RESUMEN

Fast and regulated potassium efflux by Slo1 channels is crucial in many tissues in animals including neurons, the kidney and smooth muscle. During the last decade, structures have revealed many details about the gating mechanism and regulation of these large and complex molecular machines. This review summarizes these findings and the current knowledge about the intricate regulation of these important channels. Slo1 integrates sensing of the membrane potential via a voltage-sensor domain that undergoes subtle but significant structural rearrangements with a calcium-induced expansion of parts of the intracellular gating ring. Together, these two signals synergistically lead to changes in the conformation and chemical nature of the pore domain, allowing potassium ions to be translocated. In many native tissues, Slo1 channels are assembled with at least three classes of auxiliary subunits that change the gating kinetics or allow the channel to open also in absence of one of the two signals. Finally, Slo1 is inhibited, activated or deregulated by natural toxins and synthetic compounds, underlining the importance of the channel for the organism and as a potential target for drugs and other molecules.

RESUMEN

Voltage-gated potassium (Kv) channels and hyperpolarization-activated cyclic nucleotide-gated (HCN) channels share similar structures but have opposite gating polarity. Kv channels have a strong coupling (>109) between the voltage sensor (S4) and the activation gate: when S4s are activated, the gate is open to >80% but, when S4s are deactivated, the gate is open <10-9 of the time. Using noise analysis, we show that the coupling between S4 and the gate is <200 in HCN channels. In addition, using voltage clamp fluorometry, locking the gate open in a Kv channel drastically altered the energetics of S4 movement. In contrast, locking the gate open or decreasing the coupling between S4 and the gate in HCN channels had only minor effects on the energetics of S4 movement, consistent with a weak coupling between S4 and the gate. We propose that this loose coupling is a prerequisite for the reversed voltage gating in HCN channels.

Asunto(s)

Canales Regulados por Nucleótidos Cíclicos Activados por Hiperpolarización , Activación del Canal Iónico , Canales Regulados por Nucleótidos Cíclicos Activados por Hiperpolarización/metabolismo , Canales Regulados por Nucleótidos Cíclicos Activados por Hiperpolarización/genética , Animales , Técnicas de Placa-Clamp , Humanos

RESUMEN

Mammalian Ca2+-dependent Slo K+ channels can stably associate with auxiliary γ subunits which fundamentally alter their behavior. By a so far unknown mechanism, the four γ subunits reduce the need for voltage-dependent activation and, thereby, allow Slo to open independently of an action potential. Here, using cryo-EM, we reveal how the transmembrane helix of γ1/LRRC26 binds and presumably stabilizes the activated voltage-sensor domain of Slo1. The activation is further enhanced by an intracellular polybasic stretch which locally changes the charge gradient across the membrane. Our data provide a possible explanation for Slo1 regulation by the four γ subunits and also their different activation efficiencies. This suggests a novel activation mechanism of voltage-gated ion channels by auxiliary subunits.

Asunto(s)

Microscopía por Crioelectrón , Subunidades alfa de los Canales de Potasio de Gran Conductancia Activados por Calcio , Subunidades de Proteína , Humanos , Subunidades alfa de los Canales de Potasio de Gran Conductancia Activados por Calcio/metabolismo , Subunidades alfa de los Canales de Potasio de Gran Conductancia Activados por Calcio/química , Subunidades alfa de los Canales de Potasio de Gran Conductancia Activados por Calcio/genética , Subunidades de Proteína/metabolismo , Subunidades de Proteína/química , Animales , Activación del Canal Iónico , Modelos Moleculares , Células HEK293 , Unión Proteica , Dominios Proteicos

