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Title (Primary) Integration of conventional quantitative and phospho-proteomics reveals new elements in activated Jurkat T-cell receptor pathway maintenance
Author Jouy, F.; Müller, S.A.; Wagner, J.; Otto, W.; von Bergen, M.; Tomm, J.M.;
Journal Proteomics
Year 2015
Department PROTEOM;
Volume 15
Issue 1
Language englisch;
POF III (all) F11;
Keywords Jurkat T-cell; Kinase substrate identification; Phosphoproteomics; Quantitative proteomics; Technology
UFZ wide themes RU3;
Abstract Recent years have seen a constant development of tools for the global assessment of phosphoproteins. Here, we outline a concept for integrating approaches for quantitative proteomics and phosphoproteomics. The strategy was applied to the analysis of changes in signalling and protein synthesis occurring after activation of the T-cell receptor (TCR) pathway in a T-cell line (Jurkat cells). For this purpose, peptides were obtained from four biological replicates of activated and control Jurkat T-cells and phosphopeptides enriched via a TiO2-based chromatographic step. Both phosphopeptide-enriched and flow-through fractions were analyzed by LC–MS. We observed 1314 phosphopeptides in the enriched fraction whereas 19 were detected in the flow-through, enabling the quantification of 414 and eight phosphoproteins in the respective fractions. Pathway analysis revealed the differential regulation of many metabolic pathways. Among the quantified proteins, 11 kinases with known TCR-related function were detected. A kinase-substrate database search for the phosphosites identified also confirmed the activity of a further ten kinases. In total, these two approaches provided evidence of 19 unique TCR-related kinases. The combination of phosphoproteomics and conventional quantitative shotgun analysis leads to a more comprehensive assessment of the signalling networks needed for the maintenance of the activated status of Jurkat T-cells.
ID 15706
Persistent UFZ Identifier https://www.ufz.de/index.php?en=20939&ufzPublicationIdentifier=15706
Jouy, F., Müller, S.A., Wagner, J., Otto, W., von Bergen, M., Tomm, J.M. (2015):
Integration of conventional quantitative and phospho-proteomics reveals new elements in activated Jurkat T-cell receptor pathway maintenance
Proteomics 15 (1), 25 - 33