Collaborative Research: Uncovering How Riboswitches Exploit Out-of-Equilibrium RNA Folding Pathways to Make Genetic Decisions

合作研究：揭示核糖开关如何利用非平衡 RNA 折叠途径做出遗传决策

• 批准号: 1914567
• 负责人: Julius Lucks
• 金额: $ 35.16万
• 依托单位: Northwestern University
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-08-15 至 2024-03-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1914567&HistoricalAwards=false
• 关键词: Collaborative; Research; Uncovering; How; Riboswitches; Collaborative; Research; Uncovering; How; Riboswitches

This work will uncover new principles by which riboswitch RNAs exploit out-of-equilibrium RNA folding and ligand binding to make genetic decisions. The tools developed will also be applicable to understanding the role of co-transcriptional folding and assembly of ribozymes, regulatory RNA structures within non-coding and coding RNAs, and assembly of RNA-protein complexes such as the ribosome and spliceosome. These studies will also contribute understanding of the physical principles of out-of- equilibrium co-transcriptional RNA folding, help address long-standing questions about how dynamic RNA structures coordinate genetic processes, and shed light on how natural RNAs exploit out-of- equilibrium mechanisms to efficiently fold on extremely rugged free-energy landscapes. Since co-transcriptional RNA folding happens every time an RNA is synthesized, the broader impacts of this research include developing general principles and techniques that can be used to understand a wide array of fundamental cellular processes from gene expression to regulation. The study of riboswitches also has several broader impacts towards societal goals, since they can be used as biosensors within new molecular diagnostics, and they are important targets for new classes of antibiotics. Broader impacts of integrated research and education will come from a multi-pronged plan including conducting demonstrations of riboswitch diagnostics to school-age groups, mentorship of undergraduate researchers, and delivering hands-on tutorials of computational RNA folding approaches to broader scientific communities.The overarching goals of this proposal are to: (i) Uncover detailed mechanisms of how ligand binding bifurcates out-of-equilibrium RNA cotranscriptional folding pathways to enact genetic decisions in riboswitch RNAs; and (ii) Develop and apply new hybrid experimental-computational frameworks that can reconstruct RNA cotranscriptional folding pathways at the secondary and tertiary structure levels. The education plan focuses on integrating this research into hands-on demonstration activities targeted towards school age children, undergraduate researcher mentorship, and hands-on training tutorials for the broader scientific community. The post-genomic era has ushered in a new appreciation that RNAs play central roles in regulating, maintaining and defending the genomes of all organisms. However, a critical knowledge gap remains: we have relatively little understanding of the dynamic folding pathways that RNAs undergo as they are being synthesized during transcription, thus hindering our fundamental understanding of how RNA structures enact critical cellular functions such as catalysis, gene expression regulation, and cellular sensing. To address this gap, the PIs recently innovated and validated a hybrid experimental-computational approach that uses high-throughput RNA structure chemical probing data with computational algorithms to generate two and three-dimensional models of RNA cotranscriptional folding pathways. One central objective of this proposal is to extend this approach to incorporate more complex RNA structures and interactions such as pseudoknots relevant to a broad range of functional cellular RNAs. The second is to uncover biophysical principles of how out-of-equilibrium RNA fluctuations during cotranscriptional folding influence RNA function. The latter will be pursued through the use of riboswitch RNAs as model systems, which make ligand-mediated genetic decisions, use the dynamic formation of broadly utilized RNA structures to do so, and have broader impact relevance for fundamental biology and biotechnologies.This project is being jointly supported by the Physics of Living Systems program in the Division of Physics and the Molecular Biophysics program in the Division of Molecular and Cellular Biosciences.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.

这项工作将发现新的原理，核糖开关RNA利用不平衡的RNA折叠和配体结合以做出遗传决策。开发的工具还适用于了解核酶的共同折叠和组装，非编码和编码RNA中的调节RNA结构以及RNA蛋白复合物（例如核糖体和剪接体）的组装。这些研究还将有助于理解不平衡的共同转录RNA折叠的物理原理，有助于解决有关动态RNA结构如何协调遗传过程的长期问题，并阐明了自然RNA如何利用过平衡机制，以在极度蓄积的自由型自由式的景观上有效地折叠。由于每次合成RNA时都会发生共转录RNA折叠，因此这项研究的更广泛的影响包括开发一般原理和技术，可用于了解从基因表达到调节的广泛基本细胞过程。对核糖开关的研究也对社会目标产生了更广泛的影响，因为它们可以用作新分子诊断中的生物传感器，并且它们是新型抗生素类别的重要目标。综合研究和教育的更广泛的影响将来自一个多管齐下的计划，包括向学院群体进行核糖开关诊断，对本科研究人员的指导以及提供计算RNA折叠方法的动手教程，以实现更广泛的科学社区。 RNA共转录折叠途径在核糖开关RNA中制定遗传决策； （ii）开发和应用新的混合实验计算框架，可以在次级和第三纪结构水平上重建RNA共转录折叠途径。该教育计划的重点是将这项研究纳入针对学龄儿童，本科研究人员指导以及为更广泛的科学界的动手培训教程的动手演示活动。后的时代已经引发了一种新的欣赏，即RNA在调节，维持和捍卫所有生物的基因组中起着核心作用。但是，临界知识差距仍然存在：我们对RNA在转录过程中的合成时经历的动态折叠途径的了解相对较少，从而阻碍了我们对RNA结构如何实现关键细胞功能（例如催化，基因表达调节和细胞传感）的基本了解。为了解决这一差距，PI最近创新并验证了一种混合实验计算方法，该方法使用了使用计算算法的高通量RNA结构化学探测数据来生成RNA共表征折叠途径的两个和三维模型。该提案的一个核心目的是扩展这种方法，以结合更复杂的RNA结构和相互作用，例如与广泛的功能性细胞RNA相关的伪诺。第二个是发现在共转录折叠过程中如何平衡的RNA波动影响RNA功能的生物物理原理。后者将通过使用核糖开关RNA作为模型系统来追求，从而做出配体介导的遗传决策，使用广泛使用的RNA结构的动态形成，并具有更广泛的影响，并且对基本生物学和生物技术的基本生物学和生物技术具有更广泛的影响。这些项目在生物学方面和摩尔克式计划中都在物理学方面和分子的分子计划，并具有分子的分子计划，该奖项反映了NSF的法定任务，并被认为是值得通过基金会的智力优点和更广泛的影响审查标准的评估来支持的。