RESUMEN

Ion channels are ubiquitous throughout all forms of life. Potassium channels are even found in viruses. Every cell must communicate with its surroundings, so all cells have them, and excitable cells, in particular, especially nerve cells, depend on the behavior of these channels. Every channel must be open at the appropriate time, and only then, so that each channel opens in response to the stimulus that tells that channel to open. One set of channels, including those in nerve cells, responds to voltage. There is a standard model for the gating of these channels that has a section of the protein moving in response to the voltage. However, there is evidence that protons are moving, rather than protein. Water is critical as part of the gating process, although it is hard to see how this works in the standard model. Here, we review the extensive evidence of the importance of the role of water and protons in gating these channels. Our principal example, but by no means the only example, will be the Kv1.2 channel. Evidence comes from the effects of D2O, from mutations in the voltage sensing domain, as well as in the linker between that domain and the gate, and at the gate itself. There is additional evidence from computations, especially quantum calculations. Structural evidence comes from X-ray studies. The hydration of ions is critical in the transfer of ions in constricted spaces, such as the gate region and the pore of a channel; we will see how the structure of the hydrated ion fits with the structure of the channel. In addition, there is macroscopic evidence from osmotic experiments and streaming current measurements. The combined evidence is discussed in the context of a model that emphasizes the role of protons and water in gating these channels.

RESUMEN

Gap junction channels are regulated by gates sensitive to cytosolic acidification and trans-junctional voltage (Vj). We propose that the chemical gate is a calmodulin (CaM) lobe. The fast-Vj gate is made primarily by the connexin's NH2-terminus domain (NT). The chemical gate closes the channel slowly and completely, while the fast-Vj gate closes the channel rapidly but incompletely. The chemical gate closes with increased cytosolic calcium concentration [Ca2+]i and with Vj gradients at Vj's negative side. In contrast, the fast-Vj gate closes at the positive or negative side of Vj depending on the connexin (Cx) type. Cxs with positively charged NT close at Vj's negative side, while those with negatively charged NT close at Vj's positive side. Cytosolic acidification alters in opposite ways the sensitivity of the fast-Vj gate: it increases the Vj sensitivity of negative gaters and decreases that of positive gaters. While the fast-Vj gate closes and opens instantaneously, the chemical gate often shows fluctuations, likely to reflect the shifting of the gate (CaM's N-lobe) in and out of the channel's pore.

Asunto(s)

Conexinas , Canales Iónicos , Calmodulina , Citosol , Uniones Comunicantes

RESUMEN

KCNQ (Kv7) channels are voltage-gated, phosphatidylinositol 4,5-bisphosphate- (PIP2-) modulated potassium channels that play essential roles in regulating the activity of neurons and cardiac myocytes. Hundreds of mutations in KCNQ channels are closely related to various cardiac and neurological disorders, such as long QT syndrome, epilepsy, and deafness, which makes KCNQ channels important drug targets. During the past several years, the application of single-particle cryo-electron microscopy (cryo-EM) technique in the structure determination of KCNQ channels has greatly advanced our understanding of their molecular mechanisms. In this review, we summarize the currently available structures of KCNQ channels, analyze their special voltage gating mechanism, and discuss their activation mechanisms by both the endogenous membrane lipid and the exogenous synthetic ligands. These structural studies of KCNQ channels will guide the development of drugs targeting KCNQ channels.

Asunto(s)

Epilepsia , Síndrome de QT Prolongado , Humanos , Canales de Potasio KCNQ/genética , Canales de Potasio KCNQ/química , Microscopía por Crioelectrón , Corazón , Síndrome de QT Prolongado/genética

RESUMEN

To fire action-potential-like electrical signals, the vacuole membrane requires the two-pore channel TPC1, formerly called SV channel. The TPC1/SV channel functions as a depolarization-stimulated, non-selective cation channel that is inhibited by luminal Ca2+. In our search for species-dependent functional TPC1 channel variants with different luminal Ca2+ sensitivity, we found in total three acidic residues present in Ca2+ sensor sites 2 and 3 of the Ca2+-sensitive AtTPC1 channel from Arabidopsis thaliana that were neutral in its Vicia faba ortholog and also in those of many other Fabaceae. When expressed in the Arabidopsis AtTPC1-loss-of-function background, wild-type VfTPC1 was hypersensitive to vacuole depolarization and only weakly sensitive to blocking luminal Ca2+. When AtTPC1 was mutated for these VfTPC1-homologous polymorphic residues, two neutral substitutions in Ca2+ sensor site 3 alone were already sufficient for the Arabidopsis At-VfTPC1 channel mutant to gain VfTPC1-like voltage and luminal Ca2+ sensitivity that together rendered vacuoles hyperexcitable. Thus, natural TPC1 channel variants exist in plant families which may fine-tune vacuole excitability and adapt it to environmental settings of the particular ecological niche.

