title: Single-Molecule Imaging Reveals the Activation Dynamics of Intracellular Protein Smad3 on Cell Membrane.
abstract: Smad3 is an intracellular protein that plays a key role in propagating transforming growth factor β (TGF-β) signals from cell membrane to nucleus. However whether the transient process of Smad3 activation occurs on cell membrane and how it is regulated remains elusive. Using advanced live-cell single-molecule fluorescence microscopy to image and track fluorescent protein-labeled Smad3, we observed and quantified, for the first time, the dynamics of individual Smad3 molecules docking to and activation on the cell membrane. It was found that Smad3 docked to cell membrane in both unstimulated and stimulated cells, but with different diffusion rates and dissociation kinetics. The change in its membrane docking dynamics can be used to study the activation of Smad3. Our results reveal that Smad3 binds with type I TGF-β receptor (TRI) even in unstimulated cells. Its activation is regulated by TRI phosphorylation but independent of receptor endocytosis. This study offers new information on TGF-β/Smad signaling, as well as a new approach to investigate the activation of intracellular signaling proteins for a better understanding of their functions in signal transduction.
authors.affiliation: Beijing National Laboratory for Molecular Sciences, Key Laboratory of Molecular Nanostructure and Nanotechnology, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190, P.R. China.|University of Chinese Academy of Sciences, Beijing, 100049, P.R. China.
authors.forename: Nan
authors.initials: N
authors.lastname: Li
authors.affiliation: Beijing National Laboratory for Molecular Sciences, Key Laboratory of Molecular Nanostructure and Nanotechnology, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190, P.R. China.
authors.forename: Yong
authors.initials: Y
authors.lastname: Yang
authors.affiliation: Beijing National Laboratory for Molecular Sciences, Key Laboratory of Molecular Nanostructure and Nanotechnology, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190, P.R. China.
authors.forename: Kangmin
authors.initials: K
authors.lastname: He
authors.affiliation: Key Laboratory of Protein and Peptide Drugs, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, P.R. China.
authors.forename: Fayun
authors.initials: F
authors.lastname: Zhang
authors.affiliation: Beijing National Laboratory for Molecular Sciences, Key Laboratory of Molecular Nanostructure and Nanotechnology, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190, P.R. China.
authors.forename: Libo
authors.initials: L
authors.lastname: Zhao
authors.affiliation: Beijing National Laboratory for Molecular Sciences, Key Laboratory of Molecular Nanostructure and Nanotechnology, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190, P.R. China.|University of Chinese Academy of Sciences, Beijing, 100049, P.R. China.
authors.forename: Wei
authors.initials: W
authors.lastname: Zhou
authors.affiliation: Beijing National Laboratory for Molecular Sciences, Key Laboratory of Molecular Nanostructure and Nanotechnology, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190, P.R. China.
authors.forename: Jinghe
authors.initials: J
authors.lastname: Yuan
authors.affiliation: Key Laboratory of Protein and Peptide Drugs, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, P.R. China.
authors.forename: Wei
authors.initials: W
authors.lastname: Liang
authors.affiliation: Beijing National Laboratory for Molecular Sciences, Key Laboratory of Molecular Nanostructure and Nanotechnology, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190, P.R. China.|University of Chinese Academy of Sciences, Beijing, 100049, P.R. China.
authors.forename: Xiaohong
authors.initials: X
authors.lastname: Fang
mesh_terms: D002462:Cell Membrane / Q000378:metabolism*; D004705:Endocytosis; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D042541:Intracellular Space / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D008954:Models, Biological; D062105:Molecular Docking Simulation; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D000077293:Receptor, Transforming Growth Factor-beta Type I; D018125:Receptors, Transforming Growth Factor beta / Q000378:metabolism; D000072760:Single Molecule Imaging / Q000379:methods*; D051900:Smad3 Protein / Q000378:metabolism*
chemical_list: D018125:Receptors, Transforming Growth Factor beta; D051900:Smad3 Protein; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D017346:Protein Serine-Threonine Kinases; D000077293:Receptor, Transforming Growth Factor-beta Type I