Fig. 3
From: CaVβ-subunit dependence of forward and reverse trafficking of CaV1.2 calcium channels
CaV1.2 forward trafficking and recycling are not CaVβ-subtype dependent. a Confocal images of tsA-201 cells expressing CaV1.2-BBS and labelled with BTX-AF488 (top panels). CaV1.2-BBS was co-expressed with CaVα2δ-1 and CaVβ1b. Cells were incubated at 17 °C with untagged BTX for 30 min and then incubated at 37 °C with BTX-AF488. The cells were fixed at different time point after incubation at 37 °C, from zero (T0) to 40 min (T40). The cells were then permeabilized and stained with a rabbit anti-CaV1.2 Ab and a secondary Ab anti-rabbit AF594 (bottom panels). Scale bar 20 µm. b Time course of insertion of CaV1.2-BBS at the cell surface when co-expressed with CaVα2δ-1 and either CaVβ1b (filled circle), CaVβ2a (open circle) or empty vector (open triangle). The results are shown as the mean ± SEM (n numbers correspond to independent experiments). Data were fitted with single exponentials. The time constants of the fits were 16.8 ± 12.6 min and 13.0 ± 14.2 min for CaVβ1b and CaVβ2a (n = 7), respectively. c and d Effect of Brefeldin A (BFA) treatment on CaV1.2 forward trafficking. tsA-201 cells were co-transfected with CaV1.2-BBS, CaVα2δ-1 and either CaVβ1b (c) or CaVβ2a (d). Cells were treated with BFA for 4 h before undergoing the forward trafficking protocol described in a. BTX-AF488 fluorescence was normalized to the mean fluorescence at T40 for the control condition (open circle). The results are shown as the mean ± SEM. The n numbers correspond to independent experiments (average fluorescence from at least 25 cells per time point). The data were compared using an unpaired t-test. The data were fitted with single exponentials. The time constants of the fits were 10.6 ± 6.6 min and 11.6 ± 8.1 min for CaVβ1b and CaVβ2a (n = 7), respectively