Translational quality control by trans-editing domains

摘要 氨基酰-tRNA合成酶(ARs)建立了遗传密码的规则，由此每个氨基酸都是 附着在同源tRNA上的。这一过程中的错误会导致误译，这可能会对细胞产生毒性。近期 研究表明，细胞特定的要求和环境条件施加的选择性力量 潜在地塑造了质量控制机制。大约一半的ARS拥有校对(或编辑)功能 对错误荷电的AA-tRNA的作用和非蛋白质氨基酸构成最大的证据 对忠诚度的威胁开始显现。Musier-Forsyth实验室在翻译质量领域的早期工作 控制集中在我们发现的第二类脯氨酰-tRNA合成酶(ProRS)编辑上。这导致了详细的 细菌ProRS新的转移后编辑结构域(INS)的机理理解与论证 INS结构域，当在ARS环境外单独纯化时，在tRNA中是完全功能的 脱酰化。我们随后发现，单结构域INS同源物广泛存在于细菌和 近年来，我们在这一领域的重点几乎完全转向了解这些类INS的功能 TRNA编辑中的结构域。然而，许多关于这些推定的生理功能的悬而未决的问题 反式编辑蛋白质仍然存在。此Mira应用程序中描述的研究的总体目标是 揭示一个称为INS超家族的不断增长的反式编辑蛋白家族的特定功能。这 多样化但普遍保守的家族现在拥有坚实的和积累的体外结构-功能知识 BASE，它强烈支持在保持翻译保真度方面发挥作用。我们对更广泛的知识的了解 这些蛋白质的生理作用，特别是在真核生物中的作用，仍处于初级阶段，刚刚开始揭示。 扮演的角色比之前预期的更广泛。这一重大差距将在这项工作中得到解决。而古典的敲门声- 仅定义必需基因和非必需基因的向下筛选不会立即识别编辑结构域 至关重要的是，这些领域的强大保守性意味着它们发挥着重要作用，而且在大多数情况下仍然如此 未被发现的，在细胞生存和竞争力中的作用。拟议的研究旨在解决一些 关于生理转译功能和潜在兼职的许多悬而未决的问题 INS超家族的功能。这些结构域在真核生物中很大程度上是未被探索的，包括一个新的亚 在许多单细胞真核病原体中编码的家族簇。转译编辑的治疗潜力 域名还没有被利用，这是该领域的另一个重大差距，我们希望通过我们的 有计划的研究。从长远来看，将针对新的翻译保真机制的药物与 已知的核糖体靶向蛋白质合成抑制剂，如氨基糖苷类，可能会导致更有效的 治疗策略。

Summary Aminoacyl-tRNA synthetases (ARSs) establish the rules of the genetic code, whereby each amino acid is attached to a cognate tRNA. Errors in this process lead to mistranslation, which can be toxic to cells. Recent studies suggest that the selective forces exerted by cell-specific requirements and environmental conditions potentially shape quality control mechanisms. Approximately half of the ARSs possess a proofreading (or editing) function to hydrolyze mischarged aa-tRNAs and evidence that non-proteinaceous amino acids pose the greatest threat to fidelity is beginning to emerge. Early work in the Musier-Forsyth lab in the field of translational quality control focused on our discovery of Class II prolyl-tRNA synthetase (ProRS) editing. This led to a detailed mechanistic understanding of the novel bacterial ProRS posttransfer editing domain (INS) and the demonstration that the INS domain, when purified on its own outside the context of the ARS, was fully functional in tRNA deacylation. We subsequently discovered that single-domain INS homologs are widespread in Bacteria and in recent years, our focus in this area has turned almost entirely to understanding the function of these INS-like domains in tRNA editing. However, many open questions regarding the physiological function of these putative trans-editing proteins remain. The overarching goal of the research described in this MIRA application is to uncover the specific functions of a growing family of trans-editing proteins known as the INS superfamily. This diverse yet universally conserved family now has a solid and accumulating in vitro structure-function knowledge base, which strongly supports a role in maintaining translational fidelity. Our knowledge of the broader physiological roles of these proteins, especially in eukaryotes, is still in its infancy and is just beginning to reveal wider roles than previously anticipated. This major gap will be addressed in this work. While classical knock- down screens that only define essential versus non-essential genes do not immediately identify editing domains as essential, the strong conservation of these domains implies they play important, and in most cases still undiscovered, roles in cell survival and competitiveness. Proposed studies are designed to address some of the many open questions with regard to both physiological trans-editing functions and potential moonlighting functions of the INS superfamily. These domains are largely unexplored in eukaryotes, including a novel sub- family cluster that is encoded in many unicellular eukaryotic pathogens. The therapeutic potential of trans-editing domains has not been exploited and represents another major gap in the field that we hope to address by our planned studies. In the long term, combining drugs that target novel translational fidelity mechanisms along with known ribosome-targeting protein synthesis inhibitors such as aminoglycosides, may results in more effective therapeutic strategies.