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1.
Cells ; 13(17)2024 Aug 23.
Artigo em Inglês | MEDLINE | ID: mdl-39272981

RESUMO

The KCNE2 protein is encoded by the kcne2 gene and is a member of the KCNE protein family, also known as the MinK-related protein 1 (MiRP1). It is mostly present in the epicardium of the heart and gastric mucosa, and it is also found in the thyroid, pancreatic islets, liver and lung, among other locations, to a lesser extent. It is involved in numerous physiological processes because of its ubiquitous expression and partnering promiscuity, including the modulation of voltage-dependent potassium and calcium channels involved in cardiac action potential repolarization, and regulation of secretory processes in multiple epithelia, such as gastric acid secretion, thyroid hormone synthesis, generation and secretion of cerebrospinal fluid. Mutations in the KCNE2 gene or aberrant expression of the protein may play a critical role in cardiovascular, neurological, metabolic and multisystem disorders. This article provides an overview of the advancements made in understanding the physiological functions in organismal homeostasis and the pathophysiological consequences of KCNE2 in multisystem diseases.


Assuntos
Arritmias Cardíacas , Canais de Potássio de Abertura Dependente da Tensão da Membrana , Humanos , Canais de Potássio de Abertura Dependente da Tensão da Membrana/metabolismo , Canais de Potássio de Abertura Dependente da Tensão da Membrana/genética , Arritmias Cardíacas/genética , Arritmias Cardíacas/metabolismo , Animais
2.
Circ Res ; 135(7): 722-738, 2024 Sep 13.
Artigo em Inglês | MEDLINE | ID: mdl-39166328

RESUMO

BACKGROUND: The KCNQ1+KCNE1 (IKs) potassium channel plays a crucial role in cardiac adaptation to stress, in which ß-adrenergic stimulation phosphorylates the IKs channel through the cyclic adenosine monophosphate (cAMP)/PKA (protein kinase A) pathway. Phosphorylation increases the channel current and accelerates repolarization to adapt to an increased heart rate. Variants in KCNQ1 can cause long-QT syndrome type 1 (LQT1), and those with defective cAMP effects predispose patients to the highest risk of cardiac arrest and sudden death. However, the molecular connection between IKs channel phosphorylation and channel function, as well as why high-risk LQT1 mutations lose cAMP sensitivity, remain unclear. METHODS: Regular patch clamp and voltage clamp fluorometry techniques were utilized to record pore opening and voltage sensor movement of wild-type and mutant KCNQ1/IKs channels. The clinical phenotypic penetrance of each LQT1 mutation was analyzed as a metric for assessing their clinical risk. The patient-specific-induced pluripotent stem-cell model was used to test mechanistic findings in physiological conditions. RESULTS: By systematically elucidating mechanisms of a series of LQT1 variants that lack cAMP sensitivity, we identified molecular determinants of IKs channel regulation by phosphorylation. These key residues are distributed across the N-terminus of KCNQ1 extending to the central pore region of IKs. We refer to this pattern as the IKs channel PKA phosphorylation axis. Next, by examining LQT1 variants from clinical databases containing 10 579 LQT1 carriers, we found that the distribution of the most high-penetrance LQT1 variants extends across the IKs channel PKA phosphorylation axis, demonstrating its clinical relevance. Furthermore, we found that a small molecule, ML277, which binds at the center of the phosphorylation axis, rescues the defective cAMP effects of multiple high-risk LQT1 variants. This finding was then tested in high-risk patient-specific induced pluripotent stem cell-derived cardiomyocytes, where ML277 remarkably alleviates the beating abnormalities. CONCLUSIONS: Our findings not only elucidate the molecular mechanism of PKA-dependent IKs channel phosphorylation but also provide an effective antiarrhythmic strategy for patients with high-risk LQT1 variants.


