α-Synuclein facilitates clathrin assembly in synaptic vesicle endocytosis

Abstract

Abstractα-Synuclein and family members β-, and γ-synuclein, are presynaptic proteins that sense and generate membrane curvature, properties important for synaptic vesicle (SV) cycling. αβγ-synuclein triple knockout (KO) neurons exhibit SV endocytosis (SVE) deficits. Here, we investigate how SVE is regulated by α-synuclein. Immuno-electron microscopy (EM) of synaptosomes reveals that α-synuclein relocalizes from SVs to the synaptic membrane upon stimulation, allowing α-synuclein to function on presynaptic membranes during or after stimulation. On cell membranes, we observe that α-synuclein is colocalized with clathrin and its adaptor AP180. Clathrin patches that contain both α-synuclein and AP180 were significantly larger than clathrin patches containing either protein alone. We also find that recruitment of clathrin and AP180 recruitment to membranes are altered in the absence of synucleins. Visualizing clathrin assembly on membranes using an in vitro endocytosis reconstitution system reveals that α-synuclein increases clathrin patch size and enhances clathrin lattice curvature, facilitating normal clathrin coated pit maturation. Thus, α-synuclein is an endocytic accessory protein that acts at early stages of SVE to controls the size and curvature of clathrin structures on the membrane.