Bioorthogonal labeling of transmembrane proteins with non-canonical amino acids unveils masked epitopes in live neurons

Diogo Bessa-Neto, Gerti Beliu, Alexander Kuhlemann, Valeria Pecoraro, Sören Doose, Natacha Retailleau, Nicolas Chevrier, David Perrais, Markus Sauer () and Daniel Choquet ()
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Diogo Bessa-Neto: University of Bordeaux, CNRS, Interdisciplinary Institute for Neuroscience, IINS, UMR 5297
Gerti Beliu: University of Würzburg, Biocenter, Am Hubland
Alexander Kuhlemann: University of Würzburg, Biocenter, Am Hubland
Valeria Pecoraro: University of Bordeaux, CNRS, Interdisciplinary Institute for Neuroscience, IINS, UMR 5297
Sören Doose: University of Würzburg, Biocenter, Am Hubland
Natacha Retailleau: University of Bordeaux, CNRS, Interdisciplinary Institute for Neuroscience, IINS, UMR 5297
Nicolas Chevrier: University of Bordeaux, CNRS, Interdisciplinary Institute for Neuroscience, IINS, UMR 5297
David Perrais: University of Bordeaux, CNRS, Interdisciplinary Institute for Neuroscience, IINS, UMR 5297
Markus Sauer: University of Würzburg, Biocenter, Am Hubland
Daniel Choquet: University of Bordeaux, CNRS, Interdisciplinary Institute for Neuroscience, IINS, UMR 5297

Nature Communications, 2021, vol. 12, issue 1, 1-16

Abstract: Abstract Progress in biological imaging is intrinsically linked to advances in labeling methods. The explosion in the development of high-resolution and super-resolution imaging calls for new approaches to label targets with small probes. These should allow to faithfully report the localization of the target within the imaging resolution – typically nowadays a few nanometers - and allow access to any epitope of the target, in the native cellular and tissue environment. We report here the development of a complete labeling and imaging pipeline using genetic code expansion and non-canonical amino acids in neurons that allows to fluorescently label masked epitopes in target transmembrane proteins in live neurons, both in dissociated culture and organotypic brain slices. This allows us to image the differential localization of two AMPA receptor (AMPAR) auxiliary subunits of the transmembrane AMPAR regulatory protein family in complex with their partner with a variety of methods including widefield, confocal, and dSTORM super-resolution microscopy.

Date: 2021
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DOI: 10.1038/s41467-021-27025-w

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