Ca2+-dependent regulation of sodium channels NaV1.4 and NaV1.5 is controlled by the post-IQ motif

Yoder, Jesse; Ben-Johny, Manu; Farinelli, Federica; Srinivasan, Lakshmi; Shoemaker, Sophie R.; Tomaselli, Gordon F.; Gabelli, Sandra B.; Amzel, L.M.

doi:10.1038/s41467-019-09570-7

Cited by 33 publications

(61 citation statements)

References 40 publications

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“…During review of this manuscript, a paper was published describing complexes of apoCaM and Ca 2+ /CaM with the Na 1.4 CT, containing both the EF-hand domain and IQ domain (61). The authors found both the apoC-lobe and Ca 2+ /C-lobe to bind to a site nearly identical to the previously identified apoC-lobe binding site in Na V 1.5.…”

Section: Discussionmentioning

confidence: 89%

Crystal structures of Ca ²⁺ –calmodulin bound to Na _V C-terminal regions suggest role for EF-hand domain in binding and inactivation

Gardill

Rivera-Acevedo

Tung

et al. 2019

Proc. Natl. Acad. Sci. U.S.A.

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Voltage-gated sodium (Na V ) and calcium channels (Ca V ) form targets for calmodulin (CaM), which affects channel inactivation properties. A major interaction site for CaM resides in the Cterminal (CT) region, consisting of an IQ domain downstream of an EF-hand domain. We present a crystal structure of fully Ca 2+occupied CaM, bound to the CT of Na V 1.5. The structure shows that the C-terminal lobe binds to a site ∼90°rotated relative to a previous site reported for an apoCaM complex with the Na V 1.5 CT and for ternary complexes containing fibroblast growth factor homologous factors (FHF). We show that the binding of FHFs forces the EF-hand domain in a conformation that does not allow binding of the Ca 2+ -occupied C-lobe of CaM. These observations highlight the central role of the EF-hand domain in modulating the binding mode of CaM. The binding sites for Ca 2+ -free and Ca 2+ -occupied CaM contain targets for mutations linked to long-QT syndrome, a type of inherited arrhythmia. The related Na V 1.4 channel has been shown to undergo Ca 2+ -dependent inactivation (CDI) akin to Ca V s. We present a crystal structure of Ca 2+ /CaM bound to the Na V 1.4 IQ domain, which shows a binding mode that would clash with the EF-hand domain. We postulate the relative reorientation of the EFhand domain and the IQ domain as a possible conformational switch that underlies CDI.

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Section: Discussionmentioning

confidence: 89%

Crystal structures of Ca ²⁺ –calmodulin bound to Na _V C-terminal regions suggest role for EF-hand domain in binding and inactivation

Gardill

Rivera-Acevedo

Tung

et al. 2019

Proc. Natl. Acad. Sci. U.S.A.

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“…2019; Yoder et al . 2019). Our structural studies of CaM disease mutants show that different mutations have distinct effects on the structure in the presence of the Ca V 1.2 IQ domain, despite the fact that most, but not all, have a similar functional outcome on CDI: Permanent loss of Ca 2+ binding to EF3, resulting in a distorted C‐lobe.…”

Section: Discussionmentioning

confidence: 99%

Arrhythmia mutations in calmodulin can disrupt cooperativity of Ca²⁺ binding and cause misfolding

Wang

Brohus

Holt

et al. 2020

The Journal of Physiology

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Key points Mutations in the calmodulin protein (CaM) are associated with arrhythmia syndromes. This study focuses on understanding the structural characteristics of CaM disease mutants and their interactions with the voltage‐gated calcium channel CaV1.2. Arrhythmia mutations in CaM can lead to loss of Ca2+ binding, uncoupling of Ca2+ binding cooperativity, misfolding of the EF‐hands and altered affinity for the calcium channel. These results help us to understand how different CaM mutants have distinct effects on structure and interactions with protein targets to cause disease. Abstract Calmodulinopathies are life‐threatening arrhythmia syndromes that arise from mutations in calmodulin (CaM), a calcium sensing protein whose sequence is completely conserved across all vertebrates. These mutations have been shown to interfere with the function of cardiac ion channels, including the voltage‐gated Ca2+ channel CaV1.2 and the ryanodine receptor (RyR2), in a mutation‐specific manner. The ability of different CaM disease mutations to discriminate between these channels has been enigmatic. We present crystal structures of several C‐terminal lobe mutants and an N‐terminal lobe mutant in complex with the CaV1.2 IQ domain, in conjunction with binding assays and complementary structural biology techniques. One mutation (D130G) causes a pathological conformation, with complete separation of EF‐hands within the C‐lobe and loss of Ca2+ binding in EF‐hand 4. Another variant (Q136P) has severely reduced affinity for the IQ domain, and shows changes in the CD spectra under Ca2+‐saturating conditions when unbound to the IQ domain. Ca2+ binding to a pair of EF‐hands normally proceeds with very high cooperativity, but we found that N98S CaM can adopt different conformations with either one or two Ca2+ ions bound to the C‐lobe, possibly disrupting the cooperativity. An N‐lobe variant (N54I), which causes severe stress‐induced arrhythmia, does not show any major changes in complex with the IQ domain, providing a structural basis for why this mutant does not affect function of CaV1.2. These findings show that different CaM mutants have distinct effects on both the CaM structure and interactions with protein targets, and act via distinct pathological mechanisms to cause disease.

