Figure 4

High-temperature cultivation of an rhIL-10 expression strain impairs ER folding capacity. (A) Total RNA was extracted from yeast cells at the 24-h time point as in Figure 1B, and the mRNA levels of primary UPR genes were examined by qRT-PCR. (B) Changes in UPR induction in a non-producing strain. The non-producing strain was methanol-induced for 24 hours before extraction of total RNA. The mRNA levels of primary UPR genes were examined by qRT-PCR. (C) Whole-cell lysates from the 24-h time point as in Figure 2A were examined by Western blotting using an anti-Kar2 antibody, with α-actin as a loading control. (D) The rhIL-10 expression strains were methanol-induced for 24 hours at 30°C in shaking flask culture. Whole-cell lysates (extracted from a volume of cells equivalent to 0.1 OD600 units) were examined by Western blotting. The labels `L,' `M' and `H' indicate 1, 5 and 10 gene copies, respectively. (E) Total RNA was extracted from the cells as in Figure 4D, and the mRNA levels of primary UPR genes were examined by qRT-PCR. The label `V' indicates the vector control. All the experiments were repeated three times and showed similar results. The statistical results presented represent the mean ± SD. (*P <0.05, **P <0.01, ***P <0.001).