Journal article
Involvement of a Heptad Repeat in the Carboxyl Terminus of the Dihydropyridine Receptor β1a Subunit in the Mechanism of Excitation-Contraction Coupling in Skeletal Muscle
Biophysical journal, Vol.87(2), pp.929-942
08/2004
DOI: 10.1529/biophysj.104.043810
PMCID: PMC1304501
PMID: 15298900
Abstract
Chimeras consisting of the homologous skeletal dihydropyridine receptor (DHPR)
β
1a subunit and the heterologous cardiac/brain
β
2a subunit were used to determine which regions of
β
1a were responsible for the skeletal-type excitation-contraction (EC) coupling phenotype. Chimeras were transiently transfected in
β
1 knockout myotubes and then voltage-clamped with simultaneous measurement of confocal fluo-4 fluorescence. All chimeras expressed a similar density of DHPR charge movements, indicating that the membrane density of DHPR voltage sensors was not a confounding factor in these studies. The data indicates that a
β
1a-specific domain present in the carboxyl terminus, namely the D5 region comprising the last 47 residues (
β
1a 478–524), is essential for expression of skeletal-type EC coupling. Furthermore, the location of
β
1aD5 immediately downstream from conserved domain D4 is also critical. In contrast, chimeras in which
β
1aD5 was swapped by the D5 region of
β
2a expressed Ca
2+
transients triggered by the Ca
2+
current, or none at all. A hydrophobic heptad repeat is present in domain D5 of
β
1a (L478, V485, V492). To determine the role of this motif, residues in the heptad repeat were mutated to alanines. The triple mutant
β
1a(L478A/V485A/V492A) recovered weak skeletal-type EC coupling (Δ
F
/
F
max
= 0.4 ± 0.1 vs. 2.7 ± 0.5 for wild-type
β
1a). However, a triple mutant with alanine substitutions at positions out of phase with the heptad repeat,
β
1a(S481A/L488A/S495A), was normal (Δ
F
/
F
max
= 2.1 ± 0.4). In summary, the presence of the
β
1a-specific D5 domain, in its correct position after conserved domain D4, is essential for skeletal-type EC coupling. Furthermore, a heptad repeat in
β
1aD5 controls the EC coupling activity. The carboxyl terminal heptad repeat of
β
1a might be involved in protein-protein interactions with ryanodine receptor type 1 required for DHPR to ryanodine receptor type 1 signal transmission.
Details
- Title: Subtitle
- Involvement of a Heptad Repeat in the Carboxyl Terminus of the Dihydropyridine Receptor β1a Subunit in the Mechanism of Excitation-Contraction Coupling in Skeletal Muscle
- Creators
- David C Sheridan - Department of Physiology, University of Wisconsin School of Medicine, Madison, Wisconsin 53706Weijun Cheng - Department of Physiology, University of Wisconsin School of Medicine, Madison, Wisconsin 53706Leah Carbonneau - Department of Physiology, University of Wisconsin School of Medicine, Madison, Wisconsin 53706Chris A Ahern - Department of Physiology, University of Wisconsin School of Medicine, Madison, Wisconsin 53706Roberto Coronado - Department of Physiology, University of Wisconsin School of Medicine, Madison, Wisconsin 53706
- Resource Type
- Journal article
- Publication Details
- Biophysical journal, Vol.87(2), pp.929-942
- Publisher
- Biophysical Society
- DOI
- 10.1529/biophysj.104.043810
- PMID
- 15298900
- PMCID
- PMC1304501
- ISSN
- 0006-3495
- eISSN
- 1542-0086
- Language
- English
- Date published
- 08/2004
- Academic Unit
- Molecular Physiology and Biophysics; Iowa Neuroscience Institute
- Record Identifier
- 9984070405802771
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