Structural Interactomics of virus host relationships

病毒宿主关系的结构相互作用组学

• 批准号: 454970352
• 负责人: Dr.-Ing. Boris Bogdanow
• 金额: --
• 依托单位: Arbeitsgruppe Structural Interactomics
• 依托单位国家: 德国
• 项目类别: WBP Position
• 财政年份: 2021
• 资助国家: 德国
• 起止时间: 2020-12-31 至 2022-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/454970352?language=en
• 关键词: Structural; Interactomics; virus; host; relationships

Protein-protein interactions (PPIs) facilitate essential processes during viral infections. Despite their significant importance, a method to globally study host-virus PPIs and corresponding interaction interfaces from intact infected cells is lacking. I am here planning to address this shortcoming by using cross-linking reagents on infected cells followed by mass-spectrometry of cross-linked peptides. I will combine this method with pulse labeling using biorthogonal amino acids, which increases the sensitivity of the approach towards detecting interactions of viral proteins with host proteins. As a model system, I chose Herpes-simplex virus type 1 due to its medical relevance, complex proteome, numerous interactions with host cellular proteins and its ability to induce shutoff of host cellular protein synthesis.The method I am proposing has numerous advantages compared to current approaches to systematically study virus-host interactions. First, since crosslinks are only formed between proteins that are in proximity, identified crosslinks hint towards interaction interfaces. Second, the use of membrane permissive crosslinking reagents allows to capture interactions in intact cells. Third, the approach works with genetically unaltered viruses and captures interactions of proteins in their native cellular environment. The overarching focus of the presented research proposal is to establish a valuable method for the systematic discovery of direct PPIs based on combining bio-orthogonal labeling and XL-MS. My preliminary data revealed an extensive interaction network involving hundreds of host and virus proteins based on thousands of crosslinks. I will build on these preliminary data and aim to further increase the depth and sensitivity of detecting PPIs in order to achieve near-comprehensive coverage of virus-host interactions. Further, I aim to assess how virus-host and virus-virus interactions dynamically change in the infected cell. Therefore, I will establish a proteomic and bioinformatics pipeline, allowing me to quantitatively capture the dynamics of PPIs during HSV-1 infection. Novel interactions will be validated and the functional importance of notable interactions will be demonstrated. The presented project thus integrates crosslinking mass-spectrometry with functional studies and integrative bioinformatics analyses. The results will deliver unmatched insights into the structural complexity of virus-host protein interactions for an important human pathogen.

蛋白质-蛋白质相互作用(PPI)促进了病毒感染过程中的基本过程。尽管它们非常重要，但缺乏一种从完整的感染细胞中全局研究宿主病毒PPI和相应的相互作用界面的方法。我在这里计划通过在感染细胞上使用交联剂，然后对交联肽进行质谱分析来解决这一缺点。我将把这种方法与使用双正交氨基酸的脉冲标记结合起来，这增加了检测病毒蛋白与宿主蛋白相互作用的方法的灵敏度。作为模型系统，我选择了单纯疱疹病毒1型，因为它与医学相关，复杂的蛋白质组，与宿主细胞蛋白的大量相互作用，以及它诱导宿主细胞蛋白质合成关闭的能力。与目前系统研究病毒与宿主相互作用的方法相比，我提出的方法具有许多优势。首先，由于交联键只在邻近的蛋白质之间形成，识别出的交联键提示相互作用界面。其次，膜许可交联剂的使用允许捕获完整细胞中的相互作用。第三，该方法适用于基因未改变的病毒，并捕捉到蛋白质在其天然细胞环境中的相互作用。提出的研究建议的重点是建立一种基于生物正交标记和XL-MS相结合的系统发现直接PPI的有价值的方法。我的初步数据显示了一个广泛的相互作用网络，涉及数以千计的基于数千个交联链的宿主和病毒蛋白。我将以这些初步数据为基础，旨在进一步提高检测PPI的深度和灵敏度，以实现对病毒-宿主相互作用的近乎全面的覆盖。此外，我的目标是评估病毒-宿主和病毒-病毒相互作用如何在受感染的细胞中动态变化。因此，我将建立一个蛋白质组和生物信息学流水线，使我能够定量地捕捉HSV-1感染期间PPI的动态。新的相互作用将得到验证，显着相互作用的功能重要性将得到证明。因此，本项目将交联质谱学与功能研究和综合生物信息学分析相结合。这一结果将对一种重要的人类病原体的病毒-宿主蛋白相互作用的结构复杂性提供无与伦比的见解。