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. 2006 Nov 13;27(3):1027–1043. doi: 10.1128/MCB.00408-06

FIG. 5.

FIG. 5.

Effects of ER stress inducers on oligomeric status of ATF6. (A) CHO cells were treated with 1 mM DTT, 2 μg/ml Tm, or 1 μM Tg for the indicated periods, and then cell lysates were prepared and analyzed as described for Fig. 3A(b) (with NEM). Monomer* denotes the nonglycosylated form of monomer. The same set of data for Tm treatment is shown in the left and right panels with different exposure times to emphasize the difference in the extent of ATF6α cleavage between DTT and Tg. (B) CHO cells were treated with 1 mM DTT for the indicated periods (1st DTT, lanes 1 to 5). Cells treated with 1 mM DTT for 90 min were washed and then incubated in the absence of DTT for the indicated periods (wash, lanes 6 to 8). Cells incubated without DTT for 60 min were treated again with 1 mM DTT for the indicated periods (2nd DTT, lanes 9 to 11). Cell lysates were prepared and analyzed as described for Fig. 3A(a) (without NEM). (C) CHO cells were pretreated with (+) or without (−) 300 μM AEBSF for 5 min and then treated with 1 mM DTT for the indicated periods without removing AEBSF. Cell lysates were prepared and analyzed as described for Fig. 3A(b) (with NEM). The bottom panel shows the result of shorter exposure of the region around the monomer.