Critical determinants of Ca2+-dependent inactivation within an EF-hand motif of L-type Ca2+ channels

Blaise Z. Peterson, Joanna S. Lee, Jennifer G. Mulle, Van Wang, Marita De Leon, David T. Yue

Research output: Contribution to journalArticle

166 Citations (Scopus)

Abstract

L-type (α(1c)) calcium channels inactivate rapidly in response to localized elevation of intracellular Ca2+, providing negative Ca2+ feedback in a diverse array of biological contexts. The dominant Ca2+ sensor for such Ca2+dependent inactivation has recently been identified as calmodulin, which appears to be constitutively tethered to the channel complex. This Ca2+ sensor induces channel inactivation by Ca2+-dependent CaM binding to an IQ-like motif situated on the carboxyl tail of α1c. Apart from the IQ region, another crucial site for Ca2+ inactivation appears to be a consensus Ca2+-binding, EF-hand motif, located ~100 amino acids upstream on the carboxyl terminus. However, the importance of this EF-hand motif for channel inactivation has become controversial since the original report from our lab implicating a critical role for this domain. Here, we demonstrate not only that the consensus EF hand is essential for Ca2+ inactivation, but that a four-amino acid cluster (VVTL) within the F helix of the EF-hand motif is itself essential for Ca2+ inactivation. Mutating these amino acids to their counterparts in non-inactivating α(1E) calcium channels (MYEM) almost completely ablates Ca2+ inactivation. In fact, only a single amino acid change of the second valine within this cluster to tyrosine (V1548Y) supports much of the functional knockout. However, mutations of presumed Ca2+-coordinating residues in the consensus EF hand reduce Ca2+ inactivation by only ~2-fold, fitting poorly with the EF hand serving as a contributory inactivation Ca2+ sensor, in which Ca2+ binds according to a classic mechanism. We therefore suggest that while CaM serves as Ca2+ sensor for inactivation, the EF-hand motif of α(1C) may support the transduction of Ca2+-CaM binding into channel inactivation. The proposed transduction role for the consensus EF hand is compatible with the detailed Ca2+-inactivation properties of wild-type and mutant V1548Y channels, as gauged by a novel inactivation model incorporating multivalent Ca2+ binding of CaM.

Original languageEnglish (US)
Pages (from-to)1906-1920
Number of pages15
JournalBiophysical journal
Volume78
Issue number4
DOIs
StatePublished - Apr 2000

Fingerprint

EF Hand Motifs
Amino Acids
L-Type Calcium Channels
Valine
Calmodulin
Calcium Channels
Tyrosine
Mutation

All Science Journal Classification (ASJC) codes

  • Biophysics

Cite this

Peterson, Blaise Z. ; Lee, Joanna S. ; Mulle, Jennifer G. ; Wang, Van ; De Leon, Marita ; Yue, David T. / Critical determinants of Ca2+-dependent inactivation within an EF-hand motif of L-type Ca2+ channels. In: Biophysical journal. 2000 ; Vol. 78, No. 4. pp. 1906-1920.
@article{ced67ce423b949d4b638673ba12dc0f2,
title = "Critical determinants of Ca2+-dependent inactivation within an EF-hand motif of L-type Ca2+ channels",
abstract = "L-type (α(1c)) calcium channels inactivate rapidly in response to localized elevation of intracellular Ca2+, providing negative Ca2+ feedback in a diverse array of biological contexts. The dominant Ca2+ sensor for such Ca2+dependent inactivation has recently been identified as calmodulin, which appears to be constitutively tethered to the channel complex. This Ca2+ sensor induces channel inactivation by Ca2+-dependent CaM binding to an IQ-like motif situated on the carboxyl tail of α1c. Apart from the IQ region, another crucial site for Ca2+ inactivation appears to be a consensus Ca2+-binding, EF-hand motif, located ~100 amino acids upstream on the carboxyl terminus. However, the importance of this EF-hand motif for channel inactivation has become controversial since the original report from our lab implicating a critical role for this domain. Here, we demonstrate not only that the consensus EF hand is essential for Ca2+ inactivation, but that a four-amino acid cluster (VVTL) within the F helix of the EF-hand motif is itself essential for Ca2+ inactivation. Mutating these amino acids to their counterparts in non-inactivating α(1E) calcium channels (MYEM) almost completely ablates Ca2+ inactivation. In fact, only a single amino acid change of the second valine within this cluster to tyrosine (V1548Y) supports much of the functional knockout. However, mutations of presumed Ca2+-coordinating residues in the consensus EF hand reduce Ca2+ inactivation by only ~2-fold, fitting poorly with the EF hand serving as a contributory inactivation Ca2+ sensor, in which Ca2+ binds according to a classic mechanism. We therefore suggest that while CaM serves as Ca2+ sensor for inactivation, the EF-hand motif of α(1C) may support the transduction of Ca2+-CaM binding into channel inactivation. The proposed transduction role for the consensus EF hand is compatible with the detailed Ca2+-inactivation properties of wild-type and mutant V1548Y channels, as gauged by a novel inactivation model incorporating multivalent Ca2+ binding of CaM.",
author = "Peterson, {Blaise Z.} and Lee, {Joanna S.} and Mulle, {Jennifer G.} and Van Wang and {De Leon}, Marita and Yue, {David T.}",
year = "2000",
month = "4",
doi = "10.1016/S0006-3495(00)76739-7",
language = "English (US)",
volume = "78",
pages = "1906--1920",
journal = "Biophysical Journal",
issn = "0006-3495",
publisher = "Biophysical Society",
number = "4",

}

Critical determinants of Ca2+-dependent inactivation within an EF-hand motif of L-type Ca2+ channels. / Peterson, Blaise Z.; Lee, Joanna S.; Mulle, Jennifer G.; Wang, Van; De Leon, Marita; Yue, David T.

