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 exper...