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Precise Temporal Profiling of Signaling Complexes in Primary Cells Using SWATH Mass Spectrometry


Abstract

Spatiotemporal organization of protein interactions in cell signaling is a fundamental process that drives cellular functions. Given differential protein expression across tissues and developmental stages, the architecture and dynamics of signaling interaction proteomes is, likely, highly context dependent. However, current interaction information has been almost exclusively obtained from transformed cells. In this study, we applied an advanced and robust workflow combining mouse genetics and affinity purification (AP)-SWATH mass spectrometry to profile the dynamics of 53 high-confidence protein interactions in primary T cells, using the scaffold protein GRB2 as a model. The workflow also provided a sufficient level of robustness to pinpoint differential interaction dynamics between two similar, but functionally distinct, primary T cell populations. Altogether, we demonstrated that precise and reproducible quantitative measurements of protein interaction dynamics can be achieved in primary cells isolated from mammalian tissues, allowing resolution of the tissue-specific context of cell-signaling events.

Abstract

Spatiotemporal organization of protein interactions in cell signaling is a fundamental process that drives cellular functions. Given differential protein expression across tissues and developmental stages, the architecture and dynamics of signaling interaction proteomes is, likely, highly context dependent. However, current interaction information has been almost exclusively obtained from transformed cells. In this study, we applied an advanced and robust workflow combining mouse genetics and affinity purification (AP)-SWATH mass spectrometry to profile the dynamics of 53 high-confidence protein interactions in primary T cells, using the scaffold protein GRB2 as a model. The workflow also provided a sufficient level of robustness to pinpoint differential interaction dynamics between two similar, but functionally distinct, primary T cell populations. Altogether, we demonstrated that precise and reproducible quantitative measurements of protein interaction dynamics can be achieved in primary cells isolated from mammalian tissues, allowing resolution of the tissue-specific context of cell-signaling events.

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Additional indexing

Item Type:Journal Article, refereed, original work
Communities & Collections:04 Faculty of Medicine > Functional Genomics Center Zurich
Dewey Decimal Classification:570 Life sciences; biology
610 Medicine & health
Language:English
Date:28 March 2017
Deposited On:26 Jan 2018 12:51
Last Modified:19 Aug 2018 13:16
Publisher:Cell Press (Elsevier)
ISSN:2211-1247
OA Status:Gold
Free access at:PubMed ID. An embargo period may apply.
Publisher DOI:https://doi.org/10.1016/j.celrep.2017.03.019
PubMed ID:28355572
Project Information:
  • : FunderFP7
  • : Grant ID322465
  • : Project TitleINTEGRATE - Integrative biology of T cells and dendritic cells in vivo.
  • : FunderH2020
  • : Grant ID670821
  • : Project TitlePROTEOMICS4D - Proteomics 4D: The proteome in context
  • : FunderFP7
  • : Grant ID115766
  • : Project TitleULTRA-DD - Unrestricted Leveraging of Targets for Research Advancement and Drug Discovery

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