Fig. 6

Cells lacking Pkh are sensitive to UPR inducers and ionic stress. a Effect of partial depletion of Pkh on the sensitivity to the UPR inducers tunicamycin and β-mercaptoethanol. Cultures of CML476 cells (WT) and its specified derivatives were serially diluted and spotted onto YPD plates containing 10 μg/ml doxycycline (DOX), or the vehicle, and the indicated concentrations of tunicamycin (Tn) or β-mercaptoethanol (β-ME). Control plate in the absence of doxycycline of cells incubated with β-ME is shown in the Fig. 5b. Cell growth was recorded after 3 days. b Left panel. β-Galactosidase activity of SDP8 cells containing the UPRE-LacZ reporter gene. SDP8 were transformed with the pMCZ-Y plasmid and incubated in medium without uracil in the presence of 10 μg/ml of doxycycline (“SDP8-Dox”) or the vehicle (“SDP8 + Dox”) for a total time of 24 h. Six h before the treatment with tunicamycin, new cells cultures at OD₆₆₀ ~ 0.05 were prepared containing fresh doxycycline. Finally, cells were resuspended in medium containing fresh doxycycline (or the vehicle) and 2.5 μg/ml tunicamycin (or DMSO). β-Galactosidase activity was measured for cells non-treated (empty bars) or treated with tunicamycin (filled bars). Data are mean ± SEM from six independent clones. Right panel: Representation of the cell density of the cultures used in the left panel obtained by measuring their absorbance at 650 nm in a microplate reader, after the treatment of SDP8 cells with tunicamycin (filled bars) or with the vehicle (empty bars). Data are mean ± SEM from six independent clones. c Effect of partial depletion of Pkh on the salts sensitivity. Four dilutions (1:5) of cells from the strains specified in a were spotted onto YPD plates containing 10 μg/ml doxycycline (DOX) and the indicated concentrations of NaCl or LiCl. Control plate for the absence of doxycycline is shown in the Fig. 5b. Cell growth was recorded after 3 days