Molecular localization of regions in the L-type calcium channel critical for dihydropyridine action

doi:10.1016/0896-6273(93)90215-D

Neuron

Volume 11, Issue 6, December 1993, Pages 1013-1021

https://doi.org/10.1016/0896-6273(93)90215-D Get rights and content

Abstract

Sensitivity to dihydropyridines (DHPs) is a distinct characteristic that differentiates L-type Ca²⁺ channels from T-, N-, and P-type Ca²⁺ channels. To identify regions necessary for the functional effects of DHPs, chimeric Ca²⁺ channels were constructed in which portions of motif III or motif IV of a DHP-insensitive brain Ca²⁺ channel, BI-2, were introduced into the DHP-sensitive cardiac L-type Ca²⁺ channel. The resultant chimeric Ca²⁺ channels were expressed in Xenopus oocytes, and the effects of a DHP agonist and antagonist were studied. The results show that the linker region between S5 and S6 in motif IV of the L-type Ca²⁺ channel is a major site for DHP action. The DHP agonist and antagonist molecules interact with distinct sites on the α₁ subunit of the L-type Ca²⁺ channel. The data further show that the SS2–S6 region of motif III is not involved in DHP action but may be an important structural component of inactivation.

References (51)

T. Abe et al.
Identification of the receptor for ω-conotoxin in brain
J. Biol. Chem.
(1987)
M.K. Ahlijanian et al.
Subunit structure and localization of dihydropyridine-sensitive calcium channels in mammalian brain, spinal cord, and retina
Neuron
(1990)
K.P. Campbell et al.
The biochemistry and molecular biology of the dihydropyridine-sensitive calcium channel
Trends Neurosci.
(1988)
W.A. Catterall et al.
Receptor sites for Ca²⁺ channel antagonists
Trends Pharmacol. Sci.
(1992)
T. Hoshi et al.
Two types of inactivation in Shaker K channels: effects of alterations in the carboxy-terminal region
Neuron
(1991)
K. Itagaki et al.
Native-type DHP-sensitive calcium channel currents are produced by cloned rat aortic smooth muscle and cardiac α₁ subunits expressed in Xenopus laevis oocytes and are regulated by α₂- and β-subunits
FEBS Lett.
(1992)
R.A. Janis et al.
Specific binding of a Ca channel agonist to membranes from cardiac muscle and brain
Biochem. Biophys. Res. Commun.
(1984)
S.M. King et al.
Strategies and reagents for photoaffinity labeling of mechanochemical proteins
Meth. Enzymol.
(1991)
M. Kozak
Structural features in eukaryotic mRNAs that modulate the initiation of translation
J. Biol. Chem.
(1991)
P. Lory et al.
Characterization of β subunit modulation of a rabbit cardiac L-type Ca ²⁺ channel α₁ subunit as expressed in mouse L cells
FEBS Lett.
(1993)

E. Perez-Reyes et al.

Cloning and expression of a cardiac/brain β subunit of the L-type calcium channel

J. Biol. Chem.

(1992)

D.F. Slish et al.

Evidence for the existence of a cardiac specific isoform of the α₁ subunit of the voltage-dependent calcium channel

FEBS Lett.

(1989)

S. Tang et al.

Molecular localization of ion-selectivity sites within the pore of a human L-type cardiac calcium channel

J. Biol. Chem.

(1993)

R.W. Tsien et al.

Molecular diversity of voltage-dependent Ca²⁺ channels

Trends Pharmacol. Sci.

(1991)

M.E. Williams et al.

Structure and functional expression of α₁, α₂ and β subunits of a novel human neuronal calcium channel subtype

Neuron

(1992)

P.H. Backx et al.

Molecular localization of an ion-binding site within the pore of mammalian sodium channels

Science

(1992)

A.M. Brown et al.

Dual effects of dihydropyridines on whole cell and unitary calcium currents in single ventricular cells of guinea-pig

J. Physiol.

(1986)

W.A. Catterall

Structure and function of voltage-sensitive ion channels

Science

(1988)

V. Chowdry et al.

Photoaffinity labeling of biological systems

Ann. Rev. Biochem.

(1979)

T. Collin et al.

Molecular cloning of three isoforms of the L-type voltage-dependent calcium channel β subunit from normal human heart

Circ. Res.

(1993)

G.P. Dube et al.

Nitrendipine potentiates Bay K 8644-induced contraction of isolated porcine coronary artery: evidence for functionally distinct dihydropyridine receptor subtypes

Biochem. Biophys. Res. Commun.

(1985)

S.B. Ellis et al.

Sequence and expression of mRNAs encoding the α₁ and α₂ subunits of a DHP-sensitive calcium channel

Science

(1988)

Y. Fujita et al.

Primary structure and functional expression of the ω-conotoxin-sensitive N-type calcium channel from rabbit brain

Neuron

(1993)

H. Glossmann et al.

Calcium channels: introduction into their molecular pharmacology

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Neuron

ArticleMolecular localization of regions in the L-type calcium channel critical for dihydropyridine action

Abstract

J. Biol. Chem.

Neuron

Trends Neurosci.

Trends Pharmacol. Sci.

Neuron

FEBS Lett.

Biochem. Biophys. Res. Commun.

Meth. Enzymol.

J. Biol. Chem.

FEBS Lett.

J. Biol. Chem.

FEBS Lett.

J. Biol. Chem.

Trends Pharmacol. Sci.

Neuron

Molecular localization of an ion-binding site within the pore of mammalian sodium channels

Science

Dual effects of dihydropyridines on whole cell and unitary calcium currents in single ventricular cells of guinea-pig

J. Physiol.

Structure and function of voltage-sensitive ion channels

Science

Photoaffinity labeling of biological systems

Ann. Rev. Biochem.

Molecular cloning of three isoforms of the L-type voltage-dependent calcium channel β subunit from normal human heart

Circ. Res.

Nitrendipine potentiates Bay K 8644-induced contraction of isolated porcine coronary artery: evidence for functionally distinct dihydropyridine receptor subtypes

Biochem. Biophys. Res. Commun.

Sequence and expression of mRNAs encoding the α1 and α2 subunits of a DHP-sensitive calcium channel

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Primary structure and functional expression of the ω-conotoxin-sensitive N-type calcium channel from rabbit brain

Neuron

Calcium channels: introduction into their molecular pharmacology

Article
Molecular localization of regions in the L-type calcium channel critical for dihydropyridine action

Sequence and expression of mRNAs encoding the α₁ and α₂ subunits of a DHP-sensitive calcium channel