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. 2013 Oct 1;24(19):3133–3144. doi: 10.1091/mbc.E13-05-0269

FIGURE 8:

FIGURE 8:

UPR is induced in ypt1 and trs85∆ but not ypt31∆/32ts, trs130ts, and sec7-4 mutant cells expressing Snc1-GFP. (A) Snc1-GFP accumulates in internal puncta in ypt1-1, trs85∆, ypt31∆/32ts, and trs130ts mutant cells. Cells expressing Snc1-GFP from a 2 μ plasmid were grown at 26°C (top), shifted to 37°C for 90 min (bottom), and visualized by live-cell microscopy. Representative cells are shown (a minimum of 24 cells were visualized for each panel). All mutant cells accumulate internal Snc1-GFP at 37°C and to a lesser degree at 26°C. (B) UPR is induced in ypt1-1 and trs85∆ but not ypt31∆/32ts and trs130ts mutant cells. Cells expressing Snc1-GFP (as in A) were transformed with a second plasmid carrying the LacZ gene under a UPR-inducible promoter (Kruse et al., 2006). Cells were grown at 26°C (light bars) and shifted to 37°C for 90 min (dark bars), and the induction of the UPR response was determined using the βGAL assay. UPR is expressed as percentage of the response of wild type at 26°C. The UPR response in trs85∆ and ypt1-1 mutant cells is induced at 26°C and is even higher at 37°C. No such induction is observed in ypt31∆/32ts or trs130ts mutant cells even under conditions in which these cells accumulate intracellular Snc1-GFP. (C) UPR is induced in ypt1-T40K and ypt1-T40A mutant cells expressing Snc1-GFP. UPR was determined in wild type and ypt1 mutant cells expressing LacZ under a UPR-inducible promoter and Snc1-GFP (as in B). (D) Intracellular Snc1-GFP accumulates in sec7-4 mutant cells. The experiment was done as described in A (50 cells were visualized for each panel). sec7-4 mutant cells accumulate internal Snc1-GFP at 37°C and to a much lesser degree at 26°C. (E) UPR is not induced in sec7-4 mutant cells. The experiment was done as described in B. No UPR induction is observed in sec7-4 mutant cells even under conditions in which these cells accumulate intracellular Snc1-GFP. (F) UPR can be induced in wild-type and sec7-4 mutant cells by tunicamycin. Cells were grown without tunicamycin (light bars) or with 5 μg/ml tunicamycin for 90 min (dark bars), and the induction of the UPR response was determined using the βGAL assay. Bar, 5 μm; error bars, SD (B, C, E, F). Results represent at least two independent experiments.