Structure-function relationship between multiple translation initiation factor 3

多个翻译起始因子3之间的结构-功能关系

• 批准号: 8324352
• 负责人: April Dawn Nesbit
• 金额: $ 5.22万
• 依托单位: TRUSTEES OF INDIANA UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-07-21 至 2013-07-20
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8324352
• 关键词: Affect; Animal Model; Antibiotics; Bacteria; C-terminal; Cell Survival; Cell physiology; Color; Complement; Cyanobacterium; Development; Escherichia coli; Eubacterium; Foundations; Gene Expression; Genes; Genetic Screening; Lead; Light; Messenger RNA; N-terminal; Operon; Organism; Peptide Initiation Factors; Post-Transcriptional Regulation; Process; Prokaryotic Cells; Prokaryotic Initiation Factor-3; Proteins; Regulation; Research; Resistance; Ribosomes; Role; Structural Protein; Structure; Structure-Activity Relationship; Transcript; Translating; Translation Initiation; Translational Regulation; Translations; Variant; X-Ray Crystallography; attenuation; cell type; comparative; genome sequencing; genome-wide; improved; innovation; insight; novel; response

DESCRIPTION (provided by applicant): The recent discovery of two translation initiation factor IF3s, IF3a and IF3b, in the cyanobacterium Fremyella diplosiphon provides an exciting long-term opportunity to better understand how these proteins regulate gene expression at the level of transcriptional attenuation and translation. IF3s are an essential component of the translation initiation process, and therefore cell survival, in many bacteria such as Escherichia coli. However, a genetic screen has revealed that IF3a in F. diplosiphon is not essential for viability, and analysis of its sequenced genome uncovered a second gene encoding an IF3 (IF3b). In addition, both IF3a and IF3b complement an E. coli strain lacking IF3, indicating that both are functional IF3 proteins. Furthermore, IF3a regulates the abundance of transcripts from the cpeC operon at the post-transcriptional level in response to changes in ambient light color. To understand the role of these two IF3s in translational regulation, I propose to initially use ribosome profiling of wild-type strains, strains lacking IF3a, and strains lacking IF3b to identify the mRNAs that are being translated in each strain. In addition, I propose to use deep sequencing of the total mRNA in each strain to study the possible roles of IF3a and IF3b in the genome-wide regulation of transcript abundance in this organism. The combined analyses of global translational activity and transcript levels in these strains will provide a comprehensive view of how IF3a and IF3b affect central cellular processes within F. diplosiphon. These functional studies will be complemented by structural studies. The existence of IF3a and IF3b in this organism provides a unique opportunity to conduct a comparative analysis of their structures using X-ray crystallography and/or NMR to relate any structural differences to differences uncovered in the functional analysis. Overall, this research will provide important insights into the relationship between structure and function for this important class of proteins, results that have not been possible to obtain from any other group of eubacteria, which contain a single, essential IF3. Translation is an important cellular process, and the results of my research will expand our understanding the role of the protein IF3 in bacterial translation initiation. Because IF3 is known to be an essential protein in bacteria and has a structure that is distinct from its equivalent eukaryotic protein, called eIF1, the study of how IF3 function relates to its structure has the potential to lead to the development of novel antibiotics, which is important as more bacteria become resistant to known antibiotics.

描述(申请人提供)：最近在蓝藻Fremyella diplosiphon中发现了两个翻译起始因子IF3a和IF3b，这为更好地了解这些蛋白质如何在转录衰减和翻译水平上调节基因表达提供了一个令人兴奋的长期机会。IF3是翻译起始过程的重要组成部分，因此在许多细菌中，如大肠杆菌中，细胞生存。然而，遗传筛查显示，双虹吸虫中的IF3a对生存并不是必不可少的，对其测序基因组的分析发现了第二个编码IF3(IF3b)的基因。此外，IF3a和IF3b都补充了缺乏IF3的大肠杆菌菌株，表明两者都是功能性的IF3蛋白。此外，IF3a在转录后水平上调节CPEC操纵子转录产物的丰度，以响应环境光颜色的变化。为了了解这两个IF3在翻译调控中的作用，我建议首先使用野生型菌株、缺乏IF3a的菌株和缺乏IF3b的菌株的核糖体图谱来鉴定每个菌株中正在翻译的mRNAs。此外，我建议对每个菌株的总mRNA进行深度测序，以研究IF3a和IF3b在该生物体中转录丰度的全基因组调控中的可能作用。对这些菌株的整体翻译活性和转录水平的联合分析将提供关于IF3a和IF3b如何影响双管吸虫体内的中央细胞过程的全面看法。这些功能研究将得到结构研究的补充。该生物体中IF3a和IF3b的存在为利用X射线结晶学和/或核磁共振对其结构进行比较分析提供了一个独特的机会，以将任何结构差异与功能分析中发现的差异联系起来。总体而言，这项研究将为这一重要类别的蛋白质的结构和功能之间的关系提供重要的见解，这是从任何其他真细菌组中无法获得的结果，这些真细菌组包含一个单一的、必要的IF3。翻译是一个重要的细胞过程，我的研究结果将扩大我们对IF3蛋白在细菌翻译起始中的作用的理解。由于IF3是已知的细菌必需蛋白质，其结构不同于同等的真核蛋白质eIF1，因此研究IF3的功能与其结构的关系有可能导致新型抗生素的开发，这一点随着更多的细菌对已知抗生素产生抗药性而变得重要。