The II-III Loop of the Skeletal Muscle Dihydropyridine Receptor Is Responsible for the Bi-directional Coupling with the Ryanodine Receptor

doi:10.1074/jbc.274.31.21913

The II-III Loop of the Skeletal Muscle Dihydropyridine Receptor Is Responsible for the Bi-directional Coupling with the Ryanodine Receptor*

From the Department of Anatomy and Neurobiology College of Veterinary Medicine and Biomedical Sciences, Colorado State University, Fort Collins, Colorado 80523

Abstract

The dihydropyridine receptor (DHPR) in the skeletal muscle plasmalemma functions as both voltage-gated Ca²⁺ channel and voltage sensor for excitation-contraction (EC) coupling. As voltage sensor, the DHPR regulates intracellular Ca²⁺ release via the skeletal isoform of the ryanodine receptor (RyR-1). Interaction with RyR-1 also feeds back to increase the Ca²⁺ current mediated by the DHPR. To identify regions of the DHPR important for receiving this signal from RyR-1, we expressed in dysgenic myotubes a chimera (SkLC) having skeletal (Sk) DHPR sequence except for a cardiac (C) II-III loop (L). Tagging with green fluorescent protein (GFP) enabled identification of expressing myotubes. Dysgenic myotubes expressing GFP-SkLC or SkLC lacked EC coupling and had very small Ca²⁺currents. Introducing a short skeletal segment (α_1Sresidues 720–765) into the cardiac II-III loop (replacing α_1C residues 851–896) of GFP-SkLC restored both EC coupling and Ca²⁺ current densities like those of the wild type skeletal DHPR. This 46-amino acid stretch of skeletal sequence was recently shown to be capable of transferring strong, skeletal-type EC coupling to an otherwise cardiac DHPR (Nakai, J., Tanabe, T., Konno, T., Adams, B., and Beam, K.G. (1998) J. Biol. Chem.273, 24983–24986). Thus, this segment of the skeletal II-III loop contains a motif required for both skeletal-type EC coupling and RyR-1-mediated enhancement of Ca²⁺ current.

Footnotes

↵* This work was supported by a Schrödinger scholarship from the Fonds zur Förderung der Wissenschaftlichen Forschung, Austria (J01242-GEN) (to M. G.), by the Muscular Dystrophy Association (to R. T. D.), by a long term fellowship from the Human Frontier Science Program (to N. S.), and by National Institutes of Health Grants NS 24444 and AR 44750 (to K. G. B.).The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵‡ To whom correspondence and reprint requests should be addressed. Tel.: 970-491-5277; Fax: 970-491-7907; E-mail: kbeam@lamar.colostate.edu.
Abbreviations:

EC

excitation-contraction

DHPR

dihydropyridine receptor

RyR-1

skeletal ryanodine receptor

GFP

green fluorescent protein

SR

sarcoplasmic reticulum

nt

nucleotide number

Ω

ohms

F

farads

S

siemens

C

coulombs
- Received October 26, 1998.
The American Society for Biochemistry and Molecular Biology, Inc.