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“…In a previous report, the CTerm in both Nav1.4 and Nav1.5 showed functional alteration in a Ca 2+ /CAM‐dependent manner, and disease mutations in CTerm substantially diminished Ca 2+ ‐dependent regulation . In addition, a recent study demonstrated that Ca 2+ ‐dependent inactivation in Nav1.4 could be regulated through an affinity between EFL and CaM . Although not investigated in this study, the possibility of Ca 2+ ‐dependence or CaM expression‐dependence is an important subject that needs to be explored in future studies.…”

Section: Discussionmentioning

confidence: 77%

“…Recently, the structure of human Nav1.4 in complex with the β1 subunit was elucidated by cryo‐EM, but the CTerm structure has not yet been solved. However, the crystal structure of the CTerm 168 residues alone was elucidated, showing the structural components of the CTerm in detail. The CTerm is composed of an EF hand‐like motif (EFL) consisting of helices αI‐αV followed by a long α‐helix (helix αVI).…”

Section: Discussionmentioning

confidence: 99%

“…For example, some studies have suggested that an IFM (Ile‐Phe‐Met) residue in the linker between domains DIII and DIV (DIII‐DIV linker) is the putative fast inactivation gate . CTerm consists of a five‐helix EF hand‐like motif (EFL, helices αI‐αV) followed by a long α‐helix (helix αVI) (Figure A) . An IQ (isoleucine‐glutamine) motif in helix αVI shows high‐affinity for calmodulin (CaM) in Ca 2+ ‐dependent inactivation .…”

Section: Introductionmentioning

confidence: 99%

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EF hand‐like motif mutations of Nav1.4 C‐terminus cause myotonic syndrome by impairing fast inactivation

et al. 2020

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Introduction: Mutations of the voltage-gated sodium channel gene (SCN4A), which encodes Nav1.4, cause nondystrophic myotonia that occasionally is associated with severe apnea and laryngospasm. There are case reports of nondystrophic myotonia due to mutations in the C-terminal tail (CTerm) of Nav1.4, but the functional analysis is scarce.Methods: We present two families with nondystrophic myotonia harboring a novel heterozygous mutation (E1702del) and a known heterozygous mutation (E1702K). Results:The proband with E1702K exhibited repeated rhabdomyolysis, and the daughter showed laryngospasm and cyanosis. Functional analysis of the two mutations as well as another known heterozygous mutation (T1700_E1703del), all located on EF hand-like motif in CTerm, was conducted with whole-cell recording of heterologously expressed channel. All mutations displayed impaired fast inactivation.

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Ca2+-dependent regulation of sodium channels NaV1.4 and NaV1.5 is controlled by the post-IQ motif

Cited by 33 publications

References 40 publications

Crystal structures of Ca ²⁺ –calmodulin bound to Na _V C-terminal regions suggest role for EF-hand domain in binding and inactivation

Crystal structures of Ca ²⁺ –calmodulin bound to Na _V C-terminal regions suggest role for EF-hand domain in binding and inactivation

Arrhythmia mutations in calmodulin can disrupt cooperativity of Ca²⁺ binding and cause misfolding

EF hand‐like motif mutations of Nav1.4 C‐terminus cause myotonic syndrome by impairing fast inactivation

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Ca2+-dependent regulation of sodium channels NaV1.4 and NaV1.5 is controlled by the post-IQ motif

Cited by 33 publications

References 40 publications

Crystal structures of Ca 2+ –calmodulin bound to Na V C-terminal regions suggest role for EF-hand domain in binding and inactivation

Crystal structures of Ca 2+ –calmodulin bound to Na V C-terminal regions suggest role for EF-hand domain in binding and inactivation

Arrhythmia mutations in calmodulin can disrupt cooperativity of Ca2+ binding and cause misfolding

EF hand‐like motif mutations of Nav1.4 C‐terminus cause myotonic syndrome by impairing fast inactivation

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Crystal structures of Ca ²⁺ –calmodulin bound to Na _V C-terminal regions suggest role for EF-hand domain in binding and inactivation

Crystal structures of Ca ²⁺ –calmodulin bound to Na _V C-terminal regions suggest role for EF-hand domain in binding and inactivation

Arrhythmia mutations in calmodulin can disrupt cooperativity of Ca²⁺ binding and cause misfolding