CAREER: Uncovering Quantitative Design Principles of RNA Regulators For Synthetic Biology

非技术描述：细胞有惊人的能力来处理信息，做出决定，并改变自己的状态，以应对不断变化的环境。这种能力编码在细胞DNA基因组中，通过基因表达的基本过程转化为RNA和蛋白质分子。在这些RNA和蛋白质发挥的众多功能中，有一项是调节自己的表达。事实上，现在已知RNA可以调控基因表达的几乎所有方面，并在控制生命的一些最基本的过程中发挥核心作用。生物技术和合成生物学中的一个关键问题是：如何设计RNA分子来控制目标基因在细胞内的表达，以促进从将细胞用作化工厂到将它们用作环境传感器的各种应用？与许多生物分子一样，RNA的功能与其结构密切相关。在这项工作中，这位研究人员建立在他开发的一种新的小合成RNA机制的基础上，称为小转录激活RNA，或STAR。假设STAR可以构建独特的基因表达控制技术，因为它们代表了sRNA的一种全新功能。该项目寻求将STAR作为一个试验台，以揭示设计RNA分子以精确控制细胞内基因表达的原理。这项工作将有助于实现RNA作为一种强大的细胞工程底物的潜力。它还将为我们科学理解RNA在自然生物系统中的核心作用所依据的基本序列/结构/功能关系提供新的线索。因此，预计开展的研究和研究培训活动将对社会产生广泛影响，包括细胞基因调控科学和可与广泛的生物技术应用直接相关的核糖核酸基因调控工程科学。该项目还将培养下一代训练有素的合成生物学研究生和教师，他们将被介绍给生物技术研究的广泛、跨学科性质。此外，该计划将积极参与更广泛的社区，帮助创造一个有能力对合成生物学的未来做出重要决定的知情公众。技术描述：该项目的总体目标是建立一个综合的研究和教育计划，专注于揭示将小RNA序列、结构和功能联系起来的量化设计原理，并利用这些原理设计能够精确调控基因表达的合成RNA。该教育计划的重点是将研究整合到下一代合成生物学学生和教师的培训中，并向更广泛的公众宣传和激发合成生物学。反式细菌小RNA(SRNA)通过与靶信使RNA(MRNAs)的直接RNA-RNA相互作用来实施调控，从而导致靶标的结构变化。这些变化反过来调节基因表达的许多方面，包括转录、翻译和mRNA降解。该项目的中心假设是：1)可以发现定量的sRNA结构/功能设计原理，并将其用于合理优化和扩展RNA调节器的功能；2)可以发现电路级设计规则，并将其用于设计新的合成RNA遗传模块，以控制基因表达的时间和模式。这个职业项目的中心目标是使用小转录激活RNA(STAR)作为试验台，揭示合成生物学中RNA调节剂的定量设计原理。这将使用多方面的方法来实现，其中包括使用尖端的RNA结构测量技术来阐明恒星的分子水平设计原则。一旦学习了分子层面的原理，该项目将专注于使用这些技术来学习将恒星整合到决策监管网络中的设计规则。此外，可以对STAR介导的基因表达动力学进行定量建模的新实验和计算工具将被用于了解STAR功能的不同方面如何通过RNA网络传播。这些研究的成功完成将推动建立生物设计的量化学科的更广泛目标，该学科可用于对细胞系统进行编程，以解决可持续能源和生物制造方面的重要问题。此外，这项工作可能为了解自然发生的生物系统的天然小RNA调节机制提供洞察。教育活动包括制定长期计划，继续举办冷泉港合成生物学暑期课程，作为全球范围内广泛学生和教师的培训中心；开发用于培训未来几代合成生物学家的量化课程材料；以及开展针对有学龄儿童的家庭的动手活动，以激发和告知他们关于合成生物学的知识。

Nontechnical Description: Cells have an amazing ability to process information, make decisions, and change their state in response to changing environments. This ability is encoded within the cellular DNA genome, which is converted into RNA and protein molecules through the basic processes of gene expression. Among the many functions these RNAs and proteins perform is regulating their own expression. In fact RNAs are now known to regulate almost all aspects of gene expression, and play central roles in controlling some of life's most basic processes. A key question in biotechnology and synthetic biology is then: How can RNA molecules be designed to control the expression of target genes inside cells to facilitate applications ranging from using cells as chemical factories all the way to using them as environmental sensors? As with many biomolecules, RNA function is intimately related to its structure. In this work, the investigator builds off of his development of a new small synthetic RNA mechanism called Small Transcription Activating RNAs, or STARs. STARs are hypothesized to allow the construction of unique gene expression control techniques since they represent a brand new function for sRNAs. This project seeks to use STARs as a test-bed for uncovering the principles for engineering RNA molecules to precisely control gene expression inside cells. This work will help realize the potential of RNA as a powerful substrate for cellular engineering. It will also shed new light on our scientific understanding of the fundamental sequence/structure/function relationship that underlies RNA's central role in natural biological systems. The projected studies and research training activities are thus expected to have a broad impact on society, ranging from the science of cellular gene regulation and the engineering science of RNA gene regulation that can directly connect to a broad array of biotechnological applications. This project will also cultivate the next generation of highly trained graduate students and teachers of synthetic biology who will be introduced to the broad, interdisciplinary nature of biotechnology research. Moreover, this program will actively engage the broader community to help create an informed public that is equipped to make important decisions about the future of synthetic biology.Technical Description: The overall goal of this project is to build an integrated research and education program focused on uncovering quantitative design principles that link small RNA sequence, structure, and function, and to use these principles to design synthetic RNAs that can precisely regulate gene expression. The education plan focuses on integrating research into the training of the next generation of synthetic biology students and teachers, and informing and exciting the broader public about synthetic biology.Trans-acting bacterial small RNAs (sRNA) exert regulation via direct RNA-RNA interactions with target messenger RNAs (mRNAs) that cause structural changes in the target. These changes in turn regulate many aspects of gene expression including transcription, translation and mRNA degradation. The central hypotheses of this project are that: 1) quantitative sRNA structure/function design principles can be discovered and used to rationally optimize and expand the functionality of RNA regulators, and 2) circuit-level design rules can be discovered and used to engineer new synthetic RNA genetic modules that control the timing and pattern of gene expression. The central goal of this CAREER project is to use Small Transcription Activating RNAs (STARs) as a test-bed for uncovering the quantitative design principles of RNA regulators for synthetic biology. This will be pursued using a multi-faceted approach that includes using cutting-edge RNA structure measurement technologies to elucidate the molecular level design principles of STARs. Once molecular-level principles are learned, the project will focus on using these techniques to learn the design rules for integrating STARs into decision making regulatory networks. In addition, new experiments and computational tools that can quantitatively model the kinetics of gene expression mediated by STARs will be used to understand how different aspects of STAR function propagate through RNA networks. Successful completion of these studies will forward the broader goal of creating a quantitative discipline of biological design that can be used to program cellular systems to solve important problems in sustainable energy, and biomanufacturing. In addition, the work may provide insight to mechanisms of native small RNA regulation of naturally occurring biological systems.Educational activities include developing a long term plan for the continuation of the Cold Spring Harbor Synthetic Biology summer course as a training center for a broad range of students and teachers across the globe; developing quantitative curricular materials for training future generations of synthetic biologists; and performing hands-on activities aimed towards families with school-age children to excite and inform them about synthetic biology.