Internet-scale discovery of RNA bioengineering rules

描述(申请人提供)：生物基础非编码RNA的最新发现鼓舞着新的抗菌、抗肿瘤和抗病毒疗法，这些疗法可能会使相关分子失效或操纵。然而，在电子折叠模型中，还不能在体外自信地预测RNA的构象，从而减缓了这些潜在的挽救生命的努力的发展。为了制定下一代稳健且经过严格测试的RNA生物工程规则，我们正在探索一种非常规战略：去年，我们发布了一个公民科学项目，使非专家能够在互联网规模的RNA设计比赛中开发和测试折叠假设，这些假设通过湿实验室反馈严格评分。2011年初启动的2.5万人参与的EteRNA项目已经超过了现有的设计在体外正确折叠的新型RNA结构的方法。关键的下一阶段是维持和正式化保存在EteRNA社区内的RNA纳米工程见解。我们建议采取措施，将社区经过实验验证的人类计算整合到一套可供整个RNA纳米工程社区使用的自动化设计算法(EteRNABot)中。除了汇编这样的案例，我们还将通过数千项新的实验，将严格的搜索和解决硅胶和体外设计失败的问题众包出去。最后，我们将利用不断增长的3D构建块数据库和Rosetta RNA折叠/设计算法，为专家和众包三维设计部署EteRNA-3D界面。我们将通过我们的高通量湿法实验室管道对新设计的合成和单核苷酸分辨化学图谱进行评估，以评估每个目标的成功。更广泛地说，我们将通过评估自动化设计工具的使用和引用程度、为该项目生成的公开可用RNA图谱数据库的挖掘和引用程度以及在核酸设计以外的生物医学计算问题中采用互联网规模的科学发现采用EteRNA范式来评估成功与否。 与公共卫生相关：RNA分子在所有生命系统中传递和调节遗传信息方面发挥着基本作用，包括致病细菌、艾滋病毒等逆转录病毒和肿瘤细胞。针对这些RNA的潜在挽救生命的新疗法正受到设计具有新折叠和相互作用的RNA序列的缓慢速度的阻碍。这项拟议的研究旨在解决这一瓶颈，使数以万计的公民科学家能够假设稳健的RNA设计的新规则，并通过高通量RNA合成和化学实验来测试这些想法。

DESCRIPTION (provided by applicant): Recent discoveries of biologically fundamental non-coding RNAs are inspiring novel antibacterial, antitumor, and antiviral therapies that might disable or manipulate the molecules involved. However, in silico folding models cannot yet confidently predict RNA conformations in vitro, slowing the development of these potentially life-saving efforts. To develop the next-generation of robust and rigorously tested RNA bioengineering rules, we are exploring an unconventional strategy: last year, we released a citizen science project enabling non-experts to develop and test folding hypotheses in internet-scale RNA design competitions that are rigorously scored through wet-lab feedback. Launched in early 2011, the 25,000-player EteRNA project already outperforms existing previous methods for designing novel RNA structures which fold properly in vitro. The critical next stage is to sustain and formalize the RNA nanoengineering insights preserved within the EteRNA community. We propose steps to consolidate the community's experimentally validated human computation into a suite of automated design algorithms (EteRNAbot) usable by the entire RNA nanoenginering community. Beyond compiling such cases, we will crowd-source the rigorous search and resolution of in silico and in vitro design failures with thousands of new experiments. Finally, we will deploy the EteRNA- 3D interface for both expert and crowd-sourced three-dimensional design, leveraging a growing database of 3D building blocks and the ROSETTA RNA folding/design algorithms. We will evaluate success in each aim via synthesis and single-nucleotide- resolution chemical mapping of novel designs through our high-throughput wet-lab pipeline. More generally, we will evaluate success by assessing the extent of utilization and citation of the automated design tools; the extent of mining and citation of the publically available RNA Mapping Database generated for the project; and the adoption of the EteRNA paradigm for Internet-scale scientific discovery in biomedical computation problems beyond nucleic acid design. PUBLIC HEALTH RELEVANCE: RNA molecules play fundamental roles in transmitting and regulating genetic information in all living systems, including disease-causing bacteria, retroviruses like HIV, and tumor cells. New potentially life-saving therapies that target these RNAs are being hindered by the slow rate of designing RNA sequences with new folds and interactions. The proposed research seeks to resolve this bottleneck by enabling tens of thousands of citizen scientists to hypothesize new rules for robust RNA design and to test these ideas via high-throughput RNA synthesis and chemical experimentation.