Fig. 4: GISTs intrinsically adapt to ER stress in an ATF6-dependent manner, which is beneficial for folding and overexpression of MT-KIT.

A Activation status of the three arms of the unfolded protein response (UPR) pathway (ATF6, IRE1α, and PERK) was measured by western blotting in a panel of cancer cell lines expressing WT-KIT or MT-KIT. B Confocal microscopy was performed to evaluate the nuclear localization of ATF6 in various cancer cell lines selected from the cell line panel used in A. C Cell viability was measured in GIST cells treated with various doses (0.1–5 µM) of an ER-stress inducer, thapsigargin (TG) for 24 h. D GIST cells were treated with 5 µM TG (strong ER-stressor) across various timepoints, and the expression of the three arms of the UPR pathway, a cell-death marker (CHOP), and KIT was measured by western blotting. E GIST cells were treated with 0.1 µM TG (mild ER-stressor) across various timepoints, and the expression of the markers measured in D and chaperones (HSP70, HSP90, BIP, and GRP94) was measured by western blotting. F Expression of ATF6 and chaperones was measured by western blotting after treatment with 30 µM PF429242 (PF), which blocks ATF6 cleavage. G Cell viability of GIST cells treated with 0.1 µM TG and PF was measured after 24 and 48 h. H Immunoprecipitation of GIST cell lysates was performed with a control IgG or KIT antibody, and western blotting was performed against chaperones. Error bars in C and G represent the SD of the mean of three independent experiments. One-way ANOVA with a post-hoc test was performed to compare multiple means (*p < 0.05, **p < 0.01, ***p < 0.001).