Asunto(s)

Arabidopsis , Vicia faba , Vacuolas , Arabidopsis/genética , Potenciales de Acción , Ecosistema

RESUMEN

The intricate voltage regulation presented by lysenin channels reconstituted in artificial lipid membranes leads to a strong hysteresis in conductance, bistability, and memory. Prior investigations on lysenin channels indicate that the hysteresis is modulated by multivalent cations which are also capable of eliciting single-step conformational changes and transitions to stable closed or sub-conducting states. However, the influence on voltage regulation of Cu2+ ions, capable of completely closing the lysenin channels in a two-step process, was not sufficiently addressed. In this respect, we employed electrophysiology approaches to investigate the response of lysenin channels to variable voltage stimuli in the presence of small concentrations of Cu2+ ions. Our experimental results showed that the hysteretic behavior, recorded in response to variable voltage ramps, is accentuated in the presence of Cu2+ ions. Using simultaneous AC/DC stimulation, we were able to determine that Cu2+ prevents the reopening of channels previously closed by depolarizing potentials and the channels remain in the closed state even in the absence of a transmembrane voltage. In addition, we showed that Cu2+ addition reinstates the voltage gating and hysteretic behavior of lysenin channels reconstituted in neutral lipid membranes in which lysenin channels lose their voltage-regulating properties. In the presence of Cu2+ ions, lysenin not only regained the voltage gating but also behaved like a long-term molecular memory controlled by electrical potentials.

Asunto(s)

Electrofisiología Cardíaca , Electricidad , Iones , Membranas Artificiales , Lípidos

RESUMEN

Voltage-gated ion channels, whose first identified function was to generate action potentials, are divided into subfamilies with numerous members. The family of voltage-gated proton channels (HV ) is tiny. To date, all species found to express HV have exclusively one gene that codes for this unique ion channel. Here we report the discovery and characterization of three proton channel genes in the classical model system of neural plasticity, Aplysia californica. The three channels (AcHV 1, AcHV 2, and AcHV 3) are distributed throughout the whole animal. Patch-clamp analysis confirmed proton selectivity of these channels but they all differed markedly in gating. AcHV 1 gating resembled HV in mammalian cells where it is responsible for proton extrusion and charge compensation. AcHV 2 activates more negatively and conducts extensive inward proton current, properties likely to acidify the cytosol. AcHV 3, which differs from AcHV 1 and AcHV 2 in lacking the first arginine in the S4 helix, exhibits proton selective leak currents and weak voltage dependence. We report the expansion of the proton channel family, demonstrating for the first time the expression of three functionally distinct proton channels in a single species.

Asunto(s)

Activación del Canal Iónico , Protones , Animales , Activación del Canal Iónico/fisiología , Canales Iónicos/metabolismo , Arginina , Citosol/metabolismo , Mamíferos/metabolismo

