Contribution of Alanine-76 and Serine Phosphorylation inα-Synuclein Membrane Association and Aggregation in Yeasts

Abstract

In Parkinson's disease (PD), misfolded and aggregated α-synuclein protein accumulates in degenerating midbrain dopaminergic neurons. The amino acid alanine-76 in α-synuclein and phosphorylation at serine-87 and serine-129 are thought to regulate its aggregation and toxicity. However, their exact contributions to α-synuclein membrane association are less clear. We found that α-synuclein is indeed phosphorylated in fission yeast and budding yeast, the two models that we employed for assessing α-synuclein aggregation and membrane association properties, respectively. Surprisingly, blocking serine phosphorylation (S87A, S129A, and S87A/S129A) or mimicking it (S87D, S129D) altered α-synuclein aggregation in fission yeast. Either blocking or mimicking this phosphorylation increased endomembrane association in fission yeast, but only mimicking it decreased plasma membrane association in budding yeast. Polar substitution mutations of alanine-76 (A76E and A76R) decreased α-synuclein membrane association in budding yeast and decreased aggregation in fission yeast. These yeast studies extend our understanding of serine phosphorylation and alanine-76 contributions to α-synuclein aggregation and are the first to detail their impact on α-synuclein's plasma membrane and endomembrane association.

Assessing the Subcellular Dynamics of Alpha-synuclein Using Photoactivation Microscopy

“…This tendency might at least partially explain the cytoplasmic inclusions detected in the cells expressing this mutant aSyn and suggest that the phosphorylation status on S129 is crucial for aggregation, in agreement with recent findings in yeast, in which S129A mutant potentiates the formation of aSyn foci [46]. GRKs and PLKs modulate the dynamics of aSyn in different ways and we did not find a consistent pattern that can explain the role of S129 phosphorylation on the distribution of aSyn.…”

Phosphorylation of α-Synuclein Protein at Ser-129 Reduces Neuronal Dysfunction by Lowering Its Membrane Binding Property in Caenorhabditis elegans

“…In addition, S129D-␣-synuclein expressed in budding yeast exhibited decreased plasma membrane association (37). These observations are consistent with our results, suggesting the conserved mechanism of regulation of neurotoxicity and membrane binding by Ser-129 phosphorylation.…”

Implication of Alpha-Synuclein Phosphorylation at S129 in Synucleinopathies: What Have We Learned in the Last Decade?

“…On the other hand, cellular and animal studies, based on the use of the phospho-mimic strategy, reported an inhibitory effect of S129 phosphorylation on α -syn-membrane interaction. In yeast and worm models of PD, S129→A substitution (to block phosphorylation) increases α -syn membrane-bound fraction, however the phospho-mimic mutation (S129D) inhibits its association with membranes [ 46, 47 ]. A similar observation has been reported in an adeno-associated virus (AAV)-based rat genetic model of PD where immuno-electron microscopy analysis detected the majority of the non-phosphorylatable α -syn mutant (S129A) associated with cellular membranes [ 48 ].…”