Figure 1.
Knockout of RB1CC1 results in loss of photoreceptor outer segment (POS)-induced autophagy in ARPE-19 cells. (A) Quantitative western blot analysis shows that siRNA against the RB1CC1 transcript (si-RB1 and si-RB2) significantly decreased the levels of LC3-II/LC3-I ratio, with or without the presence of bafilomycin A1 (BafA), after POS feeding. As a control, cells were also treated with a nonspecific siRNA sequence (Scramble). (B) Cells treated with RB1CC1-siRNA showed increased levels of SQSTM1, with or without the presence of BafA after POS feeding. SQSTM1 levels normalized to the ACTB/actin loading control. (C) Immunofluorescence analysis showed RB1CC1 siRNA-treated ARPE-19 cells have a significantly reduced punctate pattern of LC3 staining (green), consistent with the formation of fewer autophagosomes. Treatment of cells with si-RB1 or si-RB2 alone did not induce puncta formation. (D) Quantification of the number of autophagosomes per cell.