Figure 5.

The C147F mutant, but not WT human arrestin-1, induces UPR in 661W cells. (A) Representative Western blots for indicated proteins are shown. For BIP and arrestin, 10 μg protein per lane was loaded; for GAPDH, the blots were stripped and reprobed with mouse anti-GAPDH antibody. (B) Quantification of BIP expression in control cells (CO) cells, cells treated with known inducers of UPR 1 μM of thapsigargin (tg1) or 100 μM tunicamycin (tm100), as well as cells transfected with empty vector (pcDNA3) or pcDNA3 encoding the WT human arestin-1 (DNA per well of six-well plate: 0.56 μg, WT1; 0.64 μg, WT2) or C147F mutant (DNA per well of six-well plate: 0.64 μg, CF1; 0.75 μg, CF2). The data are presented in arbitrary units. The intensity of the bands was normalized to the sum of intensities of all eight bands on the blot in each experiment (n = 3) and multiplied by 10. The data were analyzed by 1-way ANOVA (F(7,16) = 49.4, P < 0.0001), followed by Bonferroni's post hoc comparisons to corresponding controls (CO or pcDNA3). Statistical significance of the differences is shown, as follows: ***P < 0.001, #P < 0.001 to WT1/2, $P < 0.05 to tm100.