The Clathrin-Dependent Localization of Dopamine Transporter to Surface Membranes Is Affected by α-Synuclein

Parkinson's disease (PD) is a progressive agedependent neurodegenerative disorder, predominantly affecting the dopamine-producing neurons residing at the substantia nigra. Abnormalities in alpha-synuclein (alpha-Syn) and dopamine transporter (DAT) are implicated in the pathogenesis of PD. We tested the hypothesis that alpha-Syn regulates surface DAT localization and DAT activity, in cultured cells co-expressing alpha-Syn and DAT, and in brains of mice modeling PD, transgenic for the mutant A53T alpha-Syn form. The results indicate that alpha-Syn expression affects the partitioning of DAT between the cell surface and intracellular compartments, resulting in lower surface DAT levels. Accordingly, lower uptake of tritiated dopamine was measured in synaptosomes of A53T alpha-Syn transgenic mouse brains. Importantly, we show that the effect of alpha-Syn on surface DAT is mediated by clathrin. Downregulation of clathrin by specific siRNAs directed against its heavy chain abolished the effect of alpha-Syn on phorbol 12-myristate13-acetate-induced DAT internalization. These results suggest that alpha-Syn plays a role in regulating dopamine homeostasis through its involvement in clathrin-mediated endocytosis.