In: Biophysical journal, Vol. 78, No. 4, 04.2000, p. 1906-1920.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Critical determinants of Ca2+-dependent inactivation within an EF-hand motif of L-type Ca2+ channels

AU - Peterson, Blaise Z.

AU - Lee, Joanna S.

AU - Mulle, Jennifer G.

AU - Wang, Van

AU - De Leon, Marita

AU - Yue, David T.

PY - 2000/4

Y1 - 2000/4

N2 - L-type (α(1c)) calcium channels inactivate rapidly in response to localized elevation of intracellular Ca2+, providing negative Ca2+ feedback in a diverse array of biological contexts. The dominant Ca2+ sensor for such Ca2+dependent inactivation has recently been identified as calmodulin, which appears to be constitutively tethered to the channel complex. This Ca2+ sensor induces channel inactivation by Ca2+-dependent CaM binding to an IQ-like motif situated on the carboxyl tail of α1c. Apart from the IQ region, another crucial site for Ca2+ inactivation appears to be a consensus Ca2+-binding, EF-hand motif, located ~100 amino acids upstream on the carboxyl terminus. However, the importance of this EF-hand motif for channel inactivation has become controversial since the original report from our lab implicating a critical role for this domain. Here, we demonstrate not only that the consensus EF hand is essential for Ca2+ inactivation, but that a four-amino acid cluster (VVTL) within the F helix of the EF-hand motif is itself essential for Ca2+ inactivation. Mutating these amino acids to their counterparts in non-inactivating α(1E) calcium channels (MYEM) almost completely ablates Ca2+ inactivation. In fact, only a single amino acid change of the second valine within this cluster to tyrosine (V1548Y) supports much of the functional knockout. However, mutations of presumed Ca2+-coordinating residues in the consensus EF hand reduce Ca2+ inactivation by only ~2-fold, fitting poorly with the EF hand serving as a contributory inactivation Ca2+ sensor, in which Ca2+ binds according to a classic mechanism. We therefore suggest that while CaM serves as Ca2+ sensor for inactivation, the EF-hand motif of α(1C) may support the transduction of Ca2+-CaM binding into channel inactivation. The proposed transduction role for the consensus EF hand is compatible with the detailed Ca2+-inactivation properties of wild-type and mutant V1548Y channels, as gauged by a novel inactivation model incorporating multivalent Ca2+ binding of CaM.

AB - L-type (α(1c)) calcium channels inactivate rapidly in response to localized elevation of intracellular Ca2+, providing negative Ca2+ feedback in a diverse array of biological contexts. The dominant Ca2+ sensor for such Ca2+dependent inactivation has recently been identified as calmodulin, which appears to be constitutively tethered to the channel complex. This Ca2+ sensor induces channel inactivation by Ca2+-dependent CaM binding to an IQ-like motif situated on the carboxyl tail of α1c. Apart from the IQ region, another crucial site for Ca2+ inactivation appears to be a consensus Ca2+-binding, EF-hand motif, located ~100 amino acids upstream on the carboxyl terminus. However, the importance of this EF-hand motif for channel inactivation has become controversial since the original report from our lab implicating a critical role for this domain. Here, we demonstrate not only that the consensus EF hand is essential for Ca2+ inactivation, but that a four-amino acid cluster (VVTL) within the F helix of the EF-hand motif is itself essential for Ca2+ inactivation. Mutating these amino acids to their counterparts in non-inactivating α(1E) calcium channels (MYEM) almost completely ablates Ca2+ inactivation. In fact, only a single amino acid change of the second valine within this cluster to tyrosine (V1548Y) supports much of the functional knockout. However, mutations of presumed Ca2+-coordinating residues in the consensus EF hand reduce Ca2+ inactivation by only ~2-fold, fitting poorly with the EF hand serving as a contributory inactivation Ca2+ sensor, in which Ca2+ binds according to a classic mechanism. We therefore suggest that while CaM serves as Ca2+ sensor for inactivation, the EF-hand motif of α(1C) may support the transduction of Ca2+-CaM binding into channel inactivation. The proposed transduction role for the consensus EF hand is compatible with the detailed Ca2+-inactivation properties of wild-type and mutant V1548Y channels, as gauged by a novel inactivation model incorporating multivalent Ca2+ binding of CaM.

UR - http://www.scopus.com/inward/record.url?scp=0034031414&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0034031414&partnerID=8YFLogxK

U2 - 10.1016/S0006-3495(00)76739-7

DO - 10.1016/S0006-3495(00)76739-7

M3 - Article

C2 - 10733970

AN - SCOPUS:0034031414

VL - 78

SP - 1906

EP - 1920

JO - Biophysical Journal

JF - Biophysical Journal

SN - 0006-3495

IS - 4

ER -