ABI INNOVATION: Physical Bioinformatics Tools for Measuring Translation Rates from Next-Generation Sequencing Data

翻译是信使RNA中编码的基因组信息转化为蛋白质分子的过程。与翻译相关的速率决定了蛋白质合成的时间尺度，影响细胞中蛋白质分子的丰度，并且最近被证明影响所产生的蛋白质的结构和功能。这些速率很难在生物体的转录组中测量。该项目的目的是开发和应用物理生物信息学分析工具，从下一代测序技术核糖体分析（Ribo-Seq）产生的实验数据中测量翻译起始率和延伸率。物理生物信息学为生物信息学领域带来了物理科学的视角，并允许从大型生物数据集中提取比以前可能的更多信息。新的分析方法将被创造出来，从Ribo-Seq和其他数据中测量翻译起始和延伸的绝对速率，使实验学家能够测量这些数量。该项目将通过确定延伸期在全局和单基因水平上对翻译控制的贡献，以及进化编码的翻译-延伸动力学的变化是否调节伴侣结合，来推进分子生物学的知识。将举办网络研讨会和讲习班，向生物信息学家介绍该项目开发的方法的理论基础，运行分析管道，并解释结果-从而加强研究基础设施。这些事件将被记录并公开存档，以供社区将来使用。作为服务不足的K-12社区的一部分，该项目将通过宾夕法尼亚州立大学的向上数学和科学项目向少数民族高中生介绍生物信息学的科学研究，该项目安排这些学生在夏天从事大学研究经验。翻译起始率和延伸率很难在生物体的转录组中测量。该项目的目的是开发和应用扎根于化学和物理领域的物理生物信息学分析方法，从下一代测序技术Ribo-Seq生成的实验数据中测量翻译起始和延伸率。可以肯定的是，文献中已经报道了利用启发式或基于模拟的方法对这些比率进行定性测量。然而，这些方法通常不能产生绝对的比率，也不能保证提供最优的解决方案。了解绝对速率是必要的，因为它们是系统中的实际速率，通过了解它们，可以确定它们对其他共平移过程的影响。翻译涉及分子和化学反应，这意味着符合化学和物理定律的分析方法更有可能产生准确的结果。利用理论化学、物理和生物信息学的跨学科专业知识，PI实验室将在该项目中开发方法：(1)在核糖核酸测序实验产生的核糖体保护片段上最佳地确定核糖体a位点的位置-这是测量绝对速率的第一步；(2)通过将翻译视为一种离散的流体流动形式，测量核糖体径流实验中密码子的平均翻译率；(3)通过稳态Ribo-Seq实验测量个体密码子翻译率；(4)通过结合Ribo-Seq、RNA-Seq和多体分析数据来测量起始率。这些方法将用于回答基本的生物学问题：在翻译的起始和延伸阶段，基因表达的翻译控制在多大程度上是确定的？与伴侣结合相关的进化编码翻译-延伸动力学是否会对伴侣结合产生因果影响？这些问题的答案将提供关于基因表达如何在翻译阶段被调节的基本问题的见解，并可能为某些疾病中基因表达出错时的治疗开辟新的途径。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

Translation is the process by which the genomic information encoded in a messenger RNA is converted into a protein molecule. The rates associated with translation determine the time scales of protein synthesis, influence the abundance of protein molecules in cells, and have recently been shown to influence the structure and function of the protein produced. These rates are difficult to measure across the transcriptomes of organisms. The purpose of this project is to develop and apply physical bioinformatics analysis tools to measure translation-initiation and -elongation rates from experimental data generated with the next-generation sequencing technique Ribosome Profiling (Ribo-Seq). Physical Bioinformatics brings a physical-science perspective to the field of bioinformatics and permits more information to be extracted from big biological data sets than previously possible. New analysis methods will be created to measure absolute rates of translation initiation and elongation from Ribo-Seq and other data, making it possible for experimentalists to measure these quantities. This project will advance knowledge in molecular biology by determining the contribution of the elongation phase to translation control at both the global and single-gene level and whether changes in evolutionarily encoded translation-elongation kinetics modulates chaperone binding. Webinars and workshops will be held to introduce bioinformaticians to the theory underlying the methods developed in this project, running the analysis pipelines, and interpreting the results - thereby enhancing research infrastructure. These events will be recorded and publicly archived for future use by the community. As part of outreach to underserved K-12 communities, this project will introduce minority high school students to scientific research in Bioinformatics through Penn State's Upward Bound Math and Science program, which arranges for these students to engage in university research experience over the summer.Translation initiation and elongation rates are difficult to measure across the transcriptomes of organisms. The purpose of this project is to develop and apply physical bioinformatics analysis methods, rooted in the fields of chemistry and physics, to measure translation-initiation and -elongation rates from experimental data generated with the Next-Generation Sequencing technique Ribo-Seq. To be sure, qualitative measures of these rates utilizing heuristic or simulation-based approaches have been reported in the literature. However, these methods often do not yield absolute rates nor are they guaranteed to provide optimal solutions. Knowledge of absolute rates is essential because they are the actual rates in the system, and by knowing them their impact on other co-translational processes can be determined. Translation involves molecules and chemical reactions, meaning that analysis methods consistent with the laws of chemistry and physics are more likely to yield accurate results. Utilizing the interdisciplinary expertise in theoretical chemistry, physics, and bioinformatics, the PI's lab will develop methods in this project that: (1) optimally identify the location of the ribosome's A-site on ribosome-protected fragments generated from Ribo-Seq experiments - the first step towards measuring absolute rates; (2) measure the average codon translation rates from ribosome run-off experiments by treating translation as a form of discretized fluid flow; (3) measure individual codon translation rates from steady-state Ribo-Seq experiments; (4) measure initiation rates through a combination of Ribo-Seq, RNA-Seq and polysome profiling data. The methods will be applied to answer fundamental biological questions: To what extent is translation control of gene expression determined in the initiation versus elongation phase of translation? Do evolutionarily encoded translation-elongation kinetics that correlate with chaperone binding causally influence chaperone binding? Answers to these questions will provide insight into fundamental issues concerning how gene expression is regulated at the stage of translation, and may open new avenues for treatment when gene expression goes awry in some diseases.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.