FIGURE 6.

XN suppresses clustering of the SCAP/SREBP complex into COP II vesicles. A, CHO-7 cells were depleted of sterols by incubating in medium F for 16 h. The cells were then switched to medium F in the presence or absence of 50 μm XN. After incubation for 3 h, the cell lysates were precleared with control beads and then incubated with control or XN beads. The pelleted beads were washed, and the eluted proteins were subjected to immunoblotting with anti-Sec23 (recognize Sec23A and Sec23B), anti-Sec24C, anti-SCAP, or anti-SREBP-1 (2A4) antibodies. B, CHO-7 cells were depleted of sterols by incubating in medium F for 16 h. The cells were then switched to medium F in the presence of vehicle, 1 μg/ml 25-HC, or 30 μm XN. After incubation for 3 h, cell lysates were subjected to immunoprecipitation with anti-SCAP antibody. The immunoprecipitates were subjected to immunoblotting (IB) with anti-Sec23, anti-SCAP, or anti-SREBP-1 (2A4) antibodies. The same results were obtained in more than three separate experiments.