Skip to main content
. 2022 Apr 29;298(6):101997. doi: 10.1016/j.jbc.2022.101997

Figure 3.

Figure 3

Effects of S724A mutation upon the dynamic activation of IRE1α in MEF cells.A, MEF cells of the indicated genotypes were treated with Tm at the indicated concentrations for 24 h. Quantitative RT–PCR analysis of Xbp1 mRNA splicing and the mRNA abundance of the indicated genes. B–G, MEF cells were treated with 100 ng/ml or 10 μg/ml Tm for the indicated time intervals. B and E, immunoblot analysis of IRE1α phosphorylation and XBP1s protein. Upper Phos-tag gels with long or short exposure time are shown for the band-shift analysis of IRE1α protein phosphorylation. Tubulin was used as the loading control. C and F, agarose gel analysis of Xbp1 mRNA splicing by RT–PCR. D and G, quantitative RT–PCR analysis of Xbp1 mRNA splicing and the RIDD target Blos1 mRNA. All data are presented as the mean ± SD (n = 3 independent experiments). ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001 by two-tailed unpaired Student’s t test or two-way ANOVA. IRE1α, inositol-requiring enzyme 1α; MEF, mouse embryonic fibroblast; RIDD, regulated IRE1-dependent decay; Tm, tunicamycin; Xbp1, X-box binding protein 1; XBP1s, spliced XBP1.