Assuntos
Proteínas Quinases Dependentes de AMP Cíclico , Células-Tronco Pluripotentes Induzidas , Canal de Potássio KCNQ1 , Humanos , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Fosforilação , Canal de Potássio KCNQ1/genética , Canal de Potássio KCNQ1/metabolismo , Células-Tronco Pluripotentes Induzidas/metabolismo , Síndrome de Romano-Ward/genética , Síndrome de Romano-Ward/metabolismo , AMP Cíclico/metabolismo , Miócitos Cardíacos/metabolismo , Mutação , Síndrome do QT Longo/genética , Síndrome do QT Longo/metabolismo , Células HEK293 , Canais de Potássio de Abertura Dependente da Tensão da Membrana
3.
Virol J ; 21(1): 181, 2024 Aug 08.
Artigo em Inglês | MEDLINE | ID: mdl-39118175

RESUMO

The orf virus (ORFV) poses a serious threat to the health of domestic small ruminants (i.e., sheep and goats) and humans on a global scale, causing around $150 million in annual losses to livestock industry. However, the host factors involved in ORFV infection and replication are still elusive. In this study, we compared the RNA-seq profiles of ORFV-infected or non-infected sheep testicular interstitial cells (STICs) and identified a novel host gene, potassium voltage-gated channel subfamily E member 4 (KCNE4), as a key host factor involved in the ORFV infection. Both RNA-seq data and RT-qPCR assay revealed a significant increase in the expression of KCNE4 in the infected STICs from 9 to 48 h post infection (hpi). On the other hand, the RT-qPCR assay detected a decrease in ORFV copy number in both the STICs transfected by KCNE4 siRNA and the KCNE4 knockout (KO) HeLa cells after the ORFV infection, together with a reduced fluorescence ratio of ORFV-GFP in the KO HeLa cells at 24 hpi, indicating KCNE4 to be critical for the ORFV infection. Furthermore, the attachment and internalization assays showed decreased ORFV attachment, internalization, replication, and release by the KO HeLa cells, demonstrating a potential inhibition of ORFV entry into the cells by KCNE4. Pretreatment with the KCNE4 inhibitors such as quinidine and fluoxetine significantly repressed the ORFV infection. All our findings reveal KCNE4 as a novel host regulator of the ORFV entry and replication, shedding new insight into the interactive mechanism of ORFV infection. The study also highlights the K+ channels as possible druggable targets to impede viral infection and disease.


Assuntos
Vírus do Orf , Canais de Potássio de Abertura Dependente da Tensão da Membrana , Internalização do Vírus , Animais , Humanos , Ovinos , Canais de Potássio de Abertura Dependente da Tensão da Membrana/genética , Canais de Potássio de Abertura Dependente da Tensão da Membrana/metabolismo , Células HeLa , Vírus do Orf/genética , Vírus do Orf/fisiologia , Replicação Viral , Interações Hospedeiro-Patógeno , Masculino , Ectima Contagioso/virologia
4.
Mar Drugs ; 22(8)2024 Jul 29.
Artigo em Inglês | MEDLINE | ID: mdl-39195466

RESUMO

Bioactive compounds are abundant in animals originating from marine ecosystems. Ion channels, which include sodium, potassium, calcium, and chloride, together with their numerous variants and subtypes, are the primary molecular targets of the latter. Based on their cellular targets, these venom compounds show a range of potencies and selectivity and may have some therapeutic properties. Due to their potential as medications to treat a range of (human) diseases, including pain, autoimmune disorders, and neurological diseases, marine molecules have been the focus of several studies over the last ten years. The aim of this review is on the various facets of marine (or marine-derived) molecules, ranging from structural characterization and discovery to pharmacology, culminating in the development of some "novel" candidate chemotherapeutic drugs that target potassium channels.


