title: 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.
authors.affiliation: Department of Biology, Institute of Molecular Systems Biology, ETH Zurich, 8093 Zurich, Switzerland.
authors.forename: Etienne
authors.initials: E
authors.lastname: Caron
authors.affiliation: Centre d'Immunologie de Marseille-Luminy, Aix Marseille Université, INSERM, CNRS, 13288 Marseille, France.
authors.forename: Romain
authors.initials: R
authors.lastname: Roncagalli
authors.affiliation: The Systems Biology Institute, Tokyo 108-0071, Japan.
authors.forename: Takeshi
authors.initials: T
authors.lastname: Hase
authors.affiliation: Functional Genomics Center Zurich, ETH Zurich and University of Zurich, 8057 Zurich, Switzerland.
authors.forename: Witold E
authors.initials: WE
authors.lastname: Wolski
authors.affiliation: College of Science, College of Computer and Information Science, Northeastern University, Boston, MA 02115, USA.
authors.forename: Meena
authors.initials: M
authors.lastname: Choi
authors.affiliation: Centre d'Immunologie de Marseille-Luminy, Aix Marseille Université, INSERM, CNRS, 13288 Marseille, France.
authors.forename: Marisa G
authors.initials: MG
authors.lastname: Menoita
authors.affiliation: Centre d'Immunologie de Marseille-Luminy, Aix Marseille Université, INSERM, CNRS, 13288 Marseille, France.
authors.forename: Stephane
authors.initials: S
authors.lastname: Durand
authors.affiliation: Centre d'Immunologie de Marseille-Luminy, Aix Marseille Université, INSERM, CNRS, 13288 Marseille, France.
authors.forename: Antonio
authors.initials: A
authors.lastname: García-Blesa
authors.affiliation: Department of Biology, Institute of Molecular Systems Biology, ETH Zurich, 8093 Zurich, Switzerland.
authors.forename: Ivo
authors.initials: I
authors.lastname: Fierro-Monti
authors.affiliation: Department of Biology, Institute of Molecular Systems Biology, ETH Zurich, 8093 Zurich, Switzerland.
authors.forename: Tatjana
authors.initials: T
authors.lastname: Sajic
authors.affiliation: Department of Biology, Institute of Molecular Systems Biology, ETH Zurich, 8093 Zurich, Switzerland.
authors.forename: Moritz
authors.initials: M
authors.lastname: Heusel
authors.affiliation: Department of Neurology and Brain Tumor Center, University Hospital Zurich and University of Zurich, 8091 Zurich, Switzerland.
authors.forename: Tobias
authors.initials: T
authors.lastname: Weiss
authors.affiliation: Centre d'Immunologie de Marseille-Luminy, Aix Marseille Université, INSERM, CNRS, 13288 Marseille, France.
authors.forename: Marie
authors.initials: M
authors.lastname: Malissen
authors.affiliation: Functional Genomics Center Zurich, ETH Zurich and University of Zurich, 8057 Zurich, Switzerland.
authors.forename: Ralph
authors.initials: R
authors.lastname: Schlapbach
authors.affiliation: Department of Biology, Institute of Molecular Systems Biology, ETH Zurich, 8093 Zurich, Switzerland.
authors.forename: Ben C
authors.initials: BC
authors.lastname: Collins
authors.affiliation: The Systems Biology Institute, Tokyo 108-0071, Japan.
authors.forename: Samik
authors.initials: S
authors.lastname: Ghosh
authors.affiliation: The Systems Biology Institute, Tokyo 108-0071, Japan; Okinawa Institute of Science and Technology Garuda School, 904-0495 Okinawa, Japan.
authors.forename: Hiroaki
authors.initials: H
authors.lastname: Kitano
authors.affiliation: Department of Biology, Institute of Molecular Systems Biology, ETH Zurich, 8093 Zurich, Switzerland; Faculty of Science, University of Zurich, 8006 Zurich, Switzerland.
authors.forename: Ruedi
authors.initials: R
authors.lastname: Aebersold
authors.affiliation: Centre d'Immunologie de Marseille-Luminy, Aix Marseille Université, INSERM, CNRS, 13288 Marseille, France; Centre d'Immunophénomique, Aix Marseille Université, INSERM, CNRS, 13288 Marseille, France. Electronic address: bernardm@ciml.univ-mrs.fr.
authors.forename: Bernard
authors.initials: B
authors.lastname: Malissen
authors.affiliation: Department of Biology, Institute of Molecular Systems Biology, ETH Zurich, 8093 Zurich, Switzerland. Electronic address: matthias.gstaiger@imsb.biol.ethz.ch.
authors.forename: Matthias
authors.initials: M
authors.lastname: Gstaiger
mesh_terms: D000818:Animals; D015496:CD4-Positive T-Lymphocytes / Q000378:metabolism; D002454:Cell Differentiation; D002478:Cells, Cultured; D051380:GRB2 Adaptor Protein / Q000378:metabolism; D013058:Mass Spectrometry / Q000379:methods*; D051379:Mice; D025941:Protein Interaction Mapping; D015203:Reproducibility of Results; D015398:Signal Transduction; D013997:Time Factors
chemical_list: D051380:GRB2 Adaptor Protein
keywords: DIA; GRB2; SWATH; interactome; primary T cells; targeted mass spectrometry