RESUMEN

Although human sperm is morphologically mature in the epididymis, it cannot fertilize eggs before capacitation. Cholesterol efflux from the sperm plasma membrane is a key molecular event essential for cytoplasmic alkalinization and hyperactivation, but the underlying mechanism remains unclear. The human voltage-gated proton (hHv1) channel functions as an acid extruder to regulate intracellular pHs of many cell types, including sperm. Aside from voltage and pH, Hv channels are also regulated by distinct ligands, such as Zn2+ and albumin. In the present work, we identified cholesterol as an inhibitory ligand of the hHv1 channel and further investigated the underlying mechanism using the single-molecule fluorescence resonance energy transfer (smFRET) approach. Our results indicated that cholesterol inhibits the hHv1 channel by stabilizing the voltage-sensing S4 segment at resting conformations, a similar mechanism also utilized by Zn2+. Our results suggested that the S4 segment is the central gating machinery in the hHv1 channel, on which voltage and distinct ligands are converged to regulate channel function. Identification of membrane cholesterol as an inhibitory ligand provides a mechanism by which the hHv1 channel regulates fertilization by linking the cholesterol efflux with cytoplasmic alkalinization, a change that triggers calcium influx through the CatSper channel. These events finally lead to hyperactivation, a remarkable change in the mobility pattern indicating fertilization competence of human sperm.

Asunto(s)

Colesterol , Activación del Canal Iónico , Colesterol/metabolismo , Humanos , Activación del Canal Iónico/fisiología , Canales Iónicos/metabolismo , Ligandos , Masculino , Semen/metabolismo

RESUMEN

Voltage-gated proton (Hv) channels are standalone voltage sensors without separate ion conductive pores. They are gated by both voltage and transmembrane proton gradient (i.e., ∆pH), serving as acid extruders in most cells. Like the canonical voltage sensors, Hv channels are a bundle of four helices (named S1 -S4), with the S4 segment carrying three positively charged Arg residues. Extensive structural and electrophysiological studies on voltage-gated ion channels, in general, agree on an outwards movement of the S4 segment upon activating voltage, but the real-time conformational transitions are still unattainable. With purified human voltage-gated proton (hHv1) channels reconstituted in liposomes, we have examined its conformational dynamics, including the S4 segment at different voltage and pHs using single-molecule fluorescence resonance energy transfer (smFRET). Here, we provide the first glimpse of real-time conformational trajectories of the hHv1 voltage sensor and show that both voltage and pH gradient shift the conformational dynamics of the S4 segment to control channel gating. Our results indicate that the S4 segment transits among three major conformational states and only the transitions between the inward and outward conformations are highly dependent on voltage and pH. Altogether, we propose a kinetic model that explains the mechanisms underlying voltage and pH gating in Hv channels, which may also serve as a general framework for understanding the voltage sensing and gating in other voltage-gated ion channels.

Asunto(s)

Activación del Canal Iónico , Protones , Humanos , Activación del Canal Iónico/fisiología , Canales Iónicos/metabolismo , Cinética , Estructura Secundaria de Proteína

RESUMEN

Two-pore cation channel, TPC1, is ubiquitous in the vacuolar membrane of terrestrial plants and mediates the long distance signaling upon biotic and abiotic stresses. It possesses a wide pore, which transports small mono- and divalent cations. K+ is transported more than 10-fold faster than Ca2+, which binds with a higher affinity within the pore. Key pore residues, responsible for Ca2+ binding, have been recently identified. There is also a substantial progress in the mechanistic and structural understanding of the plant TPC1 gating by membrane voltage and cytosolic and luminal Ca2+. Collectively, these gating factors at resting conditions strongly reduce the potentially lethal Ca2+ leak from the vacuole. Such tight control is impressive, bearing in mind high unitary conductance of the TPC1 and its abundance, with thousands of active channel copies per vacuole. But it remains a mystery how this high threshold is overcome upon signaling, and what type of signal is emitted by TPC1, whether it is Ca2+ or electrical one, or a transduction via protein conformational change, independent on ion conductance. Here we discuss non-exclusive scenarios for the TPC1 integration into Ca2+, ROS and electrical signaling.

