(A and B) Cells [the WT H99 strain, ire1Δ and hxl1Δ mutants, ire1Δ+IRE1 and hxl1Δ+HXL1 complemented strains, and PH3
:KAR2 strains (YSB1751, YSB1741, and YSB1745)] were spotted on a YPD agar medium containing the indicated concentrations of DNA damage inducers, including methyl methanesulfonate (MMS; A) or hydroxyl urea (HU; B), incubated at 30oC for 2–4 days, and photographed. (C) The RT-PCR analysis of UPR-induced HXL1 splicing was performed with cDNA samples prepared from total RNA samples of the WT H99 strain and ire1Δ mutants treated with HU (90 mM) or MMS (0.03%) for 1 h. (D) Using the total RNA set from (C), Northern blot assay was performed to measure KAR2 induction levels in the WT H99 strain and ire1Δ mutants. The Northern blot membrane was hybridized with the KAR2 specific probe, washed, and developed.