A-synuclein facilitates endocytosis by elevating the steady-state levels of phosphatidylinositol 4,5-bisphosphate

a-Synuclein (a-Syn) is a protein implicated in the pathogenesis of Parkinson’s disease (PD). It is an intrinsically disordered protein that binds acidic phospholipids. Growing evidence supports a role for a-Syn in membrane trafficking, including, mechanisms of endocytosis and exocytosis, although the exact role of a-Syn in these mechanisms is currently unclear. Here we investigate the associations of a-Syn with the acidic phosphoinositides (PIPs), phosphatidylinositol 4,5-bisphosphate (PI(4,5) P₂) and phosphatidylinositol 3,4-bisphosphate (PI(3,4)P₂). Our results show that a-Syn colocalizes with PIP₂ and the phosphorylated active form of the clathrin adaptor protein 2 (AP2) at clathrin-coated pits. Using endocytosis of transferrin as an indicator for clathrin-mediated endocytosis (CME), we find that a-Syn involvement in endocytosis is specifically mediated through PI(4,5)P₂ levels on the plasma membrane. In accord with their effects on PI(4,5)P₂ levels, the PD associated A30P, E46K, and A53T mutations in a-Syn further enhance CME in neuronal and nonneuronal cells. However, lysine to glutamic acid substitutions at the KTKEGV repeat domain of a-Syn, which interfere with phospholipid binding, are ineffective in enhancing CME. We further show that the rate of synaptic vesicle (SV) endocytosis is differentially affected by the a-Syn mutations and associates with their effects on PI(4,5)P₂ levels, however, with the exception of the A30P mutation. This study provides evidence for a critical involvement of PIPs in a-Syn–mediated membrane trafficking.