Assuntos
Toxinas Marinhas , Canais de Potássio de Abertura Dependente da Tensão da Membrana , Animais , Humanos , Toxinas Marinhas/farmacologia , Canais de Potássio de Abertura Dependente da Tensão da Membrana/antagonistas & inibidores , Canais de Potássio de Abertura Dependente da Tensão da Membrana/metabolismo , Canais de Potássio de Abertura Dependente da Tensão da Membrana/efeitos dos fármacos , Organismos Aquáticos , Descoberta de Drogas
5.
Biochim Biophys Acta Biomembr ; 1866(7): 184377, 2024 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-39103068

RESUMO

KCNQ1, also known as Kv7.1, is a voltage gated potassium channel that associates with the KCNE protein family. Mutations in this protein has been found to cause a variety of diseases including Long QT syndrome, a type of cardiac arrhythmia where the QT interval observed on an electrocardiogram is longer than normal. This condition is often aggravated during strenuous exercise and can cause fainting spells or sudden death. KCNE1 is an ancillary protein that interacts with KCNQ1 in the membrane at varying molar ratios. This interaction allows for the flow of potassium ions to be modulated to facilitate repolarization of the heart. The interaction between these two proteins has been studied previously with cysteine crosslinking and electrophysiology. In this study, electron paramagnetic resonance (EPR) spectroscopy line shape analysis in tandem with site directed spin labeling (SDSL) was used to observe changes in side chain dynamics as KCNE1 interacts with KCNQ1. KCNE1 was labeled at different sites that were found to interact with KCNQ1 based on previous literature, along with sites outside of that range as a control. Once labeled KCNE1 was incorporated into vesicles, KCNQ1 (helices S1-S6) was titrated into the vesicles. The line shape differences observed upon addition of KCNQ1 are indicative of an interaction between the two proteins. This method provides a first look at the interactions between KCNE1 and KCNQ1 from a dynamics perspective using the full transmembrane portion of KCNQ1.


Assuntos
Canal de Potássio KCNQ1 , Canais de Potássio de Abertura Dependente da Tensão da Membrana , Canais de Potássio de Abertura Dependente da Tensão da Membrana/metabolismo , Canais de Potássio de Abertura Dependente da Tensão da Membrana/genética , Canais de Potássio de Abertura Dependente da Tensão da Membrana/química , Canal de Potássio KCNQ1/metabolismo , Canal de Potássio KCNQ1/genética , Canal de Potássio KCNQ1/química , Espectroscopia de Ressonância de Spin Eletrônica/métodos , Ligação Proteica , Humanos , Animais , Síndrome do QT Longo/metabolismo , Síndrome do QT Longo/genética
6.
Vascul Pharmacol ; 156: 107418, 2024 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-39159736

RESUMO

Substituted catechols include both natural and synthetic compounds found in the environment and foods. Some of them are flavonoid metabolites formed by the gut microbiota which are absorbed afterwards. Our previous findings showed that one of these metabolites, 4-methylcatechol, exerts potent vasorelaxant effects in rats. In the current study, we aimed at testing of its 22 structural congeners in order to find the most potent structure and to investigate the mechanism of action. 3-methoxycatechol (3-MOC), 4-ethylcatechol, 3,5-dichlorocatechol, 4-tert-butylcatechol, 4,5-dichlorocatechol, 3-fluorocatechol, 3-isopropylcatechol, 3-methylcatechol and the parent 4-methylcatechol exhibited high vasodilatory activities on isolated rat aortic rings with EC50s ranging from ∼10 to 24 µM. Some significant sex-differences were found. The most potent compound, 3-MOC, relaxed also resistant mesenteric artery but not porcine coronary artery, and decreased arterial blood pressure in both male and female spontaneously hypertensive rats in vivo without affecting heart rate. It potentiated the vasodilation mediated by cAMP and cGMP, but did not impact L-type Ca2+-channels. By using two inhibitors, activation of voltage-gated potassium channels (KV) was found to be involved in the mechanism of action. This was corroborated by docking analysis of 3-MOC with the KV7.4 channel. None of the most active catechols decreased the viability of the A-10 rat embryonic thoracic aorta smooth muscle cell line. Our findings showed that various catechols can relax vascular smooth muscles and hence could provide templates for developing new antihypertensive vasodilator agents without affecting coronary circulation.


