Comprehensive analysis of fitness effects and epistasis along a billion-year evolutionary trajectory

十亿年进化轨迹上的适应度效应和上位性综合分析

• 批准号: 10212033
• 负责人: DANIEL N BOLON
• 金额: $ 56.58万
• 依托单位: UNIVERSITY OF CHICAGO
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-06-01 至 2025-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10212033
• 关键词: Accidents; Address; Affect; Amino Acid Sequence; Amino Acids; Animals; Architecture; Biochemical; Biological; Biological Assay; Biological Models; Biological Process; Biophysical Process; Biophysics; Catalysis; Cells; Chemicals; Client; Complement; Data; Disease; Engineering; Eukaryota; Eukaryotic Cell; Event; Evolution; Future; Gene Proteins; Genetic; Genetic Epistasis; Genetic Variation; Health; Human; Knowledge; Lead; Libraries; Ligand Binding; Measures; Mediating; Methods; Molecular Chaperones; Molecular Conformation; Molecular Evolution; Mutation; Outcome; Pattern; Phylogenetic Analysis; Phylogeny; Play; Process; Property; Protein Engineering; Proteins; Public Health; Recording of previous events; Resolution; Role; Saccharomyces cerevisiae; Sampling; Signal Transduction; Site; Structure; Structure-Activity Relationship; System; Techniques; Technology; Time; Work; Yeasts; biological systems; chemical genetics; experimental analysis; experimental study; fitness; fungus; gene therapy; improved; insight; mutant; mutation screening; new technology; prevent; protein folding; reconstruction; sample fixation; tool

Epistatic interactions within proteins can, in principle, make the paths and outcomes of evolution contingent on chance events; they can also entrench proteins with residues that appear to be optimal but are accidents of history. The extent to which epistasis actually affected the trajectory and outcomes of molecular evolution depends on the fitness effects of substitutions when they occurred in history compared to their potential effects earlier or later in time and on the temporal order in which interacting substitutions occurred. Deep mutational scanning studies have revealed pervasive epistasis among the huge number of possible mutations, but no studies have directly assessed how the fitness effects of substitutions that happened during history changed over time as the protein evolved. We will perform the first comprehensive experimental analysis of the fitness effects of all amino acid states that evolved in a protein during a long-term phylogenetic trajectory, both at the time they occurred and if they had occurred at other points in history. These data will be analyzed in the ordered temporal context of the protein's phylogeny and supplemented with biochemical experiments, enabling a deep characterization of the causes and consequences of epistasis, contingency and entrenchment across the billion-year history of an essential protein. Our model system is ideal for this purpose. Hsp90, the essential molecular chaperone in all eukary- otic cells, plays key roles in protein folding and maturation, cell signaling, and a wide range of diseases. Strong phylogenetic signal allows confident reconstruction of the billion-year evolutionary history of Hsp90's protein sequence from the last common ancestor of animals, fungi and related protists to present-day Saccharomyces cerevisiae. We will generate targeted protein libraries containing every ancestral and derived state that occurred during this phylogenetic trajectory, singly and in every possible pair, in the background of all 30 reconstructed ancestral proteins along the trajectory. Using a high- resolution bulk competition assay in yeast, we will precisely measure selection coefficients and epistatic interactions and quantify how these properties changed over time. This will reveal the fitness effects and interactions of every substitution at the approximate time it occurred, as well as the effects and interactions it would have had if it happened (or reverted to the ancestral state) at any point earlier or later during the trajectory. We will also apply biophysical and structural techniques to elucidate the underlying biochemical mechanisms that drove these genetic and evolutionary phenomena. This work will provide deep new insight into the ways in which proteins' genetic and physical architecture influences, and is influenced by, the processes by which they evolve; it will also strengthen our understanding of sequence-structure-function relationships in a biologically essential protein.

蛋白质中的上毒相互作用原则上可以使进化的路径和结果 取决于机会事件；它们还可以使蛋白质构成蛋白质，这些残留物似乎是最佳但 是历史的事故。上学实际影响的轨迹和结果的程度 分子进化取决于替代在历史上发生的适应性影响 与它们的潜在影响更早或更晚的时间和时间顺序相比 发生替换。深度突变扫描研究表明，在 大量可能的突变，但没有直接评估的研究如何影响 随着蛋白质的发展，历史上发生的替换随着时间的流逝而发生了变化。我们将表演 所有氨基酸的适应性效应的首次全面实验分析 在长期系统发育轨迹中在蛋白质中进化的状态，均在 它们发生的时间以及是否发生在历史上的其他时刻。这些数据将是 在蛋白质的系统发育的有序时间环境中分析并补充了生化 实验，能够深刻地表征上学的原因和后果， 基本蛋白质的十亿年历史的偶然性和根深蒂固。 我们的模型系统是理想的目的。 HSP90，所有真核生物中必不可少的分子伴侣 耳细胞，在蛋白质折叠和成熟，细胞信号传导以及多种疾病中起关键作用。 强大的系统发育信号可以自信重建十亿年的进化历史 HSP90的蛋白质序列来自动物，真菌和相关生物的最后一个共同祖先到 当今的酿酒酵母。我们将生成靶向蛋白质库，其中包含每个库 在这种系统发育轨迹期间发生的祖先和衍生状态 对，在沿轨迹的所有30个重建祖传蛋白的背景下。使用高 分辨率的散装竞争测定法在酵母中，我们将精确衡量选择系数和上毒 交互并量化这些属性如何随时间变化。这将揭示健身效果，并 大约发生的每个替代的相互作用，以及效果和 如果互动发生（或恢复为祖先国家），它将在任何时候或 后来在轨迹期间。我们还将应用生物物理和结构技术来阐明 驱动这些遗传和进化现象的基本生化机制。这项工作 将对蛋白质的遗传和物理结构的方式提供深刻的见解 影响并受到它们进化的过程的影响；它也将加强我们的 了解生物学上必不可少的蛋白质中序列结构功能关系。