Complex modifications of tRNA: regulatory roles and crosstalk with DNA metabolism

tRNA 的复杂修饰：调节作用以及与 DNA 代谢的串扰

• 批准号: 8884789
• 负责人: Peter C Dedon
• 金额: $ 40.34万
• 依托单位: UNIVERSITY OF FLORIDA
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-03-02 至 2019-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8884789
• 关键词: 7-deazaguanine; 7-deazaguanosine; Address; Affect; Anabolism; Anti-Bacterial Agents; Anticodon; Award; Bacteria; Cell physiology; Cells; Cessation of life; Complex; DNA; DNA Damage; DNA Modification Process; DNA Repair; Data; Deinococcus radiodurans; Disease; Enzymes; Escherichia coli; Eukaryota; Exhibits; Family; Family member; Gene Expression; Genes; Genetic Epistasis; Goals; Homeostasis; Human; Knowledge; Lead; Life; Link; Malignant Neoplasms; Mammals; Messenger RNA; Metabolism; Metals; Methods; Mission; Mitochondria; Modeling; Modification; Nucleic Acids; Nucleoside Q; Organism; Pathogenesis; Pathology; Pathway interactions; Phenotype; Physiological; Physiology; Play; Positioning Attribute; Process; Property; Proteins; RNA; Recruitment Activity; Regulation; Research; Research Support; Resistance; Role; Salmonella; Stress; System; Testing; Transfer RNA; Translations; Work; Yeasts; base; comparative genomics; genetic regulatory protein; in vivo; innovation; mutant; nervous system disorder; novel; paralogous gene; pathogenic bacteria; prevent; protein misfolding; public health relevance; repaired; research study; response; stem

 DESCRIPTION (provided by applicant): The anticodon stem loop (ASL) is critical for decoding properties of tRNA. The universal threonylcarbamoyladenosine (t6A) and the widespread 7-deazaguanosine derivative queuosine (Q) are two ASL modifications at positions 37 and 34, respectively. Our discovery of the biosynthesis pathways for these two complex modifications that had been "missing" for decades has opened the path to study the role of these modifications in vivo. The long-term goal of our research is to expand fundamental knowledge on the synthesis and function of complex tRNA modifications and related molecules, and to understand their roles in core cellular processes and physiology. The current application focuses on the in vivo role of t6A and Q. Because their pathways are complex and draw on primary metabolites and both these modifications have central roles in decoding, t6A and Q are ideal candidates for molecules that integrate metabolism and translation and could play unforeseen regulatory roles. Preliminary results suggest that the absence of t6A triggers an unfolded protein response in both yeast and Bacteria and that t6A affects the translation of specific proteins and this will be the focus of Aim 1. Preliminary phenotypic screens show that Escherichia coli mutants that lack Q have metal sensitivity or resistance phenotypes, and the role of this modification in how cells sense and adapt to metal stresses will be explored in Aim 2. In addition, it has recently been shown that RNA and DNA modifications pathways have much more in common than previously anticipated. Our unexpected discovery that Q precursors are inserted in DNA suggests that paralogs of Q synthesis enzymes have been recruited in DNA metabolism and this will be studied in Aim 3. Aim 4 will focus on the potential role in DNA repair of a paralog of the first enzyme of t6A synthesis. The approach is innovative because comparative genomic methods were used to guide the experimental effort, and this work is revealing new regulatory mechanisms linking metabolism and translation as well totally novel modifications of DNA. The proposed research is significant because it will advance our understanding of the role of critical tRNA modifications and novel DNA modifications in fundamental cellular physiology.

 描述（由申请人提供）：反密码子茎环（ASL）对于tRNA的解码特性至关重要。通用的苏氨酰氨基甲酰基腺苷（t6 A）和广泛分布的7-脱氮鸟苷衍生物鸟苷（Q）分别是37和34位的两种ASL修饰。我们发现了这两种复杂修饰的生物合成途径，这两种修饰已经“缺失”了几十年，这为研究这些修饰在体内的作用开辟了道路。我们研究的长期目标是扩大复杂tRNA修饰和相关分子的合成和功能的基础知识，并了解它们在核心细胞过程和生理学中的作用。本申请集中于t6 A和Q的体内作用。由于它们的途径是复杂的，并且依赖于初级代谢产物，并且这两种修饰在解码中具有核心作用，因此t6 A和Q是整合代谢和翻译的分子的理想候选者，并且可以发挥不可预见的调节作用。初步结果表明，t6 A的缺乏在酵母和细菌中触发未折叠的蛋白质反应，并且t6 A影响特定蛋白质的翻译，这将是目标1的焦点。初步的表型筛选显示，缺乏Q的大肠杆菌突变体具有金属敏感性或抗性表型，这种修饰在细胞如何感知和适应金属胁迫中的作用将在Aim 2中探索。 此外，最近的研究表明，RNA和DNA修饰途径的共同点比以前预期的要多得多。我们意外地发现Q前体插入DNA中，表明Q合成酶的旁系同源物已在DNA代谢中被招募，这将在目标3中进行研究。目的4：研究t6 A合成的第一个酶的一部分在DNA修复中的潜在作用。这种方法是创新的，因为比较基因组学方法被用来指导实验工作，这项工作揭示了连接代谢和翻译的新调控机制以及DNA的全新修饰。这项研究意义重大，因为它将促进我们对关键tRNA修饰和新型DNA修饰在基础细胞生理学中的作用的理解。