Abstract
Excitation-contraction coupling in skeletal muscle is believed to take place at the triads. Therefore, precise knowledge about the triadic structure and the molecular interactions involved in junction formation are essential to understand the mechanism of excitation-contraction coupling. Triads were isolated from rabbit skeletal muscle and subjected to previously recognized protocols for disruption of the triad junction: French press treatment, hypertonic salt treatment and proteolytic cleavage. Ligand binding assays for estimations of junctional breakage and identification of the subcomponents of the triads indicated that the isolated triads are a mixture of breakage-susceptible "weak" and breakage-resistant "strong" triads. Compared to the weak triads, the strong triads are enriched in the dihydropyridine receptor. Thin section views of the strong triads frequently reveal pillars and filamentous structures, in addition to the presence of the feet, joining the transverse tubule and the terminal cisternae. These biochemical and morphological observations raise the question whether the foot structure is the only true bridging structure in the triad junction.Recently, Brandt et al. (20) reported that the dihydropyridine receptor and the junctional foot protein (JFP), which have been recognized as a voltage sensor of the transverse tubule and a Ca release channel, respectively, do not directly bind to each other. Instead, an intrinsic terminal cisternae protein of M$\sb{\rm r}$ 95,000, interacting specifically with the dihydropyridine receptor, was identified. In the present study, ($\sp{125}$I) JFP overlay onto muscle organelles showed that the JFP also binds to this 95 kDa protein selectively localized in the terminal cisternae. The 95 kDa protein was purified to 87% purity by sequential column chromatography. The isolated 95 kDa protein showed the specific interaction with the dihydropyridine receptor and the JFP observed by the 95 kDa protein-affinity chromatography.These observations suggest that the 95 kDa protein links the dihydropyridine receptor to the JFP, forming a ternary complex of dihyropyridine receptor-95 kDa protein-JFP. This bridge is likely to be the structural basis for the strong triads and the functional pathway for excitation-contraction coupling in skeletal muscle.