Pleiotropy, Epistasis, and the Biophysical Adaptation of ssDNA Bacteriophages

描述(由申请人提供)： 拟议的项目将探索多效性和上位性在适应性进化中的作用，并将生物物理和生物化学的想法和问题整合到噬菌体系统的进化框架中。这些实验受到许多病原体经历的种群动态的启发，将使用一种独特的实验-进化方案，涉及快速波动的选择压力，基于生长速度和三个生物物理参数之一：衣壳稳定性、低pH耐受性和新的宿主结合。选择方案包括在宿主内的生长期，在没有噬菌体复制的情况下，不时地对三种生物物理特性中的一种进行强烈选择。对于目标1，该协议将 用来研究五种微病毒型噬菌体基因的个体有益突变对生长速度和三种生物物理特性的多效性影响，并确定这种冲突和一般的多效性如何影响可用于适应的遗传变异。对于目标2，为目标1确定的有益突变将被工程到新的遗传背景中，以揭示突变的生物物理属性在多大程度上是跨背景的相加的。目标3的结果将被用来确定长期适应是否可以通过补偿进化来克服有害的多效性效应，从而使这两个特征同时最大化。拟议项目的结果将成为生物物理和进化之间的桥梁，通过将生物物理参数纳入适合度，生长速度和稳定性之间的关系将提供对蛋白质折叠、功能和进化的基本方面的洞察。长期以来，理论家一直在寻找表征进化过程的概括性，如果存在这样的概括性，它们应该自然地从较低级别的现象中产生。拟议中的实验旨在揭示这种现象是否存在。关于蛋白质热稳定性进化的一般性还没有公布，尽管它们与许多领域有关，从极端细菌的进化到关于进化性和工业用酶的合理设计的基本问题。热稳定性实际上被认为是通过缓冲突变的有害多效性效应来促进进化的。研究结果还将有助于调整当前的适应模型，以改善其现实性，并提高其预测的准确性。多效性是Fisher几何模型的一个定义特征，拟议的实验将量化有助于适应的突变的多效性效应，提供关于多维表型空间中可能的运动类型的信息。他们还将提供突变景观模型中主要参数的估计。虽然上位性疾病已被广泛记载，但其分子基础仍然难以捉摸。这些结果将提供大量关于上位性及其有益突变原因的信息，有关这些信息的数据很少。许多病原体必须在两次感染之间的恶劣条件下生存，并可能因此进化出更强的毒力。拟议的实验系统将作为研究这类选择的影响的模型，并为传染病的进化提供见解。

DESCRIPTION (provided by applicant): The proposed project will explore the roles of pleiotropy and epistasis in adaptive evolution and integrate ideas and questions from biophysics and biochemistry into an evolutionary framework in a bacteriophage system. The experiments, inspired by the population dynamics experienced by many pathogens, will use a unique experimental-evolution protocol involving rapidly fluctuating selective pressures to induce a two-component fitness based on growth rate and one of three biophysical parameters: capsid stability, low-pH tolerance, and novel host binding. The selection protocol consists of periods of growth within hosts punctuated by strong selection for one of three biophysical properties in the absence phage replication. For Aim 1, this protocol will be used to study the pleiotropic effects of individual beneficial mutations in five microvirid bacteriophage genotypes on growth rate and the three biophysical properties and to determine how this conflict, and pleiotropy in general, affects the genetic variation available for adaptatio. For Aim 2, beneficial mutations identified for Aim 1 will be engineered into new genetic contexts to reveal the extent to which biophysical properties of mutations are additive across backgrounds. The results from Aim 3 will be used to determine whether long-term adaptation can allow deleterious pleiotropic effects to be overcome through compensatory evolution to allow the two traits to be simultaneously maximized. The results from the proposed project will serve as a bridge between biophysics and evolution by subsuming biophysical parameters within fitness, and the relationship between growth rate and stability will provide insight into basic aspects of protein folding, function, and evolution. Theoreticians have long sought generalities that characterize the evolutionary process, and if such generalities exist, they should arise naturally from lower-level phenomena. The proposed experiments are designed to reveal such phenomena if they exist. Generalities about the evolution of protein thermal stability have not been forthcoming despite their relevance to a variety of fields, ranging from the evolution of extremophiles to basic questions about evolvability and the rational design of enzymes for industrial uses. Thermal stability is thought actually to promote evolvability by buffering against deleterious pleiotropic effects of mutations. The results will also aid in adjusting current models of adaptation to improve their realism and increase the accuracy of their predictions. Pleiotropy is a defining feature of Fisher's geometric model, and the proposed experiments will quantify the pleiotropic effects of mutations contributing to adaptation, providin information about the types of movements possible in multidimensional phenotypic space. They will also provide estimates of the main parameters in the mutational landscape model. Although epistasis has been widely documented, its molecular basis remains elusive. The results will provide a wealth of information about epistasis and its causes for beneficial mutations, about which few data exist. Many pathogens must survive under harsh conditions between infections and can potentially evolve greater virulence as a result. The proposed experimental system will serve as a model for studying the implications of this type of selection and provide insights into the evolution of infectious diseases.