The effect of S129 phosphorylation on the interaction of α-synuclein with synaptic and cellular membranes

Abstract In healthy brain, less than 5% of α-synuclein (α-syn) is phosphorylated at serine 129 (pS129). However, within Parkinson disease (PD) Lewy bodies, 89% of α-syn is pS129. The effects of pS129 modification on α-syn distribution and solubility are poorly understood. As α-syn normally exists in both membrane-bound and cytosolic compartments, we examined the binding and dissociation of pS129 α-syn and analyzed the effects of manipulating pS129 levels on α-syn membrane interactions using synaptosomal membranes and neural precursor cells from α-syn-deficient mice or transgenic mice expressing human α-syn. We first evaluated the recovery of the pS129 epitope following either α-syn membrane binding or dissociation. We demonstrate a rapid turnover of pS129 during both binding to and dissociation from synaptic membranes. Although the membrane binding of WT α-syn was insensitive to modulation of pS129 levels by multiple strategies (the use of phosphomimic S129D and non-phosphorylated S129A α-syn mutants; by enzymatic dephosphorylation of pS129 or proteasome inhibitor-induced elevation in pS129; or by inhibition or stable overexpression of PLK2), PD mutant pS129 α-syn showed a preferential membrane association compared to WT pS129 α-syn. Collectively, these data suggest that phosphorylation at S129 is dynamic, and that the subcellular distribution of α-syn bearing PD-linked mutations, A30P or A53T, is influenced by the phosphorylation state of S129.