Figure 2. IPA activates the IRE1 branch of the unfolded protein response (UPR) in HEK293T cells.
(A) HEK293T cells were treated with increasing concentrations of IPA as a function of time. Tunicamycin (Tm, 2 µg/ml) was used as positive control to induce endoplasmic reticulum stress. The resulting XBP1 mRNA spliced products detected by RT-PCR (‘u’: unspliced and ‘s’: spliced) are indicated. Control cells were treated with DMSO only. The asterisk identifies a hybrid amplicon resulting from spliced and unspliced XBP1 mRNA. (B) The effects of IPAx on the splicing of XBP1 mRNA in HEK293T cells were detected by RT-PCR after 4-hr incubation ([IPA] and [IPAx] = 2 µM). (C) Inhibition of IPA-mediated XBP1 mRNA splicing in HEK293T cells by AD60 (incubation time = 4 hr; [AD60] = 1 μM). (D) IRE1-GFP foci formation in T-REx293 cells. IRE1-GFP was visualized by confocal microscopy. (E) Effects of IPA on HEK293T cell viability (LD50 = 0.83 μM).
Figure 2—figure supplement 1. AD60 inhibition of XBP1-luciferase-splicing reporter activation in HEK293T cells.