RESUMEN

Arabidopsis thaliana two-pore channel AtTPC1 is a voltage-gated, Ca2+-modulated, nonselective cation channel that is localized in the vacuolar membrane and responsible for generating slow vacuolar (SV) current. Under depolarizing membrane potential, cytosolic Ca2+ activates AtTPC1 by binding at the EF-hand domain, whereas luminal Ca2+ inhibits the channel by stabilizing the voltage-sensing domain II (VSDII) in the resting state. Here, we present 2.8 to 3.3 Å cryoelectron microscopy (cryo-EM) structures of AtTPC1 in two conformations, one in closed conformation with unbound EF-hand domain and resting VSDII and the other in a partially open conformation with Ca2+-bound EF-hand domain and activated VSDII. Structural comparison between the two different conformations allows us to elucidate the structural mechanisms of voltage gating, cytosolic Ca2+ activation, and their coupling in AtTPC1. This study also provides structural insight into the general voltage-gating mechanism among voltage-gated ion channels.

Asunto(s)

Proteínas de Arabidopsis/metabolismo , Canales de Calcio/metabolismo , Calcio/metabolismo , Secuencia de Aminoácidos , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Canales de Calcio/genética , Cationes/metabolismo , Microscopía por Crioelectrón/métodos , Citosol/metabolismo , Activación del Canal Iónico , Potenciales de la Membrana/fisiología , Vacuolas/metabolismo

RESUMEN

The voltage-dependent anion channel (VDAC) is the primary regulating pathway of water-soluble metabolites and ions across the mitochondrial outer membrane. When reconstituted into lipid membranes, VDAC responds to sufficiently large transmembrane potentials by transitioning to gated states in which ATP/ADP flux is reduced and calcium flux is increased. Two otherwise unrelated cytosolic proteins, tubulin, and α-synuclein (αSyn), dock with VDAC by a novel mechanism in which the transmembrane potential draws their disordered, polyanionic C-terminal domains into and through the VDAC channel, thus physically blocking the pore. For both tubulin and αSyn, the blocked state is observed at much lower transmembrane potentials than VDAC gated states, such that in the presence of these cytosolic docking proteins, VDAC's sensitivity to transmembrane potential is dramatically increased. Remarkably, the features of the VDAC gated states relevant for bioenergetics-reduced metabolite flux and increased calcium flux-are preserved in the blocked state induced by either docking protein. The ability of tubulin and αSyn to modulate mitochondrial potential and ATP production in vivo is now supported by many studies. The common physical origin of the interactions of both tubulin and αSyn with VDAC leads to a general model of a VDAC inhibitor, facilitates predictions of the effect of post-translational modifications of known inhibitors, and points the way toward the development of novel therapeutics targeting VDAC.

Asunto(s)

Aniones/metabolismo , Respiración de la Célula/fisiología , Proteínas Intrínsecamente Desordenadas/fisiología , Membranas Mitocondriales/efectos de los fármacos , Tubulina (Proteína)/fisiología , Canales Aniónicos Dependientes del Voltaje/antagonistas & inhibidores , alfa-Sinucleína/fisiología , Secuencia de Aminoácidos , Animales , Calcio/metabolismo , Respiración de la Célula/efectos de los fármacos , Fluoresceínas/química , Humanos , Proteínas Intrínsecamente Desordenadas/química , Activación del Canal Iónico/efectos de los fármacos , Activación del Canal Iónico/fisiología , Cinética , Membranas Mitocondriales/metabolismo , Modelos Moleculares , Concentración Osmolar , Cloruro de Potasio/farmacología , Conformación Proteica , Mapeo de Interacción de Proteínas , Procesamiento Proteico-Postraduccional , Transporte de Proteínas , Alineación de Secuencia , Ácidos Sulfónicos/química , Tubulina (Proteína)/química , Canales Aniónicos Dependientes del Voltaje/química , Canales Aniónicos Dependientes del Voltaje/fisiología , alfa-Sinucleína/química

