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A nanobody-based fluorescent reporter reveals human α-synuclein in the cell cytosol
Aggregation and spreading of α-Synuclein (αSyn) are hallmarks of several neurodegenerative diseases, thus monitoring human αSyn (hαSyn) in animal models or cell cultures is vital for the field. However, the detection of native hαSyn in such systems is challenging. We show that the nanobody NbSyn87,...
Autores principales: | , , , , , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Nature Publishing Group UK
2020
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7264335/ https://www.ncbi.nlm.nih.gov/pubmed/32483166 http://dx.doi.org/10.1038/s41467-020-16575-0 |
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author | Gerdes, Christoph Waal, Natalia Offner, Thomas Fornasiero, Eugenio F. Wender, Nora Verbarg, Hannes Manzini, Ivan Trenkwalder, Claudia Mollenhauer, Brit Strohäker, Timo Zweckstetter, Markus Becker, Stefan Rizzoli, Silvio O. Basmanav, Fitnat Buket Opazo, Felipe |
author_facet | Gerdes, Christoph Waal, Natalia Offner, Thomas Fornasiero, Eugenio F. Wender, Nora Verbarg, Hannes Manzini, Ivan Trenkwalder, Claudia Mollenhauer, Brit Strohäker, Timo Zweckstetter, Markus Becker, Stefan Rizzoli, Silvio O. Basmanav, Fitnat Buket Opazo, Felipe |
author_sort | Gerdes, Christoph |
collection | PubMed |
description | Aggregation and spreading of α-Synuclein (αSyn) are hallmarks of several neurodegenerative diseases, thus monitoring human αSyn (hαSyn) in animal models or cell cultures is vital for the field. However, the detection of native hαSyn in such systems is challenging. We show that the nanobody NbSyn87, previously-described to bind hαSyn, also shows cross-reactivity for the proteasomal subunit Rpn10. As such, when the NbSyn87 is expressed in the absence of hαSyn, it is continuously degraded by the proteasome, while it is stabilized when it binds to hαSyn. Here, we exploit this feature to design a new Fluorescent Reporter for hαSyn (FluoReSyn) by fusing NbSyn87 to fluorescent proteins, which results in fluorescence signal fluctuations depending on the presence and amounts of intracellular hαSyn. We characterize this biosensor in cells and tissues to finally reveal the presence of transmittable αSyn in human cerebrospinal fluid, demonstrating the potential of FluoReSyn for clinical research and diagnostics. |
format | Online Article Text |
id | pubmed-7264335 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2020 |
publisher | Nature Publishing Group UK |
record_format | MEDLINE/PubMed |
spelling | pubmed-72643352020-06-12 A nanobody-based fluorescent reporter reveals human α-synuclein in the cell cytosol Gerdes, Christoph Waal, Natalia Offner, Thomas Fornasiero, Eugenio F. Wender, Nora Verbarg, Hannes Manzini, Ivan Trenkwalder, Claudia Mollenhauer, Brit Strohäker, Timo Zweckstetter, Markus Becker, Stefan Rizzoli, Silvio O. Basmanav, Fitnat Buket Opazo, Felipe Nat Commun Article Aggregation and spreading of α-Synuclein (αSyn) are hallmarks of several neurodegenerative diseases, thus monitoring human αSyn (hαSyn) in animal models or cell cultures is vital for the field. However, the detection of native hαSyn in such systems is challenging. We show that the nanobody NbSyn87, previously-described to bind hαSyn, also shows cross-reactivity for the proteasomal subunit Rpn10. As such, when the NbSyn87 is expressed in the absence of hαSyn, it is continuously degraded by the proteasome, while it is stabilized when it binds to hαSyn. Here, we exploit this feature to design a new Fluorescent Reporter for hαSyn (FluoReSyn) by fusing NbSyn87 to fluorescent proteins, which results in fluorescence signal fluctuations depending on the presence and amounts of intracellular hαSyn. We characterize this biosensor in cells and tissues to finally reveal the presence of transmittable αSyn in human cerebrospinal fluid, demonstrating the potential of FluoReSyn for clinical research and diagnostics. Nature Publishing Group UK 2020-06-01 /pmc/articles/PMC7264335/ /pubmed/32483166 http://dx.doi.org/10.1038/s41467-020-16575-0 Text en © The Author(s) 2020 Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/. |
spellingShingle | Article Gerdes, Christoph Waal, Natalia Offner, Thomas Fornasiero, Eugenio F. Wender, Nora Verbarg, Hannes Manzini, Ivan Trenkwalder, Claudia Mollenhauer, Brit Strohäker, Timo Zweckstetter, Markus Becker, Stefan Rizzoli, Silvio O. Basmanav, Fitnat Buket Opazo, Felipe A nanobody-based fluorescent reporter reveals human α-synuclein in the cell cytosol |
title | A nanobody-based fluorescent reporter reveals human α-synuclein in the cell cytosol |
title_full | A nanobody-based fluorescent reporter reveals human α-synuclein in the cell cytosol |
title_fullStr | A nanobody-based fluorescent reporter reveals human α-synuclein in the cell cytosol |
title_full_unstemmed | A nanobody-based fluorescent reporter reveals human α-synuclein in the cell cytosol |
title_short | A nanobody-based fluorescent reporter reveals human α-synuclein in the cell cytosol |
title_sort | nanobody-based fluorescent reporter reveals human α-synuclein in the cell cytosol |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7264335/ https://www.ncbi.nlm.nih.gov/pubmed/32483166 http://dx.doi.org/10.1038/s41467-020-16575-0 |
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