Fig. 3: p97 and the Hsp70 system mediate aggresome disassembly.

a HeLa cells treated with Btz (1 μM, 8 h) were fixed after 15 h recovery or recovered for six additional hours in the absence (Ctrl) or presence of 100 nM Baf, 1 μM Btz, 1 μM CB-5083, or 10 μM VER, followed by immunostaining against ubiquitin and LC3. Scale bars, 50 μm. b Quantification of the percentage of cells with an aggresome in a. Shown is the mean ± SD from n = 4 biological replicates with ≥74 cells per condition and replicate. One-way ANOVA. c Same as b, but quantification of total aggregate area. d HeLa cells treated as in b or left untreated (–) were lysed in RIPA buffer. RIPA soluble and insoluble fractions were immunoblotted for ubiquitin, and the Coomassie staining of the membrane served as a loading control (CBB). Representative result from five independent experiments. e HeLa wildtype (WT) and DNAJB6 KO cells treated with Btz (1 μM, 8 h) were fixed after 15 h recovery or recovered for six additional hours in the absence or presence of 1 μM CB-5083, followed by immunostaining against ubiquitin and LC3. Scale bars, 50 μm. f Quantification of the percentage of cells with an aggresome in e. Shown is the mean ± SD from n = 3 biological replicates with ≥100 cells per condition and replicate. Two-way ANOVA. g Lysates from control and DNAJB6 KO cells were immunoblotted for DNAJB6 (isoforms a and b) to validate KO efficiency. Representative result from two independent experiments.