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.

项目总结/摘要 在过去的十年里，人们越来越清楚地认识到， 需要在系统生物学的背景下观察它们，特别是通过全面了解 细胞中分子间相互作用的网络。许多成功的应用（例如疾病基因 优先级）建立在分子网络的局部和全局结构的事实上，主要基于蛋白质- 蛋白质相互作用（PPI）提供了重要的生物信息。随着几次大规模的 自从我们的小组制作了第一个人类二元相互作用组的蛋白质组规模图以来， 2005年，遗传和临床数据与相互作用组信息的整合已经成为可能， 有意义的和关键的见解，对疾病和潜在的更深入的病理生理学理解 来革新精准医疗然而，我们还没有得到一个全面的PPI网络图， 任何模型系统或在人类中，因此临床应用将大大受益于更深入和更广泛的 人类互动组的探索 已经开发了高通量方法来确定全球范围内许多生物体的PPI，但 这些测定仍然固有地受限并且劳动密集。筛选的一个主要瓶颈是确定 二元互动伙伴的身份。OPTIMA项目旨在消除这一瓶颈，填补以下方面的差距： 通过开发一种用于高性能交互组学的新型颠覆性技术， PPI的快速筛选，以提供全面的二进制PPI图。这一创新系统将产生于 整合了两种最近验证的技术，一方面是基于PPI的生物发光检测， 在互补的分裂Nanoluc报告，另一方面，最敏感的光遗传学之一， 驱动DNA条形码融合的程序化启动子。通过利用增强的二进制PPI检测， 二进制相互作用检测策略将大大提高蛋白质组规模的整体覆盖率 相互作用基因组图谱这种新的高通量流水线将与现有的蛋白质组规模二进制 相互作用作图平台，如酵母双杂交，然后进行验证，因此能够显着 加强现有工具，以扩大现有的互动机制。