Inactivation of CaV1 and CaV2 channels.

Voltage-gated Ca2+ channels (VGCCs) are highly expressed throughout numerous biological systems and play critical roles in synaptic transmission, cardiac excitation, and muscle contraction. To perform these various functions, VGCCs are highly regulated. Inactivation comprises a critical mechanism controlling the entry of Ca2+ through these channels and constitutes an important means to regulate cellular excitability, shape action potentials, control intracellular Ca2+ levels, and contribute to long-term potentiation and depression.

Voltage Gated Calcium Channel Dysregulation May Contribute to Neurological Symptoms in Calmodulinopathies.

Calmodulinopathies are caused by mutations in calmodulin (CaM), and result in debilitating cardiac arrythmias such as long-QT syndrome (LQTS) and catecholaminergic polymorphic ventricular tachycardia (CPVT). In addition, many patients exhibit neurological comorbidities, including developmental delay and autism spectrum disorder. Until now, most work into these mutations has focused on cardiac effects, identifying impairment of Ca /CaM-dependent inactivation (CDI) of Ca 1.

A Natural History Study of Timothy Syndrome.

Timothy syndrome (OMIM #601005) is a rare disease caused by variants in the gene . Timothy syndrome patients were first identified as having a cardiac presentation of Long QT and syndactyly of the fingers and/or toes, and an identical variant in , Gly406Arg. However, since this original identification, more individuals harboring diverse variants in have been identified and have presented with various cardiac and extra-cardiac symptoms.

Rapid modeling of an ultra-rare epilepsy variant in wild-type mice by in utero prime editing.

Generating animal models for individual patients within clinically-useful timeframes holds great potential toward enabling personalized medicine approaches for genetic epilepsies. The ability to rapidly incorporate patient-specific genomic variants into model animals recapitulating elements of the patient's clinical manifestations would enable applications ranging from validation and characterization of pathogenic variants to personalized models for tailoring pharmacotherapy to individual patients. Here, we demonstrate generation of an animal model of an individual epilepsy patient with an ultra-rare variant of the NMDA receptor subunit GRIN2A, without the need for germline transmission and breeding.

Calmodulin Mutations in Human Disease.

Calcium ions (Ca) are the basis of a unique and potent array of cellular responses. Calmodulin (CaM) is a small but vital protein that is able to rapidly transmit information about changes in Ca concentrations to its regulatory targets. CaM plays a critical role in cellular Ca signaling, and interacts with a myriad of target proteins.

CACNA1C-Related Channelopathies.

The CACNA1C gene encodes the pore-forming subunit of the Ca1.2 L-type Ca channel, a critical component of membrane physiology in multiple tissues, including the heart, brain, and immune system. As such, mutations altering the function of these channels have the potential to impact a wide array of cellular functions.

Impaired Ca1.2 inactivation reduces the efficacy of calcium channel blockers in the treatment of LQT8.

Mutations in the Ca1.2 L-type calcium channel can cause a profound form of long-QT syndrome known as long-QT type 8 (LQT8), which results in cardiac arrhythmias that are often fatal in early childhood. A growing number of such pathogenic mutations in Ca1.

CaV1.2 channelopathic mutations evoke diverse pathophysiological mechanisms.

The first pathogenic mutation in CaV1.2 was identified in 2004 and was shown to cause a severe multisystem disorder known as Timothy syndrome (TS). The mutation was localized to the distal S6 region of the channel, a region known to play a major role in channel activation.

Fibroblast growth factor homologous factors serve as a molecular rheostat in tuning arrhythmogenic cardiac late sodium current.

Voltage-gated sodium (Nav1.5) channels support the genesis and brisk spatial propagation of action potentials in the heart. Disruption of Na1.

The molecular basis of the inhibition of Ca1 calcium-dependent inactivation by the distal carboxy tail.

Ca/calmodulin-dependent inactivation (CDI) of Ca channels is a critical regulatory process that tunes the kinetics of Ca entry for different cell types and physiologic responses. CDI is mediated by calmodulin (CaM), which is bound to the IQ domain of the Ca carboxy tail. This modulatory process is tailored by alternative splicing such that select splice variants of Ca1.

Infanticide vs. inherited cardiac arrhythmias.

Aims: In 2003, an Australian woman was convicted by a jury of smothering and killing her four children over a 10-year period. Each child died suddenly and unexpectedly during a sleep period, at ages ranging from 19 days to 18 months. In 2019 we were asked to investigate if a genetic cause could explain the children's deaths as part of an inquiry into the mother's convictions.

A bilobal model of Ca-dependent inactivation to probe the physiology of L-type Ca channels.

L-type calcium channels (LTCCs) are critical elements of normal cardiac function, playing a major role in orchestrating cardiac electrical activity and initiating downstream signaling processes. LTCCs thus use feedback mechanisms to precisely control calcium (Ca) entry into cells. Of these, Ca-dependent inactivation (CDI) is significant because it shapes cardiac action potential duration and is essential for normal cardiac rhythm.

Allosteric regulators selectively prevent Ca-feedback of Ca and Na channels.

