Super-Resolution Imaging of Plasma Membrane Proteins with Click Chemistry.
Besides its function as a passive cell wall, the plasma membrane (PM) serves as a platform for different physiological processes such as signal transduction and cell adhesion, determining the ability of cells to communicate with the exterior, and form tissues. Therefore, the spatial distribution of PM components, and the molecular mechanisms underlying it, have important implications in various biological fields including cell development, neurobiology, and immunology. The existence of confined compartments in the plasma membrane that vary on many length scales from protein multimers to micrometer-size domains with different protein and lipid composition is today beyond all questions. As much as the physiology of cells is controlled by the spatial organization of PM components, the study of distribution, size, and composition remains challenging. Visualization of the molecular distribution of PM components has been impeded mainly due to two problems: the specific labeling of lipids and proteins without perturbing their native distribution and the diffraction-limit of fluorescence microscopy restricting the resolution to about half the wavelength of light. Here, we present a bioorthogonal chemical reporter strategy based on click chemistry and metabolic labeling for efficient and specific visualization of PM proteins and glycans with organic fluorophores in combination with super-resolution fluorescence imaging by direct stochastic optical reconstruction microscopy (dSTORM) with single-molecule sensitivity.
Department of Biotechnology and Biophysics, Julius Maximilian University of Würzburg Würzburg, Germany.
Pablo
P
Mateos-Gil
Department of Biotechnology and Biophysics, Julius Maximilian University of Würzburg Würzburg, Germany.
Sebastian
S
Letschert
Department of Biotechnology and Biophysics, Julius Maximilian University of Würzburg Würzburg, Germany.
Sören
S
Doose
Department of Biotechnology and Biophysics, Julius Maximilian University of Würzburg Würzburg, Germany.
Markus
M
Sauer
click chemistry; dSTORM; localization microscopy; plasma membrane organization; protein clusters; super-resolution fluorescence microscopy
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