Proteomics/Metabolism Core

项目摘要 “蛋白质组学和代谢组学”核心C将由独立的Birgit Schilling博士管理。 巴克研究所的教员核心C将为衰老相关样本提供最先进的分析 这三个项目。Schilling博士也是巴克实验室质谱（MS）设施的主任。 研究所，将直接蛋白质组学和代谢组学核心，并提供所有的基础设施，协调MS 采购和培训所有参与的项目负责人和调查人员。核心的装备非常精良 共有七个现代质谱系统，四个质谱仪专用于 蛋白质组学研究和工作流程。Core C的目标是提供一个通用平台， 项目的设计，执行和分析的实验，涉及分泌的分子，是关键 衰老细胞产生的信号调节剂以及随后对我们的AD模型的影响，包括诱导的 多能干细胞（iPS）和类器官。为了实现本计划项目的目标，核心C将致力于其 i）确定存在于衰老相关分泌表型中的蛋白质 ii）从脑特异性衰老细胞中测定SASP和外来体的组分， 衰老星形胶质细胞和小胶质细胞或衰老样神经元及其对神经元的影响，iii）研究 iv）分析来自AD的iPSC模型的类器官和共培养物， 衰老细胞，以及最后v）确定tau的翻译后修饰的动态变化。芯C 专门创新，开发和实施先进的蛋白质分析技术，包括定量 蛋白质组学，蛋白质分泌体和外泌体的测定，以及监测疾病的PTM状态 蛋白质，以推进基础生物和生物医学研究。为了支持本项目规划小组的项目和目标， Core C将提供各种不同的质谱工作流程，包括依赖数据的采集 (DDA)用于蛋白质发现和鉴定，以及数据独立采集（DIA或SWATH）， SASP和细胞内蛋白质组的全面采样，同时精确相对 量化我们将使用正交四极杆飞行时间质谱仪TripleTOF 6600 和5600（SCIEX）。我们还刚刚订购了一台Orbitrap Eclipse Tribrid（Thermo）质谱仪， 以极高的分辨率、灵敏度和动态范围覆盖蛋白质组， 多路复用能力。我们的专业知识和仪器也特别适合分析SASP 在衰老过程中受到动态影响。连续系统适用性和质量控制试验 获得最高水平的数据再现性和科学严谨性。席林博士的实验室和C区 将为数据分析和生物信息学/统计处理提供所有必要的基础设施和专业知识 的结果。我们将与核心A的生物信息学专家密切互动，进行多组学系统范围的分析。

PROJECT SUMMARY The ‘Proteomics and Metabolomics’ Core C will be under the administration of Dr. Birgit Schilling, an independent faculty member at the Buck Institute. Core C will provide state-of-the-art analysis for senescence-related samples from all three Projects. Dr. Schilling, who is also the Director of the Mass Spectrometry (MS) Facility at the Buck Institute, will direct the Proteomics and Metabolomics Core and provide all infrastructure, coordination of MS acquisitions, and training for all participating project leaders and investigators. The Core is very well-equipped with a total of seven modern mass spectrometric systems, and four mass spectrometers are dedicated to proteomics research and workflows. The objective of Core C is to provide a common platform across each Project for the design, execution and analysis of experiments that involve secreted molecules that are key signaling regulators generated by senescent cells and subsequent effects on our AD models, including induced pluripotent stem cells (iPS) and organoids. To achieve the goals of this Program Project, Core C will devote its efforts to the following work: i) determine proteins present in the senescence-associated secretory phenotype (SASP) from brain specific senescent cells, ii) determine the components of SASP and exosomes from senescent astrocytes and microglia or senescent-like neurons and their effects on neurons, iii) investigate organoids and co-cultures from iPSC models of AD, iv) analyze metabolites and measure NAD levels of senescent cells, and finally v) determine dynamic changes of post-translational modifications of tau. Core C specifically innovates, develops, and implements advanced protein analytical technologies, including quantitative proteomics, determination of protein secretomes and exosomes, as well as monitoring the PTM status of disease proteins, to advance basic biological and biomedical research. To support the projects and aims of this PPG, Core C will offer a variety of different mass spectrometric workflows, including data-dependent acquisitions (DDA) for protein discovery and identification, and data-independent acquisitions (DIA or SWATH) for comprehensive sampling of the SASP and intracellular proteomes with simultaneous accurate relative quantification. We will use our orthogonal Quadrupole Time-of-Flight mass spectrometers, the TripleTOF 6600 and 5600 (SCIEX). We also just ordered an Orbitrap Eclipse Tribrid (Thermo) mass spectrometer enabling very deep proteome coverage at very high resolution, sensitivity, dynamic range and also offering excellent multiplexing capabilities. Our expertise and instrumentation are also particularly well suited for analysis of SASP that may be dynamically affected during senescence. Continuous system suitability and quality control assays are acquired to provide highest levels of data reproducibility and scientific rigor. Dr. Schilling’s lab and Core C will provide all necessary infrastructure and expertise for data analysis and bioinformatic/statistical processing of the results. We will closely interact with bioinformatics experts in Core A for multi-omics system-wide analysis.