Translational quality control by trans-editing domains

总结 氨酰-tRNA合成酶（ARSs）建立遗传密码的规则，从而每个氨基酸都是 连接到一个同源的tRNA上这个过程中的错误会导致误译，这可能对细胞有害。最近 研究表明，细胞特异性需求和环境条件所施加的选择力 可能形成质量控制机制。大约一半的ARS拥有校对（或编辑） 功能水解错误的aa-tRNA，并有证据表明，非蛋白质氨基酸构成最大的 对忠诚的威胁开始出现。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II类脯氨酰-tRNA合成酶（ProRS编辑）。这导致了详细的 新的细菌ProRS转移后编辑结构域（INS）的机制理解和演示 INS结构域，当在ARS的环境之外单独纯化时，在tRNA中是完全功能性的， 脱酰作用。我们随后发现，单结构域INS同源物广泛存在于细菌和哺乳动物中。 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， 在tRNA编辑领域。然而，许多关于这些假定的生理功能的开放性问题， 反式编辑蛋白仍然存在。本MIRA应用程序中描述的研究的总体目标是 揭示了一个不断增长的反式编辑蛋白家族的特定功能，该家族被称为INS超家族。这 多样但普遍保守的家族现在有一个坚实的和积累的体外结构-功能知识 碱基，它强烈支持维持翻译保真度的作用。我们对更广泛的知识 这些蛋白质的生理作用，特别是在真核生物中，仍处于起步阶段， 比以前预期的更广泛的角色。这项工作将处理这一重大差距。当经典的敲门声- 仅定义必需基因与非必需基因的向下筛选不能立即识别编辑域 这些结构域的高度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是重要的。 在细胞存活和竞争中的作用。拟议的研究旨在解决一些 关于生理上的翻译编辑功能和潜在的兼职， INS超家族的功能。这些领域在真核生物中基本上是未探索的，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。反式编辑的治疗潜力 域尚未被利用，代表了我们希望通过我们的 计划的研究。从长远来看，将靶向新的翻译保真度机制的药物沿着 已知的核糖体靶向蛋白质合成抑制剂，如氨基糖苷类， 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.