Additional file 9: of Network approach identifies Pacer as an autophagy protein involved in ALS pathogenesis

Figure S6. Pacer depletion results in detergent insoluble SOD1 aggregate accumulation. a-b, Densiometric quantification of p62 and Beclin1 levels in the autophagic flux as shown in Fig. 4a. NSC34 cells depleted of Pacer and a scrambled shRNA control (shCtrl) construct were compared (n=3). â-Actin 48 served as a loading control. Statistical analyses were performed using one-way ANOVA and Bonferroni’s post-hoc tests. Mean and SEM are shown. c, NSC34 cells depleted of Pacer were transiently co-transfected with expression vectors for human wild-type or mutant SOD1G93A fused to EGFP. When indicated, human Pacer (hPacer-V5) was co-expressed. NP40-detergent insoluble protein aggregates (NP40 insoluble) were prepared as described in materials and methods. The input is shown as a reference. â-Actin levels in the input serve as a loading control. d-e, Alignment of d shRNA A and e shRNA B to the corresponding target region in mRNA of mouse Pacer and human Pacer. 100% identity between shRNAs A and B to their respective mouse Pacer target sequences. No significant similarity is found to the corresponding region of human Pacer mRNA. (PPTX 2110 kb