Assuntos
Catecóis , Artérias Mesentéricas , Simulação de Acoplamento Molecular , Ratos Endogâmicos SHR , Vasodilatação , Vasodilatadores , Animais , Vasodilatação/efeitos dos fármacos , Masculino , Catecóis/farmacologia , Catecóis/química , Vasodilatadores/farmacologia , Vasodilatadores/química , Feminino , Artérias Mesentéricas/efeitos dos fármacos , Artérias Mesentéricas/metabolismo , Canais de Potássio de Abertura Dependente da Tensão da Membrana/metabolismo , Canais de Potássio de Abertura Dependente da Tensão da Membrana/antagonistas & inibidores , Canais de Potássio de Abertura Dependente da Tensão da Membrana/efeitos dos fármacos , Músculo Liso Vascular/efeitos dos fármacos , Músculo Liso Vascular/metabolismo , Suínos , Relação Dose-Resposta a Droga , Aorta Torácica/efeitos dos fármacos , Aorta Torácica/metabolismo , Hipertensão/tratamento farmacológico , Hipertensão/fisiopatologia , Hipertensão/metabolismo , Pressão Arterial/efeitos dos fármacos , Vasos Coronários/efeitos dos fármacos , Vasos Coronários/metabolismo , Ratos , Fatores Sexuais , Miócitos de Músculo Liso/efeitos dos fármacos , Miócitos de Músculo Liso/metabolismo , Modelos Animais de Doenças , Relação Estrutura-Atividade , GMP Cíclico/metabolismo
7.
Artigo em Inglês | MEDLINE | ID: mdl-39154976

RESUMO

The gram-negative toxin lipopolysaccharides (LPS) are known to trigger inflammatory cytokines in mammals, which can result in pathological responses. Upon treatment of bacterial sepsis with antibiotics, the lysing bacteria can present a surge in LPS, inducing a cytokine storm. However, LPS can also have direct cellular effects, including transient rapid hyperpolarizing of the membrane potential, blocking glutamate receptors and even promoting release of glutamate. The detailed mechanism of action for these immediate responses is still unresolved. In addressing the membrane hyperpolarization, voltage gated K+ channel blockers 4-aminopyridine (4-AP, 3 mM), quinidine hydrochloride monohydrate (0.1 mM) and tetraethylammonium (TEA, 20 mM) were examined along with RNAi knockdowns of potential calcium activated K+ channels. The immediate responses of LPS were not blocked. Even in the presence of glutamate, the membrane still hyperpolarizes with LPS. When the driving gradient for the ionotropic glutamate receptors is enhanced during hyperpolarization, spontaneous quantal responses are dampened in amplitude. Thus, glutamate receptors are blocked, and the mechanism of hyperpolarization remains unresolved. The larval Drosophila glutamatergic neuromuscular junction is used as a model synaptic preparation to address the direct rapid actions by LPS.


Assuntos
Lipopolissacarídeos , Potenciais da Membrana , Animais , Lipopolissacarídeos/farmacologia , Potenciais da Membrana/efeitos dos fármacos , Canais de Potássio Cálcio-Ativados/metabolismo , Junção Neuromuscular/efeitos dos fármacos , Junção Neuromuscular/metabolismo , Canais de Potássio de Abertura Dependente da Tensão da Membrana/metabolismo , Drosophila melanogaster , Ácido Glutâmico/metabolismo , Ácido Glutâmico/farmacologia , Membrana Celular/metabolismo , Membrana Celular/efeitos dos fármacos , Bloqueadores dos Canais de Potássio/farmacologia , Larva/efeitos dos fármacos , Larva/metabolismo
8.
Int J Mol Sci ; 25(16)2024 Aug 16.
Artigo em Inglês | MEDLINE | ID: mdl-39201613

