Fig. 1

Fig. 1

DHPR?1 but not RyR1 is reduced in T-tubule striations of stac3 mutants. (A) Immunofluorescence labeling of WT sibling and stac3^?/? disassociated myotubes with anti-pan RyR (34c). (B) Mean immunofluorescence intensity of anti-RyR in stac3^?/? compared with WT siblings showing no difference in triadic RyR (t test, P = 0.89, n = 85 WT sibling, n = 50 stac3^?/?). a.u., arbitrary units. (C) Immunofluorescence labeling of WT sibling and stac3^?/? disassociated myotubes with mAb1 1A against a cytoplasmic region of DHPR?_1S (15). (D) Mean mAb1 1A labeling in WT siblings and stac3^?/? showing a decrease in triadic DHPR?_1S (t test, ***P < 0.0001, n = 216 WT sibling, n = 264 stac3^?/?). (E) Quantification of the mean of immunofluorescence labeling of anti-DHPR?_1S in stac3^?/? expressing stac3^NAM (NAM rescue) at triads compared with stac3^?/? expressing stac3^WT (WT rescue) (n = 75 stac3^?/?; stac3^WT, n = 53 stac3^?/?; stac3^NAM, t test, ****P < 0.0001). (F) Time course for FRAP of EGFP-DHPR?_1S expressed in WT siblings and stac3^?/? myofibers. Shown are EGFP-DHPR?_1S before (prebleach), after photobleaching (T = 0, 5, 34 min), and a maximum projection (stack) of T = 30 to 34 min (Right). (G) Mean quantification of the time course of FRAP in WT siblings (thick green line and circles) and stac3^?/? (thick red line and circles). Thin lines represent nonlinear regressions from individual traces of FRAPs from WT siblings (green) and stac3^?/? (red). The vertical thick green line depicts bleaching. (H) Histogram showing that the diffusion rate (D) of EGFP-DHPR?_1S is higher in stac3^?/? (t test, ***P < 0.0001, n = 33 stac3^?/?, n = 45 WT siblings). SEMs are indicated. (Scale bars, 2 ?m.)