EMERGE Episode 2: Spatial-Temporal (Phospho)proteomics

Ana's presentation focuses on the application of high-throughput spatial-temporal proteomics workflows for the study of phospho-signaling dynamics. Dynamic changes in subcellular localization of signaling proteins is a general concept that eukaryotic cells evolved for eliciting a coordinated response to stimuli. Mass spectrometry-based proteomics in combination with subcellular fractionation can provide comprehensive maps of spatio-temporal regulation of cells, but involves laborious methods that did not include phospho-proteome analysis. The presentation includes a high-throughput workflow, based on sequential cell fractionation, to profile the global proteome and phospho-proteome dynamics across six distinct subcellular fractions. The workflow was benchmarked by studying spatio-temporal EGFR phospho-signaling dynamics in-vitro (HeLa cell cultures) and in-vivo in mouse tissues, and was also employed to assess the spatio-temporal events in response to stress related signaling, such as hyperosmotic shock, revealing cellular relocation of ribosomal proteins.