RESUMO

This study investigates novel short-lived long noncoding RNAs (lncRNAs) in mice with altered expression in metabolic dysfunction-associated steatotic liver (MASH) and liver fibrosis. LncRNAs share similarities with mRNAs in their transcription by RNA polymerase II, possession of a 5' cap structure, and presence of a polyA tail. We identified two lncRNAs, Kcnq1ot1 and Rmst, significantly decreased in both conditions. These lncRNAs showed dramatic expression changes in MASH livers induced by Western diets and CCl4, and in fibrotic livers induced by CCl4 alone. The decrease was more pronounced in liver fibrosis, suggesting their potential as biomarkers for disease progression. Our findings are consistent across different fibrosis models, indicating a crucial role for these lncRNAs in MASH and liver fibrosis in mice. With MASH becoming a global health issue and its progression to fibrosis associated with hepatocarcinogenesis and poor prognosis, understanding the underlying mechanisms is critical. This research contributes to elucidating lncRNA functions in murine liver diseases and provides a foundation for developing novel therapeutic strategies targeting lncRNAs in MASH and liver fibrosis, offering new avenues for potential therapeutic interventions.


Assuntos
Biomarcadores , Cirrose Hepática , RNA Longo não Codificante , Animais , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Camundongos , Cirrose Hepática/genética , Cirrose Hepática/metabolismo , Cirrose Hepática/patologia , Masculino , Canais de Potássio de Abertura Dependente da Tensão da Membrana/genética , Canais de Potássio de Abertura Dependente da Tensão da Membrana/metabolismo , Hepatopatias/genética , Hepatopatias/metabolismo , Hepatopatias/patologia , Camundongos Endogâmicos C57BL , Doença Crônica , Modelos Animais de Doenças , Fígado Gorduroso/genética , Fígado Gorduroso/metabolismo , Fígado Gorduroso/patologia
9.
Int J Mol Sci ; 25(16)2024 Aug 17.
Artigo em Inglês | MEDLINE | ID: mdl-39201645

RESUMO

Migraines are a common type of headache affecting around 15% of the population. The signalling pathways leading to migraines have not been fully understood, but neuronal voltage-gated ion channels, such as KCNG4, have been linked to this pathology. KCNG4 (Kv6.4) is a silent member of the superfamily of voltage-gated potassium (Kv) channels, which expresses in heterotetramers with members of the KCNB (Kv2) family. The genetic variant Kv6.4-L360P has previously been linked to migraines, but their mode of action remains unknown. Here, we characterized the molecular characteristics of Kv6.4-L360P when co-expressed with Kv2.1. We found that Kv6.4-L360P almost completely abolishes Kv2 currents, and we propose that this mechanism in the trigeminal system, linked to the initiation of migraine, leads to the pathology.


Assuntos
Transtornos de Enxaqueca , Canais de Potássio de Abertura Dependente da Tensão da Membrana , Canais de Potássio Shab , Animais , Humanos , Variação Genética , Células HEK293 , Transtornos de Enxaqueca/genética , Transtornos de Enxaqueca/metabolismo , Canais de Potássio de Abertura Dependente da Tensão da Membrana/genética , Canais de Potássio de Abertura Dependente da Tensão da Membrana/metabolismo , Canais de Potássio Shab/genética , Canais de Potássio Shab/metabolismo
10.
J Biochem Mol Toxicol ; 38(8): e23770, 2024 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-39016041

