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. 2012 Mar 1;23(5):758–770. doi: 10.1091/mbc.E11-08-0663

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© 2012 Chiang et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

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FIGURE 2: — IRE1 selectively decreases levels of P23H rhodopsin protein. (A) P23H rhodopsin was expressed in HEK293 cells bearing IRE1[I642G], and 1NM-PP1 (5 μM) was applied for the indicated durations. (B) Wild-type rhodopsin was expressed in HEK293 cells bearing IRE1[I642G], and 1NM-PP1 (5 μM) was applied as indicated. (A, B) Total rhodopsin protein levels were detected by immunoblotting and quantified. GAPDH levels were assessed as a protein loading control. Cotransfected GFP levels were assessed to control for transfection efficiency. Xbp-1 mRNA splicing was assessed by reverse transcription-PCR, and the amount of spliced Xbp-1 mRNA was quantified as a percentage of the total amounts of unspliced (u) and spliced (s) amplicons. P23H and wild-type rhodopsin mRNA levels were measured by real-time PCR and are expressed relative to levels in cells that were not treated with 1NM-PP1. Immunoblots are representative of three independent experiments. Error bars represent SDs from three experiments. The p values were determined by Student's t test analysis.