RESUMEN

The rapidly activating delayed rectifier K+ current generated by the cardiac hERG potassium channel encoded by KCNH2 is the most important reserve current for cardiac repolarization. The unique inward rectification characteristics of the hERG channel depend on the gating regulation, which involves crucial structural domains and key single amino acid residues in the full-length hERG channel. Identifying critical molecules involved in the regulation of gating kinetics for the hERG channel requires high-resolution structures and molecular dynamics simulation models. Based on the latest progress in hERG structure and molecular dynamics simulation research, summarizing the molecules involved in the changes in the channel state helps to elucidate the unique gating characteristics of the channel and the reason for its high affinity to cardiotoxic drugs. In this review, we aim to summarize the significant advances in understanding the voltage gating regulation of the hERG channel based on its structure obtained from cryo-electron microscopy and computer simulations, which reveal the critical roles of several specific structural domains and amino acid residues.

RESUMEN

Voltage-dependent anion channel (VDAC), the most abundant mitochondrial outer membrane protein, is important for a variety of mitochondrial functions including metabolite exchange, calcium transport, and apoptosis. While VDAC's role in shuttling metabolites between the cytosol and mitochondria is well established, there is a growing interest in understanding the mechanisms of its regulation of mitochondrial calcium transport. Here we review the current literature on VDAC's role in calcium signaling, its biophysical properties, physiological function, and pathology focusing on its importance in cardiac diseases. We discuss the specific biophysical properties of the three VDAC isoforms in mammalian cells-VDAC 1, 2, and 3-in relationship to calcium transport and their distinct roles in cell physiology and disease. Highlighting the emerging evidence that cytosolic proteins interact with VDAC and regulate its calcium permeability, we advocate for continued investigation into the VDAC interactome at the contact sites between mitochondria and organelles and its role in mitochondrial calcium transport.

Asunto(s)

Fenómenos Biofísicos , Señalización del Calcio , Enfermedad , Mitocondrias/metabolismo , Canales Aniónicos Dependientes del Voltaje/metabolismo , Animales , Canales de Calcio/metabolismo , Humanos

RESUMEN

While the capsaicin receptor transient receptor potential vanilloid 1 (TRPV1) channel is a polymodal nociceptor for heat, capsaicin, and protons, the channel's responses to each of these stimuli are profoundly regulated by membrane potential, damping or even prohibiting its response at negative voltages and amplifying its response at positive voltages. Therefore, voltage sensitivity of TRPV1 is anticipated to play an important role in shaping pain responses. How voltage regulates TRPV1 activation remains unknown. Here, it is shown that voltage sensitivity does not originate from the S4 segment like classic voltage-gated ion channels; instead, outer pore acidic residues directly partake in voltage-sensitive activation, with their negative charges collectively constituting the observed gating charges. Outer pore gating-charge movement is titratable by extracellular pH and is allosterically coupled to channel activation, likely by influencing the upper gate in the ion selectivity filter. Elucidating this unorthodox voltage-gating process provides a mechanistic foundation for understanding TRPV1 polymodal gating and opens the door to novel approaches regulating channel activity for pain management.

RESUMEN

Transmembrane ß-barrels of eukaryotic outer mitochondrial membranes (OMMs) are major channels of communication between the cytosol and mitochondria and are indispensable for cellular homeostasis. A structurally intriguing exception to all known transmembrane ß-barrels is the unique odd-stranded, i.e. 19-stranded, structures found solely in the OMM. The molecular origins of this 19-stranded structure and its associated functional significance are unclear. In humans, the most abundant OMM transporter is the voltage-dependent anion channel. Here, using the human voltage-dependent anion channel as our template scaffold, we designed and engineered odd- and even-stranded structures of smaller (V216, V217, V218) and larger (V220, V221) barrel diameters. Determination of the structure, dynamics, and energetics of these engineered structures in bilayer membranes reveals that the 19-stranded barrel surprisingly holds modest to low stability in a lipid-dependent manner. However, we demonstrate that this structurally metastable protein possesses superior voltage-gated channel regulation, efficient mitochondrial targeting, and in vivo cell survival, with lipid-modulated stability, all of which supersede the occurrence of a metastable 19-stranded scaffold. We propose that the unique structural adaptation of these transmembrane transporters exclusively in mitochondria bears strong evolutionary basis and is functionally significant for homeostasis.