RESUMO

This study aimed to investigate the relationship and potential mechanisms of miR-200c-5p in colorectal cancer (CRC) progression. Differentially expressed miRNAs were screened using the TCGA database. Subsequently, univariate analysis was performed to identify CRC survival-related miRNAs. Survival and receiver operator characteristic curves were generated. The target genes of miR-200c-5p and the relevant signaling pathways or biological processes were predicted by the miRNet database and enrichment analyses. The miR-200c-5p expression was detected using quantitative reverse-transcription polymerase chain reaction, Cell Counting Kit-8, Transwell, and cell apoptosis experiments were performed to determine miR-200c-5p's impact on CRC cell viability, invasiveness, and apoptosis. Finally, we constructed a CRC mouse model with inhibited miR-200c-5p to evaluate its impact on tumors. miR-200c-5p was upregulated in CRC, implying a favorable prognosis. Gene set enrichment analysis revealed that miR-200c-5p may participate in signaling pathways such as the TGF-ß signaling pathway, RIG-I-like receptor signaling pathway, renin-angiotensin system, and DNA replication. miR-200c-5p potentially targeted mRNAs, including KCNE4 and CYP1B1, exhibiting a negative correlation with their expression. Furthermore, these mRNAs may participate in biological processes like the regulation of intracellular transport, cAMP-dependent protein kinase regulatory activity, ubiquitin protein ligase binding, MHC class II protein complex binding, and regulation of apoptotic signaling pathway. Lastly, miR-200c-5p overexpression repressed the viability and invasiveness of CRC cells but promoted apoptosis. The tumor size, weight, and volume were significantly increased by inhibiting miR-200c-5p (p < 0.05). miR-200c-5p is upregulated in CRC, serving as a promising biomarker for predicting CRC prognosis.


Assuntos
Neoplasias Colorretais , MicroRNAs , MicroRNAs/genética , MicroRNAs/metabolismo , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Neoplasias Colorretais/metabolismo , Humanos , Animais , Camundongos , Prognóstico , Regulação Neoplásica da Expressão Gênica , Apoptose/genética , Masculino , Linhagem Celular Tumoral , Camundongos Nus , Canais de Potássio de Abertura Dependente da Tensão da Membrana/genética , Canais de Potássio de Abertura Dependente da Tensão da Membrana/metabolismo , Feminino , Camundongos Endogâmicos BALB C
11.
Stem Cell Res ; 80: 103512, 2024 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-39083856

RESUMO

Pathogenic variants in the KCNV2 gene can cause a rare retinal dystrophy that can be inherited recessively, known as cone dystrophy with supernormal rod response (CDSRR). CDSRR leads to specific changes in photoreceptors' electroretinogram response, especially in the rods, poor visual acuity, photophobia, and even maculopathy. The derived iPSC lines from patients with CDSRR may pave the way for apprehension of the pathogenetic mechanism and drug development using in vitro models. PBMCs were established into induced pluripotent stem cells and then characterized by confirming the expression of pluripotency markers, demonstrating the ability to differentiate into the three germ layers, and obtaining normal karyotyping.


Assuntos
Células-Tronco Pluripotentes Induzidas , Mutação , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Canais de Potássio de Abertura Dependente da Tensão da Membrana/genética , Canais de Potássio de Abertura Dependente da Tensão da Membrana/metabolismo , Linhagem Celular , Heterozigoto , Diferenciação Celular , Masculino
12.
Cell Mol Life Sci ; 81(1): 301, 2024 Jul 14.
Artigo em Inglês | MEDLINE | ID: mdl-39003683

RESUMO

Voltage-gated K+ (KV) channels govern K+ ion flux across cell membranes in response to changes in membrane potential. They are formed by the assembly of four subunits, typically from the same family. Electrically silent KV channels (KVS), however, are unable to conduct currents on their own. It has been assumed that these KVS must obligatorily assemble with subunits from the KV2 family into heterotetrameric channels, thereby giving rise to currents distinct from those of homomeric KV2 channels. Herein, we show that KVS subunits indeed also modulate the activity, biophysical properties and surface expression of recombinant KV7 isoforms in a subunit-specific manner. Employing co-immunoprecipitation, and proximity labelling, we unveil the spatial coexistence of KVS and KV7 within a single protein complex. Electrophysiological experiments further indicate functional interaction and probably heterotetramer formation. Finally, single-cell transcriptomic analyses identify native cell types in which this KVS and KV7 interaction may occur. Our findings demonstrate that KV cross-family interaction is much more versatile than previously thought-possibly serving nature to shape potassium conductance to the needs of individual cell types.


