Global analysis of uORF evolution and function

虽然基因表达的差异明显地影响着生物体的表型，但有一个基本的 在理解转录后过程的作用方面存在差距。因为转录后过程 对蛋白质生产的调控至关重要，解决这一知识差距将有助于更好地 基因型和表型之间的关系模型。上游开放阅读框 (UORF)是在大多数人类基因中发现的调控元件，以及一些与疾病相关的突变 似乎改变了uORF的存在。最近，数百个uORF以非8月开始启动 在许多生物体中都发现了密码子。尽管有大量的这些因素，但 AUG和非AUG uORF的功能和进化仍然难以捉摸。这个项目的长期目标是 是为了确定基因变异在转录后顺式调控中的功能和影响 元素。这项提案目的是确定新的uORF如何演变和管理 翻译。我们的中心假设是，8月和非8月的uORF具有不同的调节作用， 导致了不同的进化轨迹。这一假设是基于我们的初步数据，因为我们 已经鉴定出数百个具有不同基因组特性的8月和非8月uORF 酵母菌。我们将通过追求以下具体目标来验证我们的中心假设：1) 研究AUG和非AUG uORF的演变；2)确定AUG和Non-ORFs的功能 Aug uORF；以及3)确定RNA结合蛋白在uORF调节中的作用。在第一个 目的：我们将鉴定在四种条件下生长的不同菌株和种类的酵母中具有活性的uORFs。 以验证AUG和非AUG uORF具有不同进化节奏和模式的假设。这个 第二个目标是使用FACS-uORF来确定数百个uORF的基因调控功能， 我们开发的新型双荧光报告系统。我们将使用这些数据生成预测性数据 UORF功能的模型。目标3将确定全基因组RNA结合蛋白在调控中的作用 UORF，并将这些知识集成到我们的预测模型中。这种方法是创新的，因为它 结合精致的系统生物学工具和优秀的模式属来研究生物的进化 转录后基因调控。这项拟议的研究意义重大，因为预计它将 从根本上推进基因组学、进化和系统生物学领域的发展 UORF的进化和功能特性。因为翻译机制是高度 保守的话，拟议工作产生的知识预计将改进 其他真核生物中非编码遗传变异与表型的关系，包括 人类。

While variation in gene expression clearly affects the phenotypes of organisms, there is a fundamental gap in understanding the role of post-transcriptional processes. Because post-transcriptional processes are critical for the regulation of protein production, addressing this knowledge gap will facilitate better models of the relationship between genotype and phenotype. upstream Open Reading Frames (uORFs) are regulatory elements found in most human genes, and some disease-linked mutations appear to alter the presence of uORFs. Recently, hundreds of uORFs initiating with non-AUG start codons have been identified in many organisms. Despite the vast number of these elements, the functions and evolution of AUG and non-AUG uORFs remain elusive. The long-term goal of this project is to determine the functions and impact of genetic variation in post-transcriptional cis-regulatory elements. The objective of this proposal is to determine how new uORFs evolve and regulate translation. Our central hypothesis is that AUG and non-AUG uORFs have different regulatory roles, leading to different evolutionary trajectories. This hypothesis is based on our preliminary data, as we have identified hundreds of AUG and non-AUG uORFs with different genomic properties in Saccharomyces yeasts. We will test our central hypothesis by pursuing the following specific aims: 1) Investigate the evolution of AUG and non-AUG uORFs; 2) Determine the functions of AUG and non- AUG uORFs; and 3) Identify the roles of RNA binding proteins in uORF-mediated regulation. In the first aim, we will identify active uORFs in diverse strains and species of yeasts grown under four conditions to test the hypothesis that AUG and non-AUG uORFs have different evolutionary tempo and mode. The second aim will determine the gene-regulatory functions of hundreds of uORFs using FACS-uORF, a novel dual-fluorescence reporter system we developed. We will use these data to generate predictive models of uORF function. Aim 3 will identify genome-wide roles of RNA Binding Proteins in regulating uORFs, and integrate this knowledge in our predictive models. This approach is innovative, in that it combines exquisite systems biology tools with an excellent model genus to investigate the evolution of post-transcriptional gene regulation. The proposed research is significant because it is expected to fundamentally advance the fields of genomics, evolutionary, and systems biology by deciphering the evolutionary and functional properties of uORFs. Because translation mechanisms are highly conserved, the knowledge generated by the proposed work is expected to improve models of the relationship between non-coding genetic variation and phenotype in other eukaryotes, including humans.