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

ABSTRACT: Aggregation and spreading of ?-Synuclein (?Syn) are hallmarks of several neurodegenerative diseases, thus monitoring human ?Syn (h?Syn) in animal models or cell cultures is vital for the field. However, the detection of native h?Syn in such systems is challenging. We show that the nanobody NbSyn87, previously-described to bind h?Syn, also shows cross-reactivity for the proteasomal subunit Rpn10. As such, when the NbSyn87 is expressed in the absence of h?Syn, it is continuously degraded by the proteasome, while it is stabilized when it binds to h?Syn. Here, we exploit this feature to design a new Fluorescent Reporter for h?Syn (FluoReSyn) by fusing NbSyn87 to fluorescent proteins, which results in fluorescence signal fluctuations depending on the presence and amounts of intracellular h?Syn. We characterize this biosensor in cells and tissues to finally reveal the presence of transmittable ?Syn in human cerebrospinal fluid, demonstrating the potential of FluoReSyn for clinical research and diagnostics.

SUBMITTER: Gerdes C 

PROVIDER: S-EPMC7264335 | biostudies-literature | 2020 Jun

REPOSITORIES: biostudies-literature