tRNA biology in healthy ageing: Functional differentiation and expression of tRNAiMet loci in Drosophila.

老年人在我们社会中的比例正在迅速增加。对许多人来说，老年人身体虚弱，功能受损，对疾病的易感性增加。我们人口的老龄化正在造成巨大的个人和社会经济代价，迫切需要加以解决。了解这种可塑性为我们提供了一个机会，设计干预措施，以改善人类健康到老年。重要的是，直接针对衰老具有广泛的健康改善潜力，而不仅仅局限于单一疾病。该研究项目将回答与衰老密切相关的转移RNA（tRNA）生物学中的一个基本问题。tRNA是普遍用于将遗传密码翻译成蛋白质的衔接分子。我们对tRNA生物学的兴趣来自于我们对细胞蛋白质合成的关键调节因子雷帕霉素激酶复合物1（TORC 1）的活性促进动物衰老的分子机制的理解。我们发现TORC 1通过RNA聚合酶III起作用，RNA聚合酶III本身负责产生tRNA。因此，了解tRNA生物学将推进我们对衰老过程的认识。动物基因组中存在100个tRNA基因，通常是相同序列的拷贝，使其生物学分析复杂化。历史上，这种相同的拷贝被认为是简单的冗余，提供了多个模板，以促进蛋白质合成所需的高表达水平。然而，越来越多的人认识到，将tRNA拷贝置于不同的基因组环境中可以使动物微调tRNA表达模式，从而使相同或相似的tRNA的生物功能多样化。由于缺乏合适的动物模型和遗传试剂，这一假设仍然没有得到实验证实，尽管它具有根本的重要性。果蝇是一种小动物，但它是一种强大的实验模型，已被证明可用于了解包括人类在内的动物的基本生物学以及它们如何衰老。为了开始探索tRNA的生物学作用，我们专注于一种专门用于启动蛋白质合成的tRNA，即启动子tRNAMet（tRNAiMet）。我们从四个不同的基因组位置产生了一组删除tRNAiMet拷贝的果蝇突变体。我们的初步表型分析显示，至少有一个tRNAiMet基因座，有助于老化，并表明独特的和冗余的生物体功能的相互作用的四个基因座。我们将使用这组突变体，以及我们生成的一组报告基因系，来回答tRNA生物学中的一个基本问题，这将有助于我们理解衰老：为什么在动物的基因组中存在多个相同的tRNA基因拷贝？首先，我们将对单个突变体及其组合进行广泛的表型分析，以确定某些基因座所特有的生物功能以及冗余的生物功能，重点是衰老。其次，我们将使用报告基因系来评估发育期间和不同成体组织和器官中每个位点的表达。我们将把这两组发现联系起来，并通过正式测试独特的表达模式是否驱动独特的生物体功能来整合它们。第三，我们将研究tRNAiMet基因座在衰老可塑性中的作用。具体来说，我们将确定是否长寿导致减少营养摄入或TORC 1抑制是部分由tRNAiMet水平的变化引起的。该项目将回答tRNA生物学中的一个基本问题，为我们在动物生理学背景下理解tRNA提供一个步骤变化。它将为我们提供一个开拓性的洞察他们在动物衰老中的作用，以及所需的知识来破译他们的生物体功能下游的TORC 1。反过来，这种知识将为旨在确保人类一生健康的干预措施提供信息。

The proportion of older people in our societies is rapidly increasing. For many, older age comes with increased frailty, impaired function and increased susceptibility to disease. The ageing of our populations is incurring massive personal and socioeconomic costs that need to be urgently addressed.Ageing itself can be modulated. Understanding this plasticity presents us with an opportunity to devise interventions to improve human health into old age. Importantly, directly targeting ageing has the potential for broad health improvements not limited to a single disease.This research project will answer a fundamental question in the biology of transfer RNAs (tRNAs) with strong relevance to ageing. tRNAs are the adaptor molecules universally used to translate the genetic code into proteins. Our interest in tRNA biology comes from our work on understanding the molecular mechanisms whereby the activity of a key regulator of cellular protein synthesis, the Target of Rapamycin Kinase Complex 1 (TORC1), promotes animal ageing. We found that TORC1 acts though RNA polymerase III, which itself is responsible for generating tRNAs. Hence, understanding tRNA biology will advance our knowledge of the ageing process.100s of tRNA genes are present in animal genomes, often as copies of identical sequence, complicating the analysis of their biology. Historically, such identical copies were thought to be simply redundant, providing multiple templates to facilitate high expression levels required for protein synthesis. However, there is growing appreciation that placing tRNA copies in different genomic contexts has allowed animals to finetune tRNA expression patterns thus diversifying organismal functions of identical or similar tRNAs. This hypothesis remains experimentally unaddressed due to lack of suitable animal models and genetic reagents, despite its fundamental importance. The fruit fly is a small animal but a powerful experimental model that has proven utility in understanding the basic biology of animals, including humans, and how they age. To start probing into the organismal roles of tRNAs, we focused on a tRNA specialised for initiation of protein synthesis, the initiator tRNAMet (tRNAiMet). We generated a set of fly mutants deleting copies of tRNAiMet from four different genomic locations. Our preliminary phenotyping revealed at least one tRNAiMet locus that contributes to ageing and indicated an interplay of unique and redundant organismal functions for the four loci. We will use this set of mutants, together with a set of reporter lines we have generated, to answer a fundamental question in tRNA biology, which will help us understand ageing: Why are multiple, identical copies of tRNA genes present throughout an animal's genome?Firstly, we will perform extensive phenotyping of single mutants and their combinations to identify organismal functions that are unique to certain loci as well as those that are redundant, with focus on ageing. Secondly, we will use reporter lines to assess the expression from each locus during development and in different adult tissues and organs. We will correlate the two sets of findings as well as integrate them by formally testing if unique expression patterns drive unique organismal functions. Thirdly, we will examine the role of tRNAiMet loci in the plasticity of ageing. Specifically, we will determine if longevity resulting from a reduction in nutrient intake or TORC1 inhibition is in part caused by changes in tRNAiMet levels.The project will answer a fundamental question in tRNA biology to provide us with a step change in understanding of tRNAs in the context of animal physiology. It will provide us with a pioneering insight into their role in animal ageing, as well as the knowledge required to decipher their organismal functions downstream of TORC1. In turn, this knowledge will inform interventions aimed at ensuring human health throughout the life course.