COLLABORATIVE RESEARCH: The role of quality control in microbial translation

这项研究的目标是了解当细胞在遗传密码中出错时会发生什么--为什么有时错误有助于细胞更好地生长，而在其他情况下，细胞可能会死亡？该项目将整合针对本科生和研究生的实验室研究和培训计划，包括历史上代表性不足的少数民族成员。这些努力的影响将通过在地方和国家科学会议上提交研究结果、学生未来的安置情况以及学生对同行评议出版物的作者来衡量。该项目还将为高中生和社区大学生提供实验室培训和教育经验。此外，该项目还将促进其他更广泛的活动，包括高级细菌遗传学培训课程，以及初级研究人员的指导讲习班。将RNA的四个字母代码翻译成22个字母的蛋白质字母表是细胞生命的一个主要特征。这一过程的准确性，即遗传密码的翻译，决定了基因组中的信息转化为具有正确氨基酸序列的蛋白质的忠诚度。氨酰-tRNA合成酶在翻译中的作用是通过正确地将同源tRNA与其相应的氨基酸配对来定义遗传密码。由于在这些小RNA中发现了独特的序列和结构，氨基酰-tRNA合成酶可以相对容易地选择正确的tRNA。相比之下，氨基酸结构的简单性使区分它们变得困难得多。合成酶通过两种方式实现必需的氨基酸底物专一性，以将翻译错误保持在可接受的水平：优先结合同源氨基酸，选择性编辑非同源氨基酸和非蛋白质氨基酸。编辑显著减少了体外合成氨基酰tRNA过程中的错误频率，尽管反应机制和体内编辑的影响的许多细节仍不清楚。我们已经确定了不同的生长条件，在这些条件下，需要编辑才能存活，例如，在应对氧化应激和饥饿时，以及在细胞分化期间。目前尚不完全清楚为什么只在部分生长条件下需要通过AARS编辑进行质量控制(QC)，但在其中一些条件下，缺乏编辑会导致有毒的非蛋白质氨基酸在蛋白质中积累。在其他情况下，缺乏编辑可能会减少未带电的tRNA的数量，tRNA是细胞感知的一种关键分子，用于启动必要的饥饿反应。该项目的目标是利用微生物遗传、生理和生化方法来确定质量控制影响细胞功能以应对压力的机制，以及质量控制对非同源和非蛋白质氨基酸误译的影响。

The goal of this research is to understand what happens to cells when they make mistakes in their genetic codes - why is it that sometimes mistakes help cells grow better, while in other instances cells may die? The project will integrate laboratory research and training programs for undergraduate and graduate students, including members of historically underrepresented minorities. The impact of these efforts will be measured by the presentation of research findings at local and national scientific meetings, by future placements of students, and by student authorship of peer reviewed publications. This project will also provide laboratory training and education experiences for high school and community college students. In addition, the project will contribute to other broader activities including a training course in advanced bacterial genetics, and mentoring workshops for junior investigators.Translating the four-letter code of RNA into the twenty-two-letter alphabet of proteins is a central feature of cellular life. The accuracy of this process, translation of the genetic code, determines how faithfully the information in a genome is transformed into proteins with correct amino acid sequences. The role of aminoacyl-tRNA synthetases in translation is to define the genetic code by correctly pairing cognate tRNAs with their corresponding amino acids. Aminoacyl-tRNA synthetases select the correct tRNA with comparative ease due to the unique sequences and structures found in these small RNAs. In comparison, the simplicity of amino acid structures makes discrimination between them far more difficult. Synthetases achieve the amino acid substrate specificity necessary to keep errors in translation to an acceptable level in two ways: preferential binding of the cognate amino acid, and selective editing of non-cognate and non- protein amino acids. Editing significantly decreases the frequency of mistakes during aminoacyl- tRNA synthesis in vitro, although many details of the reaction mechanism and the impact of in vivo editing remain unclear. We have identified different growth conditions where editing is required for viability, for example in response to oxidative stress and starvation, and during cellular differentiation. It is not fully clear why quality control (QC) through aaRS editing is required under only a subset of growth conditions, but under some of these conditions, the lack of editing leads to the accumulation of toxic, non-protein amino acids in proteins. Under other conditions, the lack of editing may decrease the amount of uncharged tRNA, a critical molecule that cells sense in order to mount a necessary starvation response. The objectives of this project are to use microbial genetic, physiological and biochemical approaches to determine the mechanisms by which quality control impacts cellular function in response to stress, and the impact of quality control on mistranslation with non-cognate and non-protein amino acids.