Figure 5

Expression of insulin 2 (C96Y)-EGFP induces the mRNA and protein of known ER stress response genes. Clone #4S2 cells were untreated (Control) or treated with 2 μg/ml doxycycline (+Dox) for the times indicated (h, hours; d, days). A-D. Total RNA was isolated and the mRNA levels relative to cellular β-actin for Grp78 (A), Pdi (B), Chop (C), Sel1 (D) were analyzed by real-time PCR as outlined in the methods. Shown are the mean ± SE of 3 independent experiments. *p < 0.05 relative to control. E, F. Clone #4S2 cells were treated as described above, washed in PBS, lysed and equal amounts of protein were resolved by SDS-PAGE and immunoblotted as indicated. Control INS-1 cells exposed to thapsigargin (Tg) were used as a positive control in (F).