SNCA/α-synuclein aggregation plays a crucial role in synucleinopathies such as Parkinson disease and dementia with Lewy bodies. Aggregating and nonaggregating SNCA species are degraded by the autophagy-lysosomal pathway (ALP). Previously, we have shown that the ALP is not only responsible for SNCA degradation but is also involved in the intracellular aggregation process of SNCA. An additional role of extracellular SNCA in the pathology of synucleinopathies substantiating a prion-like propagation hypothesis has been suggested since released SNCA species and spreading of SNCA pathology throughout neural cells have been observed. However, the molecular interplay between intracellular pathways, SNCA aggregation, release, and response of the local microenvironment remains unknown. Here, we attributed SNCA-induced toxicity mainly to secreted species in a cell culture model of SNCA aggregation and in SNCA transgenic mice: We showed that ALP inhibition by bafilomycinA1 reduced intracellular SNCA aggregation but increased secretion of smaller oligomers that exacerbated microenvironmental response including uptake, inflammation, and cellular damage. Low-aggregated SNCA was predominantly released by exosomes and RAB11A-associated pathways whereas high-aggregated SNCA was secreted by membrane shedding. In summary, our study revealed a novel role of the ALP by linking protein degradation to nonclassical secretion for toxic SNCA species. Thus, impaired ALP in the diseased brain not only limits intracellular degradation of misfolded proteins, but also leads to a detrimental microenvironmental response due to enhanced SNCA secretion. These findings suggest that the major toxic role of SNCA is related to its extracellular species and further supports a protective role of intracellular SNCA aggregation.
Keywords: ACTB/bAct, actin, β; AIF1/Iba1, allograft inflammatory factor 1; AK, adenylate kinase; ALP, autophagy-lysosomal pathway; ANXA5, annexin A5; BafA1, bafilomycinA1; CA1, cornu ammonis field1; CASP3/aCasp3, caspase-3; CD63, CD63 molecule; CM, conditioned medium; CMA, chaperone-mediated autophagy; CSF, cerebrospinal fluid; DLB, dementia with Lewy bodies; ER, endoplasmatic reticulum; ESCRT, endosomal sorting complex required for transport; EV, empty vector; GFAP, glial fibrillary acidic protein; HRP, horseradish peroxidase; HSPA8/Hsc70, heat shock 70kDa protein 8; Hippo, hippocampus; IL6/IL-6, interleukin-6; ILVs, intraluminal vesicles; LAMP2A/Lamp2a, lysosomal-associated membrane protein 2, isoform A; LB, Lewy bodies; LN, Lewy neuritis; MAP2, microtubule-associated protein 2; ML, molecular layer; MVBs, multivesicular bodies; N, neuron; Neoctx, neocortex; PD, Parkinson disease; PDGFB/PDGFb, platelet-derived growth factor subunit b; PF, particle fraction; PS, phosphatidylserine; Parkinson disease; RAB11A/rab11, member RAS oncogene family; RBFOX3/NeuN, RNA binding protein, fox-1 homolog (C. elegans) 3; RT, room temperature; S100B/S100b, S100 calcium-binding protein B; SL, stratum lacunosum; SNCA/aSyn; SNCA-T, tagged α-synuclein; SNCAIP/Sph1, synphilin-1; SYP, synaptophysin; TNF/TNFa, tumor necrosis factor α; TUBB3/b-III-Tub, tubulin, β 3 class III; UPS, ubiquitin proteasome system; WT-SNCA, wild-type α-synuclein; inflammation; lysosomal degradation; protein aggregation; secretion; synucleinopathies; tg, transgenic; α-synuclein.