Development of an OPTogenetic InteractoMics Assay (OPTIMA)

OPTogenic InteractoMics 检测 (OPTIMA) 的开发

• 批准号: 10057519
• 负责人: Michael A Calderwood
• 金额: $ 41.79万
• 依托单位: DANA-FARBER CANCER INST
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-01 至 2023-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10057519
• 关键词: Advanced Development; Area; Automobile Driving; Bar Codes; Benchmarking; Binding; Biological; Biological Assay; Biological Models; Biological Process; Bioluminescence; Biomedical Research; Cells; Clinical Data; Communication; Communities; Complement; Coupled; DNA; Data; Data Set; Detection; Development; Disease; Eukaryotic Cell; Foundations; Functional disorder; Generations; Genes; Genetic; Genetic Recombination; Human; Maps; Mediating; Methods; Molecular Machines; Molecular Structure; Nature; Organism; Pathway interactions; Performance; Phenotype; Protein Interaction Mapping; Protein-Protein Interaction Map; Proteins; Proteome; Reporter; Research; Resources; System; Systems Biology; Technology; Time; Two-Hybrid System Techniques; Validation; Yeasts; base; clinical application; functional genomics; genomic data; human disease; human interactome; innovation; insight; next generation; next generation sequencing; novel; novel strategies; optogenetics; precision medicine; promoter; protein protein interaction; screening; tool; yeast two hybrid system

Project Summary/Abstract Over the last decade, it has become increasingly clear that a more complete understanding of human diseases requires viewing them in the context of systems biology, in particular through a comprehensive understanding of a network of molecular interactions that occur in a cell. Numerous successful applications (e.g. for disease gene prioritization) build on the fact that the local and global structures of molecular networks, mostly based on protein- protein interactions (PPIs), provide critical biological information. With the release of several large-scale systematic PPI datasets since our group produced the first proteome-scale map of the human binary interactome in 2005, integration of genetic and clinical data with interactome information has become possible and provides meaningful and critical insights towards a deeper pathophysiological understanding of diseases and the potential to revolutionize precision medicine. However, we have not reached a comprehensive map of the PPI network in any model system or in humans yet, and thus clinical applications would greatly benefit from deeper and wider explorations of the human interactome. High-throughput approaches have been developed to determine PPIs on a global scale for many organisms, but these assays remain intrinsically limited and labor intensive. A major bottleneck in screening is determining the identities of binary interacting partners. This OPTIMA project aims to eliminate that bottleneck and fill the gap in current networks by developing a novel disruptive technology for high-performance interactomics allowing en masse screening of PPIs to provide comprehensive binary PPI maps. This innovative system will result from the integration of two recently validated technologies, on the one hand the bioluminescent detection of PPIs based on complementation of a split-Nanoluc reporter and, on the other hand, one of the most sensitive optogenetically programmed promoters driving DNA barcode fusion. By leveraging en masse binary PPI detection, this new binary interaction detection strategy will dramatically enhance the overall coverage of proteome-scale interactome maps. This new high-throughput pipeline will be orthogonal to existing proteome-scale binary interaction mapping platforms, such as yeast two-hybrid followed by validation, and thus able to significantly enhance the available tools to expand existing interactomes.

项目摘要/摘要 在过去的十年中，越来越清楚的是，对人类疾病的了解更加完整 需要在系统生物学的背景下查看它们，特别是通过对 细胞中发生的分子相互作用网络。许多成功的应用（例如疾病基因 优先考虑）建立在以下事实的基础上，即分子网络的局部和全球结构，主要基于蛋白质 蛋白质相互作用（PPI）提供关键的生物学信息。随着几个大规模的发布 系统的PPI数据集自我们的小组生产了人类二进制Interactome的第一张蛋白质组规模图 2005年，遗传和临床数据与交互信息的整合变得可能成为可能，并提供 对疾病和潜力更深入的病理生理理解的有意义和批判性见解 彻底改变精度医学。但是，我们尚未达到PPI网络的全面地图 任何模型系统或人类中的任何模型系统，因此临床应用将大大受益于更深和更广泛的 人类互动的探索。 已经开发了高通量方法来确定许多生物的全球范围PPI，但 这些测定在本质上仍然有限和劳动密集型。筛查中的主要瓶颈是确定 二进制互动伙伴的身份。这个Optima项目旨在消除该瓶颈并填补空白 当前网络通过为高性能相互作用的新型破坏性技术开发允许EN的新型破坏性技术 MASSE筛选PPI以提供全面的二进制PPI地图。这个创新的系统将由 一方面，两种最近验证的技术的集成了基于PPI的生物发光检测 在互补的纳入报告者的互补时，另一方面是最敏感的光遗传学之一 驱动DNA条形码融合的程序启动子。通过利用共同的二进制PPI检测，这个新的 二元相互作用检测策略将显着增强蛋白质组规模的整体覆盖率 Interactome图。这条新的高通量管道将与现有的蛋白质组尺度二进制 相互作用映射平台，例如酵母两杂种，然后进行验证，因此能够显着 增强可用工具以扩展现有的相互作用。