Fig. 8
JIP4-dependent retrograde lysosomal transport in response to acute oxidative stress. a, b MEFs were incubated with 20 µM curcumin for the indicated times. Cells were fixed, permeabilized, and immunostained with antibodies against LAMP-1 (a) or JIP4 and p150Glued (b). c Quantification of cells with JIP4 recruitment to lysosomes in DMSO or curcumin treated MEFs. DMSO n = 745, curcumin n = 805 from three independent experiments. Error bars denote s.e.m. P value calculated using two-tailed t-test ****P < 0.0001. d MEFs treated with control or JIP4 siRNAs and incubated with curcumin for 4 h. Cells were fixed, permeabilized, and immunostained with antibody against LAMP-1. e Immunoblot of U2OS expressing GFP-TMEM55B cells untreated or treated with 150 µM NaAsO2. f Immunoblot of lysates from MEFs treated with curcumin for the indicated times. g MEFs were pre-incubated with MAPK inhibitors, followed by combination of inhibitors and curcumin. Cells were fixed, permeabilized, and immunostained with antibody against LAMP-1. h Quantification of cells with JIP4 recruitment to lysosomes in curcumin/MAPK inhibitor-treated MEFs. Control n = 568, curcumin n = 583, curcumin + p38 inhibitor (20 µM) n = 598, curcumin + JNK inhibitor (25 µM) n = 590, curcumin + ERK inhibitor (20 µM) n = 575 from two independent experiments. Error bars denote s.e.m. P value calculated using one-way ANOVA ***P < 0.001. Scale bars, 20 μm
