FIGURE 4.

Accumulation of insoluble aberrantly disulfide-bonded pro-insulin in PERKi-treated Min6 cells. Immunoblots of pro-insulin recovered in lysates of untreated Min6 cells or cells exposed to PERKi (1 μm), the IRE1 inhibitor 4μ8C (30 μm), or the ER stress-inducing agent tunicamycin (Tm, 3 μg/ml) for the indicated period prior to lysis are shown. A, immunoblot of pro-insulin content of the total lysate, resolved on a reducing SDS-PAGE. B, immunoblot of pro-insulin recovered from the pellet fraction of the lysate in A following ultracentrifugation through a glycerol cushion, solubilization in 8 m urea, and resolution by reducing SDS-PAGE. C, the samples in B resolved by nonreducing SDS-PAGE.