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Published online 16 October 2000. doi:10.1083/jcb.151.2.467
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© The Rockefeller University Press, 0021-9525/2000//467 $5.00
The Journal of Cell Biology, Volume 151, Number 2, , 2000 467-478

Original Article

The Triad Targeting Signal of the Skeletal Muscle Calcium Channel Is Localized in the Cooh Terminus of the {alpha}1S Subunit



Bernhard E. Fluchera, Nicole Kasielkea, and Manfred Grabnera

a Department of Biochemical Pharmacology, University of Innsbruck, A-6020 Innsbruck, Austria
Department of Physiology, University of Innsbruck, Fritz-Pregl-Str. 3, A-6020 Innsbruck, Austria.43-512-507-283643-512-507-3787

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 {alpha}1S subunit restores Ca2+ currents, EC coupling, and the normal localization of {alpha}1S in the triads. In contrast, expression of the neuronal {alpha}1A subunit gives rise to robust Ca2+ currents but not to triad localization. To identify regions in the primary structure of {alpha}1S involved in the targeting of the Ca2+ channel into the triads, chimeras of {alpha}1S and {alpha}1A were constructed, expressed in dysgenic myotubes, and their subcellular distribution was analyzed with double immunofluorescence labeling of the {alpha}1S/{alpha}1A chimeras and the ryanodine receptor. Whereas chimeras containing the COOH terminus of {alpha}1A were not incorporated into triads, chimeras containing the COOH terminus of {alpha}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 {alpha}1S. Transferring this triad targeting signal to {alpha}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

© 2000 The Rockefeller University Press

Abbreviations used in this paper: DHPR, dihydropyridine receptor; EC, excitation–contraction; GFP, green fluorescent protein; nt, nucleotides; RE, restriction enzyme; RyR, ryanodine receptor; SR, sarcoplasmic reticulum; t tubule, transverse tubule.


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