Abstract
Reducing α-synuclein pathology constitutes a plausible strategy against Parkinson’s disease. As we recently demonstrated, the β-wrapin protein AS69 binds an N-terminal region in monomeric α-synuclein, interferes with fibril nucleation, and reduces α-synuclein aggregation in vitro and in a fruit fly model of α-synuclein toxicity. The aim of this study was to investigate whether AS69 also reduces α-synuclein pathology in mammalian neurons. To induce α-synuclein pathology, primary mouse neurons were exposed to pre-formed fibrils (PFF) of human α-synuclein. PFF were also injected into the striatum of A30P-α-synuclein transgenic mice. The extent of α-synuclein pathology was determined by phospho-α-synuclein staining and by Triton X-100 solubility. The degeneration of neuronal somata, dendrites, and axon terminals was determined by immunohistochemistry. AS69 and PFF were taken up by primary neurons. AS69 did not alter PFF uptake, but AS69 did reduce PFF-induced α-synuclein pathology. PFF injection into mouse striatum led to α-synuclein pathology and dystrophic neurites. Co-injection of AS69 abrogated PFF-induced pathology. AS69 also reduced the PFF-induced degeneration of dopaminergic axon terminals in the striatum and the degeneration of dopaminergic dendrites in the substantia nigra pars reticulata. AS69 reduced the activation of astroglia but not microglia in response to PFF injection. Collectively, AS69 reduced PFF-induced α-synuclein pathology and the associated neurodegeneration in primary neurons and in mouse brain. Our data therefore suggest that small proteins binding the N-terminus of α-synuclein monomers are promising strategies to modify disease progression in Parkinson’s disease.
Highlights
In Parkinson’s disease (PD) and other synucleinopathies, aggregation and accumulation of α-synuclein is considered a central event
In neurons exposed to pre-formed fibrils (PFF), human accumulation of α-synuclein (aSyn) staining was detected intracellularly 24 h after adding PFF (Figures 1A,B), confirming that primary neurons take up PFF–as demonstrated previously by others (VolpicelliDaley et al, 2014)
In primary neurons exposed to PFF, staining for human aSyn was already observed after 24 h whereas the extent of phospho-aSyn staining increased mainly between 24 and 72 h, consistent with previous findings that seeding of aSyn pathology by extracellular PFF takes time (Flavin et al, 2017; Rodriguez et al, 2018)
Summary
In Parkinson’s disease (PD) and other synucleinopathies, aggregation and accumulation of α-synuclein (aSyn) is considered a central event. Reducing the extent of aSyn pathology represents an attractive neuroprotective strategy against synucleinopathies (Obeso et al, 2017). The engineered β-wrapin AS69, in contrast, binds monomeric aSyn with high effectivity and high specificity (Mirecka et al, 2014). AS69 wraps around a sequence region of monomeric aSyn comprising residues 37–54 and stabilizes a β-hairpin conformation (Mirecka et al, 2014). AS69 binding interferes with primary and secondary nucleation processes and inhibits the proliferation of aSyn fibrils (Agerschou et al, 2019). In HEK293T cells, AS69 reduces oligomerization and aggregation of aSyn; in a fruit fly model of A53T aSyn toxicity, AS69 reduces aggregation of aSyn in neurons and rescues the locomotor deficit resulting from neuronal aSyn expression (Agerschou et al, 2019)
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