Asunto(s)

Membrana Dobles de Lípidos/metabolismo , Canales Aniónicos Dependientes del Voltaje/química , Canales Aniónicos Dependientes del Voltaje/metabolismo , Animales , Evolución Molecular , Humanos , Membrana Dobles de Lípidos/química , Mitocondrias/química , Mitocondrias/genética , Mitocondrias/metabolismo , Modelos Moleculares , Mutación , Porinas/química , Porinas/genética , Porinas/metabolismo , Conformación Proteica en Lámina beta , Ingeniería de Proteínas , Estabilidad Proteica , Saccharomyces cerevisiae/química , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/química , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Termodinámica , Canal Aniónico 2 Dependiente del Voltaje/química , Canal Aniónico 2 Dependiente del Voltaje/genética , Canal Aniónico 2 Dependiente del Voltaje/metabolismo , Canales Aniónicos Dependientes del Voltaje/genética

RESUMEN

Voltage-gated ion channels feature voltage sensor domains (VSDs) that exist in three distinct conformations during activation: resting, intermediate, and activated. Experimental determination of the structure of a potassium channel VSD in the intermediate state has previously proven elusive. Here, we report and validate the experimental three-dimensional structure of the human KCNQ1 voltage-gated potassium channel VSD in the intermediate state. We also used mutagenesis and electrophysiology in Xenopus laevisoocytes to functionally map the determinants of S4 helix motion during voltage-dependent transition from the intermediate to the activated state. Finally, the physiological relevance of the intermediate state KCNQ1 conductance is demonstrated using voltage-clamp fluorometry. This work illuminates the structure of the VSD intermediate state and demonstrates that intermediate state conductivity contributes to the unusual versatility of KCNQ1, which can function either as the slow delayed rectifier current (IKs) of the cardiac action potential or as a constitutively active epithelial leak current.

Asunto(s)

Canal de Potasio KCNQ1/fisiología , Animales , Electrofisiología , Fluorometría , Humanos , Canal de Potasio KCNQ1/química , Canal de Potasio KCNQ1/metabolismo , Espectroscopía de Resonancia Magnética , Oocitos , Técnicas de Placa-Clamp , Estructura Terciaria de Proteína , Xenopus laevis

RESUMEN

Evidence that neighboring cells uncouple from each other as one dies surfaced in the late 19th century, but it took almost a century for scientists to start understanding the uncoupling mechanism (chemical gating). The role of cytosolic free calcium (Ca2+i) in cell-cell channel gating was first reported in the mid-sixties. In these studies, only micromolar [Ca2+]i were believed to affect gating-concentrations reachable only in cell death, which would discard Ca2+i as a fine modulator of cell coupling. More recently, however, numerous researchers, including us, have reported the effectiveness of nanomolar [Ca2+]i. Since connexins do not have high-affinity calcium sites, the effectiveness of nanomolar [Ca2+]i suggests the role of Ca-modulated proteins, with calmodulin (CaM) being most obvious. Indeed, in 1981 we first reported that a CaM-inhibitor prevents chemical gating. Since then, the CaM role in gating has been confirmed by studies that tested it with a variety of approaches such as treatments with CaM-inhibitors, inhibition of CaM expression, expression of CaM mutants, immunofluorescent co-localization of CaM and gap junctions, and binding of CaM to peptides mimicking connexin domains identified as CaM targets. Our gating model envisions Ca2+-CaM to directly gate the channels by acting as a plug ("Cork" gating model), and probably also by affecting connexin conformation.

Asunto(s)

Calcio/metabolismo , Calmodulina/metabolismo , Uniones Comunicantes/metabolismo , Animales , Calmodulina/genética , Citosol/metabolismo , Humanos , Activación del Canal Iónico , Transducción de Señal