CAREER: Shape-based differentiation of RNA elements using small molecules

通过这一奖项，化学部门的生命过程化学项目资助杜克大学的Amanda Hargrove教授，通过新技术的发展来研究调节RNA结构形状的关键差异。长期以来，分子生物学的核心法则一直认为，DNA编码RNA， RNA编码蛋白质，然后蛋白质执行细胞的重要功能；但人们对RNA本身在基础生物学中的功能的认识正在迅速增加。与此同时，迫切需要新的技术来研究这些功能，包括RNA如何与其他分子相互作用。在这项工作中，Hargrove及其同事开发了新技术来满足这种需求，使用基于模式的识别，这类似于人类的味觉，其中少数受体可以根据这些味道的成分如何与受体不同地相互作用来识别各种各样的味道。在这个应用中，小有机分子被用作受体来评估RNA结构的不同“口味”，并快速揭示RNA的形状如何与其功能相对应。该方法将广泛用于未来RNA功能基序分类的研究，小分子受体及其性质的文库将成为一个公开可用和可搜索的数据库。所产生的知识和技术为对调控rna的结构和功能进行新的、根本性的发现奠定了基础，从而形成了影响从环境到人类健康等所有生活领域的分子生物学的中心教条。这些创新技术将差异传感的极限推向了结构生物学，为基于结构的其他生物大分子分类铺平了道路。本研究与PI的教育计划相结合：1)在北卡罗莱纳州不同类型的本科生和高中生中实施基于研究的实验和合作项目；2)通过可视化和应用活动提高学生对非共价相互作用的理解；3)向学生介绍化学和化学生物学的跨学科研究和实际应用；4)提高学生对STEM领域研究的参与度。PI的长期研究目标是建立RNA小分子和蛋白质识别的指导原则，并利用这些原则开发用于RNA结构和功能的化学探针。CAREER的研究目标是利用精细可调的小分子来开发节省时间和简单的技术，阐明RNA识别和相关功能的关键原理。功能性非蛋白质编码RNA （ncRNA）序列的鉴定引发了分子生物学的一场革命，但围绕ncRNA的分子功能仍存在无数问题。这些知识缺口包括ncRNA分子相互作用背后的驱动原理，特别是三维形状的影响，以及这些相互作用如何指导ncRNA依赖过程。ncRNA生物化学的基础研究受到分子表征方面的挑战的阻碍，包括确定3D RNA结构所需的大量时间和专业知识。利用PI实验室最近开发的技术，提出的工作解决了一些基本的分子问题，包括：1)形状如何影响小分子：RNA识别？2)小分子受体可以区分哪些RNA结构元件？3)不同的环境条件如何影响这种分化？4)碱基修饰如何影响RNA识别？5)小分子阵列能否以与蛋白质相当或更好的方式区分功能性RNA元件？这些新颖的，基于小分子的方法来理解RNA识别，为揭示小分子RNA识别的基本原理和RNA形状在调节关键细胞功能中的关键作用提供了化学见解，最终导致ncRNA领域的新知识。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。

With this award, the Chemistry of Life Processes Program in the Chemistry Division is funding Professor Amanda Hargrove of Duke University to investigate critical differences in the shape of regulatory RNA structures through the development of novel technologies. The central dogma of molecular biology has long stated that DNA codes for RNA, which codes for protein, which then carries out a cell's important functions; but there is a rapidly growing appreciation for the functions of RNA itself in fundamental biology. At the same time, there is a pressing need for new techniques to investigate these functions, including how RNA interacts with other molecules. In this work, Hargrove and coworkers develop new technology to meet this need using pattern-based recognition, which is similar to human's sense of taste, wherein a handful of receptors can identify a wide range of flavors based on how the components of those flavors differentially interact with the receptors. In this application, small organic molecules are used as receptors to evaluate different "flavors" of RNA structure and rapidly reveal how the shape of RNA corresponds to its function. The assay developed will be widely accessible for future investigations into the classification of RNA functional motifs, and the library of small molecule receptors and their properties will be a publicly available and searchable database. The knowledge and technology produced provide the basis for novel, fundamental discoveries into the structure and function of regulatory RNAs and thus, the central dogma of molecular biology, which impacts all areas of life, from the environment to human health. These innovative technologies push the limits of differential sensing to include structural biology, paving the way for the structure-based classification of additional biomacromolecules. This research integrates with the educational plans of the PI to: 1) implement research-based experiments and collaborative projects among diverse undergraduate and high school students throughout North Carolina; 2) increase student understanding of noncovalent interactions via accessible visualization and application activities; 3) introduce students to interdisciplinary research and real-world applications of chemistry and chemical biology; and 4) increase student engagement in the study of STEM fields. The long-term research goal of the PI is to establish guiding principles for small molecule and protein recognition of RNA and to use these principles for the development of chemical probes for RNA structure and function. The CAREER research objective is to draw upon exquisitely tunable small molecules to develop timesaving and simple technologies that elucidate critical principles in RNA recognition and related function. The identification of functional yet non-protein coding RNA (ncRNA) sequences has led to a revolution in molecular biology, yet myriad questions surround the molecular function of ncRNA. These knowledge gaps include the driving principles behind ncRNA molecular interactions, particularly the influence of three-dimensional shape, and how these interactions guide ncRNA-dependent processes. Fundamental investigations of ncRNA biochemistry are hindered by challenges in molecular characterization, including the intensive time and expertise required to determine 3D RNA structure. Using techniques recently developed in the lab of the PI, the proposed work addresses a number of fundamental molecular questions, including: 1) How does shape influence small molecule:RNA recognition? 2) What RNA structural elements can small molecule receptors differentiate? 3) How do varying environmental conditions impact this differentiation? 4) How do base modifications influence RNA recognition? 5) Can arrays of small molecules differentiate functional RNA elements in a manner comparable to or better than proteins? These novel, small molecule-based methods for understanding RNA recognition provide ready access to chemical insights that reveal both the fundamental principles of small molecule:RNA recognition and the pivotal role RNA shape plays in regulating key cellular functions, ultimately leading to new knowledge in the ncRNA field.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.