title: 3D Protein Dynamics in the Cell Nucleus.
abstract: The three-dimensional (3D) architecture of the cell nucleus plays an important role in protein dynamics and in regulating gene expression. However, protein dynamics within the 3D nucleus are poorly understood. Here, we present, to our knowledge, a novel combination of 1) single-objective based light-sheet microscopy, 2) photoconvertible proteins, and 3) fluorescence correlation microscopy, to quantitatively measure 3D protein dynamics in the nucleus. We are able to acquire >3400 autocorrelation functions at multiple spatial positions within a nucleus, without significant photobleaching, allowing us to make reliable estimates of diffusion dynamics. Using this tool, we demonstrate spatial heterogeneity in Polymerase II dynamics in live U2OS cells. Further, we provide detailed measurements of human-Yes-associated protein diffusion dynamics in a human gastric cancer epithelial cell line.
authors.affiliation: Mechanobiology Institute, National University of Singapore, Singapore.
authors.forename: Anand P
authors.initials: AP
authors.lastname: Singh
authors.affiliation: Institut Interdisciplinaire de Neurosciences, University of Bordeaux, France; Centre National de la Recherche Scientifique UMR 5297, University of Bordeaux, France.
authors.forename: Rémi
authors.initials: R
authors.lastname: Galland
authors.affiliation: Mechanobiology Institute, National University of Singapore, Singapore; Department of Physiology, National University of Singapore, Singapore.
authors.forename: Megan L
authors.initials: ML
authors.lastname: Finch-Edmondson
authors.affiliation: Mechanobiology Institute, National University of Singapore, Singapore.
authors.forename: Gianluca
authors.initials: G
authors.lastname: Grenci
authors.affiliation: Institut Interdisciplinaire de Neurosciences, University of Bordeaux, France; Centre National de la Recherche Scientifique UMR 5297, University of Bordeaux, France.
authors.forename: Jean-Baptiste
authors.initials: JB
authors.lastname: Sibarita
authors.affiliation: Institut Interdisciplinaire de Neurosciences, University of Bordeaux, France; Centre National de la Recherche Scientifique UMR 5297, University of Bordeaux, France.
authors.forename: Vincent
authors.initials: V
authors.lastname: Studer
authors.affiliation: Mechanobiology Institute, National University of Singapore, Singapore; Department of Biological Sciences, National University of Singapore, Singapore; Centre National de la Recherche Scientifique UMI 3639, Singapore. Electronic address: virgile.viasnoff@espci.fr.
authors.forename: Virgile
authors.initials: V
authors.lastname: Viasnoff
authors.affiliation: Mechanobiology Institute, National University of Singapore, Singapore; Department of Biological Sciences, National University of Singapore, Singapore; Institute for Molecular and Cell Biology, A(∗)Star, Singapore. Electronic address: dbsste@nus.edu.sg.
authors.forename: Timothy E
authors.initials: TE
authors.lastname: Saunders
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism*; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism*; D004058:Diffusion; D006801:Humans; D008856:Microscopy, Fluorescence; D010750:Phosphoproteins / Q000378:metabolism*; D038761:Photobleaching; D014157:Transcription Factors; D000091102:YAP-Signaling Proteins
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D010750:Phosphoproteins; D014157:Transcription Factors; D000091102:YAP-Signaling Proteins; C088374:YAP1 protein, human