Hayden Eric J, Bendixsen Devin P, Wagner Andreas
Department of Biological Science, Boise State University, Boise, ID 83725; Biomolecular Sciences PhD Program, Boise State University, Boise, ID 83725;
Biomolecular Sciences PhD Program, Boise State University, Boise, ID 83725;
Proc Natl Acad Sci U S A. 2015 Oct 6;112(40):12444-9. doi: 10.1073/pnas.1420902112. Epub 2015 Sep 23.
Phenotypic capacitance refers to the ability of a genome to accumulate mutations that are conditionally hidden and only reveal phenotype-altering effects after certain environmental or genetic changes. Capacitance has important implications for the evolution of novel forms and functions, but experimentally studied mechanisms behind capacitance are mostly limited to complex, multicomponent systems often involving several interacting protein molecules. Here we demonstrate phenotypic capacitance within a much simpler system, an individual RNA molecule with catalytic activity (ribozyme). This naturally occurring RNA molecule has a modular structure, where a scaffold module acts as an intramolecular chaperone that facilitates folding of a second catalytic module. Previous studies have shown that the scaffold module is not absolutely required for activity, but dramatically decreases the concentration of magnesium ions required for the formation of an active site. Here, we use an experimental perturbation of magnesium ion concentration that disrupts the folding of certain genetic variants of this ribozyme and use in vitro selection followed by deep sequencing to identify genotypes with altered phenotypes (catalytic activity). We identify multiple conditional mutations that alter the wild-type ribozyme phenotype under a stressful environmental condition of low magnesium ion concentration, but preserve the phenotype under more relaxed conditions. This conditional buffering is confined to the scaffold module, but controls the catalytic phenotype, demonstrating how modularity can enable phenotypic capacitance within a single macromolecule. RNA's ancient role in life suggests that phenotypic capacitance may have influenced evolution since life's origins.
表型缓冲指的是基因组积累条件性隐藏突变的能力，这些突变只有在特定的环境或基因变化后才会显现出改变表型的效应。缓冲对于新形式和功能的进化具有重要意义，但实验研究的缓冲背后的机制大多局限于复杂的多组分系统，通常涉及几个相互作用的蛋白质分子。在这里，我们在一个简单得多的系统中证明了表型缓冲，即一个具有催化活性的单个RNA分子（核酶）。这个天然存在的RNA分子具有模块化结构，其中一个支架模块充当分子内伴侣，促进第二个催化模块的折叠。先前的研究表明，活性并非绝对需要支架模块，但它会显著降低形成活性位点所需的镁离子浓度。在这里，我们通过对镁离子浓度进行实验性扰动来破坏这种核酶某些遗传变体的折叠，并使用体外筛选和深度测序来鉴定具有改变表型（催化活性）的基因型。我们鉴定出多个条件性突变，这些突变在低镁离子浓度的应激环境条件下改变野生型核酶表型，但在更宽松的条件下保持表型。这种条件性缓冲局限于支架模块，但控制着催化表型，证明了模块化如何在单个大分子中实现表型缓冲。RNA在生命中的古老作用表明，自生命起源以来，表型缓冲可能影响了进化。
