A Mass Spectrometry-Based Approach for Mapping Protein Subcellular Localization Reveals the Spatial Proteome of Mouse Primary Neurons

• Published in: Cell reports (Cambridge) Vol. 20; no. 11; pp. 2706 - 2718
• Main Authors: Itzhak, Daniel N., Davies, Colin, Tyanova, Stefka, Mishra, Archana, Williamson, James, Antrobus, Robin, Cox, Jürgen, Weekes, Michael P., Borner, Georg H.H.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 12.09.2017; Cell Press; Elsevier
• Subjects: EGF signaling; label-free quantification; LFQ; neurons; organellar proteomics; primary cells; quantitative mass spectrometry; spatial proteomics; tandem mass tagging; TMT; spatial proteomics; quantitative mass spectrometry; LFQ; primary cells; label-free quantification; EGF signaling; tandem mass tagging; TMT; organellar proteomics; neurons
• ISSN: 2211-1247; 2211-1247
• DOI: 10.1016/j.celrep.2017.08.063

We previously developed a mass spectrometry-based method, dynamic organellar maps, for the determination of protein subcellular localization and identification of translocation events in comparative experiments. The use of metabolic labeling for quantification (stable isotope labeling by amino acids in cell culture [SILAC]) renders the method best suited to cells grown in culture. Here, we have adapted the workflow to both label-free quantification (LFQ) and chemical labeling/multiplexing strategies (tandem mass tagging [TMT]). Both methods are highly effective for the generation of organellar maps and capture of protein translocations. Furthermore, application of label-free organellar mapping to acutely isolated mouse primary neurons provided subcellular localization and copy-number information for over 8,000 proteins, allowing a detailed analysis of organellar organization. Our study extends the scope of dynamic organellar maps to any cell type or tissue and also to high-throughput screening. [Display omitted] •High-resolution organellar maps with label-free quantification (LFQ)•High-throughput organellar maps with TMT-based multiplexing•Deep mapping of EGF-induced protein localization changes with SILAC, LFQ, and TMT•A quantitative spatial proteome from mouse primary neurons Dynamic organellar maps previously provided a proteomic method for capturing protein subcellular localization changes in cultured cells. Itzhak et al. have now adapted the approach to a universal format, extending the method to all cell types. Application to primary mouse neurons provides spatial and quantitative information for more than 8,000 proteins.