Dynamic reversibility of α-synuclein serine-129 phosphorylation is impaired in synucleinopathy models

alpha-Synuclein phosphorylation at serine-129 (pS129) is a widely used surrogate marker of pathology in Parkinson's disease and other synucleinopathies. However, we recently demonstrated that phosphorylation of S129 is also a physiological activator of synaptic transmission. In a feed-forward fashion, neuronal activity triggers reversible pS129. Here, we show that Parkinson's disease-linked missense mutations in SNCA impact activity-dependent pS129. Under basal conditions, cytosol-enriched A30P, H50Q, and G51D mutant forms of alpha-synuclein exhibit reduced pS129 levels in rat primary cortical neurons. A53T pS129 levels are similar to wild-type, and E46K pS129 levels are higher. A30P and E46K mutants show impaired reversibility of pS129 after stimulation. For the engineered profoundly membrane-associated alpha-synuclein mutant 3K (E35K + E46K + E61K), de-phosphorylation was virtually absent after blocking stimulation, implying that reversible pS129 is severely compromised. Importantly, pS129 excess resulting from proteasome inhibition is also associated with reduced reversibility by neuronal inhibition, kinase inhibition, or phosphatase activation. Our findings suggest that perturbed pS129 dynamics are probably a shared characteristic of pathology-associated alpha-synuclein, with possible implications for synucleinopathy treatment and diagnosis.

Automated summary

Research shows that Parkinson's disease-linked mutations in alpha-synuclein can impact the phosphorylation of serine-129, which is important for synaptic transmission. These mutations reduce the levels of phosphorylation and impair its reversibility. Additionally, inhibiting the proteasome also reduces the reversibility of this phosphorylation.