Figure 5: Differential seeding properties of the two α-syn polymorphs.

The diffuse distribution of ChFP-α-syn fluorescence in Neuro 2a cells (a) was converted to a punctate pattern within 4 h upon exposure of the cells to Alexa Fluor 488-labelled α-syn fibrils (0.01 nM). (b) No such redistribution was observed within the same time frame upon treatment of the cells with α-syn ribbons at a 10 fold higher concentration (0.1 nM). (d) The first ChFP-α-syn puncta were apparent in cells exposed for 18 h to α-syn ribbons (0.1 nM) (c) a time at which ChFP-α-syn was almost entirely recruited into puncta in cells exposed to α-syn fibrils (0.01 nM). (e) control cells unexposed to α-syn fibrils or ribbons. The merged images (right column) show that ChFP-α-syn puncta colocalize extensively with the exogenous Alexa Fluor 488-labelled α-syn polymorphs. Scale bar, 10 μm. (f) Neuro 2a cells exposed 48 h to α-syn fibrils or ribbons (0.1 nM) were passaged 2–3 times a week. The generation time of N2A cells under our experimental conditions was 14–16 h. The presence of internalized aggregates (green fluorescence) and induced puncta (red fluorescence) was scored in a blind manner by two scientists. The time courses of exogenous α-syn fibrils or ribbons (in black) and puncta induced by ribbons (blue) or fibrils (red) disappearance are shown. Data are mean±s.d. (n=3 independent measurements).