Project description:The inflammatory functions of the cytokine tumor necrosis factor (TNF) rely on its ability to induce cytokine production and to induce cell death. Caspase dependent and independent pathways – apoptosis and necroptosis – respectively, regulate immunogenicity by the release of distinct sets of cellular proteins. To obtain an unbiased, systems-level understanding of this important process, we here applied mass spectrometry-based proteomics to dissect protein release during apoptosis and necroptosis. We report hundreds of proteins released from human myeloid cells in time-course experiments. Both cell death types induce receptor shedding, but only apoptotic cells released nucleosome components. Conversely, necroptotic cells release lysosomal components by activating lysosomal exocytosis at early stages of necroptosis- induced membrane permeabilisation and show reduced release of conventionally secreted cytokines.
Project description:Spatial tissue proteomics integrating whole-slide imaging, laser microdissection and ultrasensitive mass spectrometry is a powerful approach to link cellular phenotypes to functional proteome states in (patho)physiology. To be applicable to large patient cohorts and low sample input amounts, including single-cell applications, loss-minimized and streamlined end-to-end workflows are key. We here introduce an automated sample preparation protocol for laser microdissected samples utilizing the cellenONE® robotic system, which has the capacity to process 192 samples in three hours. Following laser microdissection collection directly into the proteoCHIP LF 48 or EVO 96 chip, our optimized protocol facilitates lysis, formalin de-crosslinking and tryptic digest of low-input archival tissue samples. The seamless integration with the Evosep ONE LC system by centrifugation allows ‘on-the-fly’ sample clean-up, particularly pertinent for laser microdissected workflows. We validate our method in human tonsil archival tissue, where we profile proteomes of spatially-defined B-cell, T-cell and epithelial microregions of 4,000 µm2 to a depth of ~2,000 proteins and with high cell type specificity. We finally provide detailed equipment templates and experimental guidelines for broad accessibility.
Project description:Mutational signatures in esophageal squamous cell carcinoma from eight countries of varying incidence – sequence data (Mutographs)
Project description:Mutational signatures in esophageal squamous cell carcinoma from eight countries of varying incidence – patient metatdata (Mutographs)
Project description:In the marine environment, surface-associated bacteria often produce an array of antimicrobial secondary metabolites (MSMs), which have predominantly been perceived as competition molecules. However, they may also affect other hallmarks of surface-associated living, such as motility and biofilm formation. Here, we investigate the ecological significance of an antibiotic secondary metabolite, tropodithietic acid (TDA), in the producing bacterium, Phaeobacter piscinae S26. We constructed a markerless in-frame deletion mutant deficient in TDA biosynthesis wherein TDA production was abolished. Molecular networking demonstrated that other chemical sulphur-containing features, likely related to TDA, were also altered in the secondary metabolome. We found dramatic changes in the physiology of the TDA-deficient mutant, S26�tdaB, compared to the wild type S26. Growth of the two strains were similar; however, S26�tdaB cells were shorter and more motile. Transcriptome and proteome profiling revealed an increase in expression of genes and relative abundance of proteins related to a type IV secretion system, a prophage, and a gene transfer agent (GTA) in S26�tdaB. All these systems may contribute to horizontal gene transfer (HGT), which may facilitate fast adaptation to novel niches. We speculate that, once a TDA-producing population has been established in a new niche, the accumulation of TDA acts as a signal of successful colonization, prompting a switch to a sessile lifestyle. This would lead to a decrease in motility and the rate of HGT, whilst filamentous cells could form the base of a biofilm. In addition, the antibiotic properties of TDA may inhibit invading competing microorganisms.