COLLABORATIVE RESEARCH: The role of quality control in microbial translation

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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.