FIGURE 8.
RIM1 interacts with native L-type channels through the CaVβ subunit and contributes to determine insulin secretion. A and B, proteins from RIN-m5F cells were immunoprecipitated with anti-CaVβ2, anti-CaVβ3, or control (IgG0) antibodies and subjected to Western blot analysis with anti-RIM antibody. The ∼180 kDa RIM1 band is visualized in the immunoprecipitation (IP) lane. In both cases, control experiments with the irrelevant antibody as a substitute for the anti-CaVβ antibodies failed to co-immunoprecipitate RIM1. Immunoprecipitation data were collected from the same experiment, and the images are shown separately because they were acquired with different time exposures. C–E, basal and high K+-induced insulin secretion from RIN-m5F cells after transfection with RIM1- and CaVβ-targeted siRNA compared with control (non-transfected) cells. RIN-m5F cells were transfected with RIM1 siRNA (open bars) and 48 h later incubated with KRB buffer containing 5 mm (low K+) or 40 mm (high K+) KCl. Insulin content in the supernatants was measured by ELISA as described under “Experimental Procedures.” The mean ± S.E. of three independent experiments is displayed.