Figure 1
Prion aggregates traffic via autolysosomes in SMB.s15 cells. All the experiments were performed either with or without 10 μM CQ. (a) Immunofluorescence staining using anti PrP antibody BC6 in SMB-PS and SMB.s15 cells (N = 3). (b) Representative immunoblot shows PK resistant PrPSc in SMB.s15 cells (N = 2). (c) Representative immunoblot for LC3 and β-tubulin in SMB-PS and SMB.s15 cells, lane 2 & 4 were treated with CQ (N = 3). (d) Representative TEM images of SMB-PS and SMB.s15 after CQ treatment, red arrows indicate vesicles with aggregates (N = 2). (e) Increased number of vesicles with aggregates in SMB.s15 cells. All SMB.PS cells imaged with TEM showed minimal aggregates (0–5/cell) whereas all SMB.s15 cells images showed increased number of aggregates (5–24/cell). (f) WB for PK resistant PrPSc after CQ treatment in SMB.s15 cells (N = 3). (g) Representative image of proteostat dye (aggregates) and LC3 immunofluorescence staining of SMB.s15 cells after CQ treatment compared to untreated control cells (N = 3). (h) Significantly increased LC3 stained proteostat positive aggregates in SMB.s15 cells on CQ treatment. (i) Representative image of PrPSc and LC3 immunofluorescence staining of SMB.s15 cells after CQ treatment compared to untreated control cells (N = 5). (j) Significantly increased LC3 stained PrPSc aggregates in SMB.s15 cells on CQ treatment. Scale bar = 10 μm. Respective full length blots are shown in Supplementary Fig. 3. Abbreviations used Chloroquine (CQ), Transmission Electron Microscopy (TEM), proteinase K (PK).