Calmodulin (CaM) serves as a pervasive regulatory subunit of Ca1, Ca2, and Na1 channels, exploiting a functionally conserved carboxy-tail element to afford dynamic Ca-feedback of cellular excitability in neurons and cardiomyocytes. Yet this modularity counters functional adaptability, as global changes in ambient CaM indiscriminately alter its targets. Here, we demonstrate that two structurally unrelated proteins, SH3 and cysteine-rich domain (stac) and fibroblast growth factor homologous factors (fhf) selectively diminish Ca/CaM-regulation of Ca1 and Na1 families, respectively.

A Precision Medicine Approach to the Rescue of Function on Malignant Calmodulinopathic Long-QT Syndrome.

Rationale: Calmodulinopathies comprise a new category of potentially life-threatening genetic arrhythmia syndromes capable of producing severe long-QT syndrome (LQTS) with mutations involving CALM1, CALM2, or CALM3. The underlying basis of this form of LQTS is a disruption of Ca/calmodulin (CaM)-dependent inactivation of L-type Ca channels.

Objective: To gain insight into the mechanistic underpinnings of calmodulinopathies and devise new therapeutic strategies for the treatment of this form of LQTS.

Following Optogenetic Dimerizers and Quantitative Prospects.

Optogenetics describes the use of genetically encoded photosensitive proteins to direct intended biological processes with light in recombinant and native systems. While most of these light-responsive proteins were originally discovered in photosynthetic organisms, the past few decades have been punctuated by experiments that not only commandeer but also engineer and enhance these natural tools to explore a wide variety of physiological questions. In addition, the ability to tune dynamic range and kinetic rates of optogenetic actuators is a challenging question that is heavily explored with computational methods devised to facilitate optimization of these systems.

Protein kinase A modulation of CaV1.4 calcium channels.

The regulation of L-type Ca(2+) channels by protein kinase A (PKA) represents a crucial element within cardiac, skeletal muscle and neurological systems. Although much work has been done to understand this regulation in cardiac CaV1.2 Ca(2+) channels, relatively little is known about the closely related CaV1.

An autism-associated mutation in CaV1.3 channels has opposing effects on voltage- and Ca(2+)-dependent regulation.

CaV1.3 channels are a major class of L-type Ca(2+) channels which contribute to the rhythmicity of the heart and brain. In the brain, these channels are vital for excitation-transcription coupling, synaptic plasticity, and neuronal firing.

Arrhythmogenesis in Timothy Syndrome is associated with defects in Ca(2+)-dependent inactivation.

Timothy Syndrome (TS) is a multisystem disorder, prominently featuring cardiac action potential prolongation with paroxysms of life-threatening arrhythmias. The underlying defect is a single de novo missense mutation in CaV1.2 channels, either G406R or G402S.

A rendezvous with the queen of ion channels: Three decades of ion channel research by David T Yue and his Calcium Signals Laboratory.

David T. Yue was a renowned biophysicist who dedicated his life to the study of Ca(2+) signaling in cells. In the wake of his passing, we are left not only with a feeling of great loss, but with a tremendous and impactful body of work contributed by a remarkable man.

Novel fluorescence resonance energy transfer-based reporter reveals differential calcineurin activation in neonatal and adult cardiomyocytes.

Novel fluorescence resonance energy transfer-based genetically encoded reporters of calcineurin are constructed by fusing the two subunits of calcineurin with P2A-based linkers retaining the expected native conformation of calcineurin. Calcineurin reporters display robust responses to calcium transients in HEK293 cells. The sensor responses are correlated with NFATc1 translocation dynamics in HEK293 cells.

Towards a Unified Theory of Calmodulin Regulation (Calmodulation) of Voltage-Gated Calcium and Sodium Channels.

Voltage-gated Na and Ca(2+) channels represent two major ion channel families that enable myriad biological functions including the generation of action potentials and the coupling of electrical and chemical signaling in cells. Calmodulin regulation (calmodulation) of these ion channels comprises a vital feedback mechanism with distinct physiological implications. Though long-sought, a shared understanding of the channel families remained elusive for two decades as the functional manifestations and the structural underpinnings of this modulation often appeared to diverge.

Apocalmodulin itself promotes ion channel opening and Ca(2+) regulation.

The Ca(2+)-free form of calmodulin (apoCaM) often appears inert, modulating target molecules only upon conversion to its Ca(2+)-bound form. This schema has appeared to govern voltage-gated Ca(2+) channels, where apoCaM has been considered a dormant Ca(2+) sensor, associated with channels but awaiting the binding of Ca(2+) ions before inhibiting channel opening to provide vital feedback inhibition. Using single-molecule measurements of channels and chemical dimerization to elevate apoCaM, we find that apoCaM binding on its own markedly upregulates opening, rivaling the strongest forms of modulation.

Calmodulin mutations associated with long QT syndrome prevent inactivation of cardiac L-type Ca(2+) currents and promote proarrhythmic behavior in ventricular myocytes.

Recent work has identified missense mutations in calmodulin (CaM) that are associated with severe early-onset long-QT syndrome (LQTS), leading to the proposition that altered CaM function may contribute to the molecular etiology of this subset of LQTS. To date, however, no experimental evidence has established these mutations as directly causative of LQTS substrates, nor have the molecular targets of CaM mutants been identified. Here, therefore, we test whether expression of CaM mutants in adult guinea-pig ventricular myocytes (aGPVM) induces action-potential prolongation, and whether affiliated alterations in the Ca(2+) regulation of L-type Ca(2+) channels (LTCC) might contribute to such prolongation.