title: Reversible Optogenetic Control of Subcellular Protein Localization in a Live Vertebrate Embryo.
abstract: We demonstrate the utility of the phytochrome system to rapidly and reversibly recruit proteins to specific subcellular regions within specific cells in a living vertebrate embryo. Light-induced heterodimerization using the phytochrome system has previously been used as a powerful tool to dissect signaling pathways for single cells in culture but has not previously been used to reversibly manipulate the precise subcellular location of proteins in multicellular organisms. Here we report the experimental conditions necessary to use this system to manipulate proteins in vivo. As proof of principle, we demonstrate that we can manipulate the localization of the apical polarity protein Pard3 with high temporal and spatial precision in both the neural tube and the embryo's enveloping layer epithelium. Our optimizations of optogenetic component expression and chromophore purification and delivery should significantly lower the barrier for establishing this powerful optogenetic system in other multicellular organisms.
authors.affiliation: MRC Centre for Developmental Neurobiology, King's College London, London SE1 1UL, UK.
authors.forename: Clare E
authors.initials: CE
authors.lastname: Buckley
authors.affiliation: MRC Centre for Developmental Neurobiology, King's College London, London SE1 1UL, UK.
authors.forename: Rachel E
authors.initials: RE
authors.lastname: Moore
authors.affiliation: Cardiovascular Research Institute, University of California, San Francisco, San Francisco, CA 94158-9001, USA; Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, CA 94158-2517, USA.
authors.forename: Anna
authors.initials: A
authors.lastname: Reade
authors.affiliation: Cardiovascular Research Institute, University of California, San Francisco, San Francisco, CA 94158-9001, USA; Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, CA 94158-2517, USA.
authors.forename: Anna R
authors.initials: AR
authors.lastname: Goldberg
authors.affiliation: Cardiovascular Research Institute, University of California, San Francisco, San Francisco, CA 94158-9001, USA; Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, CA 94158-2517, USA. Electronic address: orion.weiner@ucsf.edu.
authors.forename: Orion D
authors.initials: OD
authors.lastname: Weiner
authors.affiliation: MRC Centre for Developmental Neurobiology, King's College London, London SE1 1UL, UK. Electronic address: jon.clarke@kcl.ac.uk.
authors.forename: Jonathan D W
authors.initials: JDW
authors.lastname: Clarke
mesh_terms: D000818:Animals; D008027:Light; D062308:Optogenetics; D021381:Protein Transport; D015398:Signal Transduction / Q000502:physiology*; D015027:Zebrafish / Q000196:embryology / Q000235:genetics / Q000378:metabolism*
keywords: Pard3; apico-basal polarity; asymmetric inheritance; optogenetics; phytochrome; zebrafish