A nanobody-based fluorescent reporter reveals human α-synuclein in the cell cytosol

Christoph Gerdes, Natalia Waal, Thomas Offner, Eugenio F. Fornasiero, Nora Wender, Hannes Verbarg, Ivan Manzini, Claudia Trenkwalder, Brit Mollenhauer, Timo Strohäker, Markus Zweckstetter, Stefan Becker, Silvio O. Rizzoli, Fitnat Buket Basmanav and Felipe Opazo ()
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Christoph Gerdes: University Medical Center Göttingen
Natalia Waal: University Medical Center Göttingen
Thomas Offner: Justus-Liebig University Giessen
Eugenio F. Fornasiero: University Medical Center Göttingen
Nora Wender: University Medical Center Göttingen
Hannes Verbarg: University Medical Center Göttingen
Ivan Manzini: Justus-Liebig University Giessen
Claudia Trenkwalder: University Medical Center Göttingen
Brit Mollenhauer: Klinikstraße 16
Timo Strohäker: Von-Siebold-Str. 3a
Markus Zweckstetter: Von-Siebold-Str. 3a
Stefan Becker: Max Planck Institute for Biophysical Chemistry
Silvio O. Rizzoli: University Medical Center Göttingen
Fitnat Buket Basmanav: University Medical Center Göttingen
Felipe Opazo: University Medical Center Göttingen

Nature Communications, 2020, vol. 11, issue 1, 1-13

Abstract: Abstract 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.

Date: 2020
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DOI: 10.1038/s41467-020-16575-0

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