title: Intercellular variation in signaling through the TGF-β pathway and its relation to cell density and cell cycle phase.
abstract: Fundamental open questions in signal transduction remain concerning the sequence and distribution of molecular signaling events among individual cells. In this work, we have characterized the intercellular variability of transforming growth factor β-induced Smad interactions, providing essential information about TGF-β signaling and its dependence on the density of cell populations and the cell cycle phase. By employing the recently developed in situ proximity ligation assay, we investigated the dynamics of interactions and modifications of Smad proteins and their partners under native and physiological conditions. We analyzed the kinetics of assembly of Smad complexes and the influence of cellular environment and relation to mitosis. We report rapid kinetics of formation of Smad complexes, including native Smad2-Smad3-Smad4 trimeric complexes, in a manner influenced by the rate of proteasomal degradation of these proteins, and we found a striking cell to cell variation of signaling complexes. The single-cell analysis of TGF-β signaling in genetically unmodified cells revealed previously unknown aspects of regulation of this pathway, and it provided a basis for analysis of these signaling events to diagnose pathological perturbations in patient samples and to evaluate their susceptibility to drug treatment.
authors.affiliation: Department of Immunology, Genetics and Pathology, Uppsala University, Uppsala, Se-75185 Sweden.
authors.forename: Agata
authors.initials: A
authors.lastname: Zieba
authors.forename: Katerina
authors.initials: K
authors.lastname: Pardali
authors.forename: Ola
authors.initials: O
authors.lastname: Söderberg
authors.forename: Lena
authors.initials: L
authors.lastname: Lindbom
authors.forename: Erik
authors.initials: E
authors.lastname: Nyström
authors.forename: Aristidis
authors.initials: A
authors.lastname: Moustakas
authors.forename: Carl-Henrik
authors.initials: CH
authors.lastname: Heldin
authors.forename: Ulf
authors.initials: U
authors.lastname: Landegren
mesh_terms: D000818:Animals; D002452:Cell Count; D002453:Cell Cycle / Q000187:drug effects*; D002460:Cell Line; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D051379:Mice; D008856:Microscopy, Fluorescence; D046912:Multiprotein Complexes / Q000032:analysis / Q000378:metabolism*; D009928:Organ Specificity; D046988:Proteasome Endopeptidase Complex; D011485:Protein Binding; D020033:Protein Isoforms / Q000032:analysis / Q000378:metabolism; D059748:Proteolysis; D015398:Signal Transduction / Q000187:drug effects*; D059010:Single-Cell Analysis / Q000379:methods; D051785:Smad Proteins / Q000032:analysis / Q000378:metabolism*; D016212:Transforming Growth Factor beta / Q000494:pharmacology*
chemical_list: D046912:Multiprotein Complexes; D020033:Protein Isoforms; D051785:Smad Proteins; D016212:Transforming Growth Factor beta; D046988:Proteasome Endopeptidase Complex