Analyzing HCMV infection and immune evasion by mass spectrometry

通过质谱分析 HCMV 感染和免疫逃避

• 批准号: 421449146
• 负责人: Dr. Andreas Schlosser
• 金额: --
• 依托单位: Rudolf-Virchow-Zentrum - Center for Integrative and Translational Bioimaging
• 依托单位国家: 德国
• 项目类别: Research Units
• 资助国家: 德国
• 项目状态: 未结题
• 来源: https://gepris.dfg.de/gepris/projekt/421449146?language=en
• 关键词: Analyzing; HCMV; infection; immune; evasion

Cytomegaloviruses (CMV) express a variety of immunoevasive factors and manipulate MHC peptide presentation in infected cells in various ways to escape elimination by the immune system. Although several targets of CMV immunoevasins have been identified, it remains largely elusive how immunoevasins modulate the immunopeptidome of the infected cells to suppress the presentation of viral antigens. We will combine mass spectrometry-based immunopeptidomics with metabolic pulse-labeling by isotopically labeled amino acids (pSILAC) to accurately quantify the perturbation of the MHC-I antigen processing pathway by HCMV immunoevasins. First, we will study the effect of ectopic expression of individual immunoevasins on the immunopeptidome of fibroblasts. This kind of experiments will not only enable us to study the suppression of antigen presentation by immunoevasins, but also to identify T-cell epitopes associated with impaired peptide processing (TEIPPs) that might be induced by the action of immunoevasins. Next, we will study the modulation of the immunopeptidome by individual immunoevasins in fibroblasts infected with ΔUS2-11 HCMV-infected fibroblasts. This should reveal how individual immunoevasins suppress the presentation of viral T-cell epitopes. Finally, we will perform quantitative immunopeptidome and proteome experiments with wild type and ΔUS2-11 HCMV-infected fibroblasts. These experiments will be complemented by quantitative proteome analyses of different cell types, such as macrophages and endothelial cells, in collaboration with project P01. Together with project P03, we will conduct quantitative immunopeptidome as well as interactome analyses to explore the function of the immunoevasins US10. In addition, we will characterize the ligandomes of various HLA-C alleles and of HLA E in HCMV-infected cells. In collaboration with project P02, we will determine the immunopeptidome of MCMV-infected fibroblasts and its manipulation by the viral m04 and MATp1 proteins. These studies will provide profound insights in the molecular and cellular function of CMV immunoevasins, and potentially in the long run enable the development of new strategies for therapeutic intervention against the immune evasion of HCMV.

巨细胞病毒（CMV）表达多种免疫逃避因子，并以各种方式操纵MHC肽在感染细胞中的呈现，以逃避免疫系统的消除。尽管已经确定了CMV免疫逃避蛋白的几个靶点，但免疫逃避蛋白如何调节感染细胞的免疫肽丘以抑制病毒抗原的呈递，在很大程度上仍然是难以捉摸的。我们将结合基于质谱的免疫肽组学和代谢脉冲标记的同位素标记氨基酸（pSILAC）来准确量化HCMV免疫球蛋白对MHC-I抗原加工途径的扰动。首先，我们将研究个体免疫球蛋白异位表达对成纤维细胞免疫肽的影响。这种实验不仅使我们能够研究免疫逃避素对抗原呈递的抑制作用，而且还可以鉴定出与免疫逃避素作用可能诱导的受损肽加工（TEIPPs）相关的t细胞表位。接下来，我们将研究个体免疫球蛋白对ΔUS2-11 hcmv感染成纤维细胞中免疫肽的调节。这应该揭示个体免疫逃避素如何抑制病毒t细胞表位的呈现。最后，我们将对野生型和ΔUS2-11感染hcmv的成纤维细胞进行定量免疫肽和蛋白质组实验。这些实验将通过与P01项目合作，对不同细胞类型（如巨噬细胞和内皮细胞）进行定量蛋白质组分析来补充。我们将与project P03一起进行定量免疫肽球和相互作用组分析，探索免疫球蛋白US10的功能。此外，我们将表征hcmv感染细胞中各种HLA- c等位基因和HLA- E的配体。在与P02项目的合作中，我们将确定mcmv感染成纤维细胞的免疫肽球及其被病毒m04和MATp1蛋白操纵的情况。这些研究将为CMV免疫逃避蛋白的分子和细胞功能提供深刻的见解，并有可能在长期内开发针对HCMV免疫逃避的治疗干预新策略。