Dynamic superresolution imaging of endogenous proteins on living cells at ultra-high density.
Versatile superresolution imaging methods, able to give dynamic information of endogenous molecules at high density, are still lacking in biological science. Here, superresolved images and diffusion maps of membrane proteins are obtained on living cells. The method consists of recording thousands of single-molecule trajectories that appear sequentially on a cell surface upon continuously labeling molecules of interest. It allows studying any molecules that can be labeled with fluorescent ligands including endogenous membrane proteins on living cells. This approach, named universal PAINT (uPAINT), generalizes the previously developed point-accumulation-for-imaging-in-nanoscale-topography (PAINT) method for dynamic imaging of arbitrary membrane biomolecules. We show here that the unprecedented large statistics obtained by uPAINT on single cells reveal local diffusion properties of specific proteins, either in distinct membrane compartments of adherent cells or in neuronal synapses.
Giannone
Gregory
G
Centre National de la Recherche Scientifique UMR 5091, Cellular Physiology of the Synapse, Bordeaux, France.
Hosy
Eric
E
Levet
Florian
F
Constals
Audrey
A
Schulze
Katrin
K
Sobolevsky
Alexander I
AI
Rosconi
Michael P
MP
Gouaux
Eric
E
Tampé
Robert
R
Choquet
Daniel
D
Cognet
Laurent
L
D000818:Animals; D019556:COS Cells; D002470:Cell Survival; D002522:Chlorocebus aethiops; D005347:Fibroblasts / Q000166:cytology* / Q000378:metabolism*; D049452:Green Fluorescent Proteins / Q000378:metabolism; D008024:Ligands; D051379:Mice; D057054:Molecular Imaging / Q000379:methods*; D009474:Neurons / Q000166:cytology / Q000378:metabolism; D021381:Protein Transport; D011506:Proteins / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D018091:Receptors, AMPA / Q000378:metabolism
D008024:Ligands; D011506:Proteins; D018091:Receptors, AMPA; D049452:Green Fluorescent Proteins