Project
387-Ando-StayGold
Title
Super resolution cellular images using fluorescent-tags of StayGold variants
Description
Although StayGold is a bright and highly photostable fluorescent protein, its propensity for obligate dimer formation may hinder applications in molecular fusion and membrane targeting. To attain monovalent as well as bright and photostable labeling, the authors engineered tandem dimers of StayGold to promote dispersibility. On the basis of the crystal structure of this fluorescent protein, they disrupted the dimerization to generate a monomeric variant that offers improved photostability and brightness compared to StayGold. The authors applied the new monovalent StayGold tools to live-cell imaging experiments using spinning-disk laser-scanning confocal microscopy or structured illumination microscopy. They achieved cell-wide, high-spatiotemporal resolution and sustained imaging of dynamic subcellular events, including the targeting of endogenous condensin I
to mitotic chromosomes, the movement of the Golgi apparatus and its membranous derivatives along microtubule networks, the distribution of cortical filamentous actin and the remolding of cristae membranes within mobile mitochondria.
Funding
This work was supported in part by Grant-in-Aid for Scientific Research (S) (21H05041 to A.M.), Grant-in-Aid for Innovative Areas: Resonance Bio (15H05948 to A.M.) and Information Physics of
Living Matters (19H05794 and 19H05795 to Y.O.), Japan Science and Technology Agency Core Research for Evolutionary Science and Technology program, ‘Spatiotemporal dynamics of intracellular components’ (JPM JCR20E2 to Y.O.) and Moonshot R&D program ‘Realization of ultra-early disease prediction and intervention by
2050’ (JPM JMS2025-14 to Y.O.), Marine Biomass Innovation Project (NFRFT-2020-00452 to A.M.), the Brain Mapping by Integrated Neurotechnologies for Disease Studies from AMED (Brain/MINDS, JP15dm0207001 to A.M.), RIKEN Collaboration Seed Fund (to M.T. and R.A.) and Platform Project for Supporting Drug Discovery and Life Science Research (Basis for Supporting Innovative Drug Discovery and Life Science Research (BINDS)) from AMED (JP21am0101070 to M.Y.).
Title
Z-series images of td5StayGold(c4)=GianCreg and TGN46 in fixed HeLa cells
Description
Z-series images of td5StayGold(c4)=GianCreg and a Golgi marker, TGN46 in fixed HeLa cells.
Description
Fixed HeLa cells expressing td5StayGold(c4)=GianCreg
Extrinsic variables
td5StayGold(c4)=GianCreg was transfected
Gene ontology: Biological processes
Gene ontology: Cellular components
Gene ontology: Molecular functions
Protein names
Golgin subfamily B member 1 (
Q14789)
Trans-Golgi network integral membrane protein 2 (
O43493)
Dimensions
512x512x34x3x1
Z scales
0.52 micrometeer
Microscopy types
spinning disk confocal microscopy (
FBbi:00000253)
(confocal microscopy (LSCM))
Visualization methods
primary antibody plus labeled secondary antibody
td5StayGold(c4), a StayGold derivative
Contrast enhancing methods
Resolution enhancing methods
Sample preparation methods
fixation method
Body
Olympus-Evident IX83
Detector
Hamamatsu ORCA-Flash4.0 CL, USB
Objective
Olympus-Evident UPLAPO100XOHR