GRP78 clustering at the cell surface of neurons transduces the action of exogenous alpha-synuclein

Mutation or multiplication of the alpha-synuclein (Syn)-encoding gene is frequent cause of early onset Parkinson''s disease (PD). Recent evidences point to the pathogenic role of excess Syn also in sporadic PD. Syn is a cytosolic protein, which has been shown to be released from neurons. Here we provide evidence that extracellular Syn induces an increase in surface-exposed glucose-related protein of 78 kDa (GRP78), which becomes clustered in microdomains of the neuronal plasma membrane. Upon interacting with Syn, GRP78 activates a signaling cascade leading to cofilin 1 inactivation and stabilization of microfilaments, thus affecting morphology and dynamics of actin cytoskeleton in cultured neurons. Downregulation of GRP78 abolishes the activity of exogenous Syn, indicating that it is the primary target of Syn. Inactivation of cofilin 1 and stabilization of actin cytoskeleton are present also in fibroblasts derived from genetic PD patients, which show a dramatic increase in stress fibers. Similar changes are displayed by control cells incubated with the medium of PD fibroblasts, only when Syn is present. The accumulation of Syn in the extracellular milieu, its interaction with the plasma membrane and Syn-driven clustering of GRP78 appear, therefore, responsible for the dysregulation of actin turnover, leading to early deficits in synaptic function that precede neurodegeneration.Alpha-synuclein (Syn), a small soluble protein abundant in the brain,¹ is implicated in the pathogenesis of various neurodegenerative disorders, among them Parkinson''s disease (PD).² Syn is the major component of intraneuronal inclusions found in the brain of PD patients.³ Distinct mutations⁴ as well as multiplications of the Syn-encoding gene (SNCA)^{5, 6, 7} have been identified in families with genetic forms of PD. Moreover, genome-wide association studies correlated polymorphisms in regions of the SNCA gene to sporadic cases of PD.⁸Syn is a natively unfolded protein enriched in presynaptic terminals, where it dynamically associates with synaptic vesicle membrane and lipid rafts.^{9, 10} A plethora of possible Syn interactors has been recognized¹¹ and among them cytoskeletal proteins. The influence of Syn on the microtubule-based cytoskeleton has been characterized in vitro.¹² Recently,¹³ Syn has been shown to depolymerize actin cytoskeleton, a master regulator of synaptic function,¹⁴ raising the possibility that the effects of Syn on synaptic vesicle dynamics are mediated by its interaction with actin.Syn is localized in the cytosolic compartment, is released from cultured neurons and is present in biological fluids¹⁵ indicating that Syn may also act extracellularly, participating in cell-to-cell transmission of pathology. Intraneuronal inclusions were found in healthy engrafted cells decades after transplantation in PD patients, and the host-to-graft transmission of Syn pre-formed fibrils has been demonstrated to occur in the mouse brain.¹⁶Here we show that exposure of neurons to extracellularly delivered wild-type (wt) or A30P-mutant monomeric Syn leads to cofilin 1 inactivation with the ensuing stabilization of microfilaments. These effects depend on Syn-driven clustering at the cell surface of the endoplasmic reticulum (ER) protein glucose-related protein of 78 kDa (GRP78). We propose actin cytoskeleton modification as the initial step in Syn-induced neurodegeneration, thereby actin and actin-binding proteins might constitute an effective target for screening and therapy of PD.