Original Article

The Triad Targeting Signal of the Skeletal Muscle Calcium Channel Is Localized in the COOH Terminus of the _1S Subunit

Correspondence to: Bernhard E. Flucher, Department of Physiology, University of Innsbruck, Fritz-Pregl-Str. 3, A-6020 Innsbruck, Austria. Tel:43-512-507-3787 Fax:43-512-507-2836

The specific localization of L-type Ca²⁺ channels in skeletal muscle triads is critical for their normal function in excitation–contraction (EC) coupling. Reconstitution of dysgenic myotubes with the skeletal muscle Ca²⁺ channel _1S subunit restores Ca²⁺ currents, EC coupling, and the normal localization of _1S in the triads. In contrast, expression of the neuronal _1A subunit gives rise to robust Ca²⁺ currents but not to triad localization. To identify regions in the primary structure of _1S involved in the targeting of the Ca²⁺ channel into the triads, chimeras of _1S and _1A were constructed, expressed in dysgenic myotubes, and their subcellular distribution was analyzed with double immunofluorescence labeling of the _1S/_1A chimeras and the ryanodine receptor. Whereas chimeras containing the COOH terminus of _1A were not incorporated into triads, chimeras containing the COOH terminus of _1S were correctly targeted. Mapping of the COOH terminus revealed a triad-targeting signal contained in the 55 amino-acid sequence (1607–1661) proximal to the putative clipping site of _1S. Transferring this triad targeting signal to _1A was sufficient for targeting and clustering the neuronal isoform into skeletal muscle triads and caused a marked restoration of Ca²⁺-dependent EC coupling.

Key Words: calcium channel, dihydropyridine receptor, excitation–contraction coupling, immunofluorescence, skeletal muscle

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