RNA interactions and reconstruct the evolutionary history in the phylogeny through a combination of CLIP footprints, perturbation of RBPs and comparative genomic analysis. In Aim 3, we will focus on MAPT alternative exon 10, which is implicated in frontotemporal dementia (FTD), and perform detailed analysis to reveal the mechanism of its divergent splicing in primates and the associated functions at the cellular level. If successful, this study will advance our understanding of the contribution of AS evolution to potential phenotypic differences among different mammalian species and the underlying mechanisms. The strategy established in this study will also provide a new and generalizable paradigm to better understand the splicing code.