Assuntos
Subunidades Proteicas , Humanos , Animais , Subunidades Proteicas/metabolismo , Células HEK293 , Potenciais da Membrana , Isoformas de Proteínas/metabolismo , Isoformas de Proteínas/genética , Canais de Potássio de Abertura Dependente da Tensão da Membrana/metabolismo , Canais de Potássio de Abertura Dependente da Tensão da Membrana/genética , Canal de Potássio KCNQ1/metabolismo , Canal de Potássio KCNQ1/genética
13.
J Gen Physiol ; 156(7)2024 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-38832889

RESUMO

Voltage-gated ion channels are responsible for the electrical excitability of neurons and cardiomyocytes. Thus, they are obvious targets for pharmaceuticals aimed to modulate excitability. Compounds activating voltage-gated potassium (KV) channels are expected to reduce excitability. To search for new KV-channel activators, we performed a high-throughput screen of 10,000 compounds on a specially designed Shaker KV channel. Here, we report on a large family of channel-activating compounds with a carboxyl (COOH) group as the common motif. The most potent COOH activators are lipophilic (4 < LogP <7) and are suggested to bind at the interface between the lipid bilayer and the channel's positively charged voltage sensor. The negatively charged form of the COOH-group compounds is suggested to open the channel by electrostatically pulling the voltage sensor to an activated state. Several of the COOH-group compounds also activate the therapeutically important KV7.2/7.3 channel and can thus potentially be developed into antiseizure drugs. The COOH-group compounds identified in this study are suggested to act via the same site and mechanism of action as previously studied COOH-group compounds, such as polyunsaturated fatty acids and resin acids, but distinct from sites for several other types of potassium channel-activating compounds.


Assuntos
Ativação do Canal Iônico , Animais , Ativação do Canal Iônico/efeitos dos fármacos , Superfamília Shaker de Canais de Potássio/metabolismo , Canal de Potássio KCNQ2/metabolismo , Canal de Potássio KCNQ2/agonistas , Canais de Potássio de Abertura Dependente da Tensão da Membrana/metabolismo , Canais de Potássio de Abertura Dependente da Tensão da Membrana/efeitos dos fármacos , Canal de Potássio KCNQ3/metabolismo , Humanos , Xenopus laevis
14.
Molecules ; 29(12)2024 Jun 20.
Artigo em Inglês | MEDLINE | ID: mdl-38931004

RESUMO

Potassium channels have recently emerged as suitable target for the treatment of epileptic diseases. Among potassium channels, KCNT1 channels are the most widely characterized as responsible for several epileptic and developmental encephalopathies. Nevertheless, the medicinal chemistry of KCNT1 blockers is underdeveloped so far. In the present review, we describe and analyse the papers addressing the issue of KCNT1 blockers' development and identification, also evidencing the pros and the cons of the scientific approaches therein described. After a short introduction describing the epileptic diseases and the structure-function of potassium channels, we provide an extensive overview of the chemotypes described so far as KCNT1 blockers, and the scientific approaches used for their identification.


Assuntos
Química Farmacêutica , Epilepsia , Bloqueadores dos Canais de Potássio , Humanos , Bloqueadores dos Canais de Potássio/química , Bloqueadores dos Canais de Potássio/uso terapêutico , Bloqueadores dos Canais de Potássio/farmacologia , Química Farmacêutica/métodos , Epilepsia/tratamento farmacológico , Epilepsia/metabolismo , Relação Estrutura-Atividade , Animais , Anticonvulsivantes/química , Anticonvulsivantes/farmacologia , Anticonvulsivantes/uso terapêutico , Proteínas do Tecido Nervoso/antagonistas & inibidores , Proteínas do Tecido Nervoso/química , Proteínas do Tecido Nervoso/metabolismo , Canais de Potássio de Domínios Poros em Tandem/antagonistas & inibidores , Canais de Potássio de Domínios Poros em Tandem/metabolismo , Canais de Potássio de Domínios Poros em Tandem/química , Canais de Potássio de Abertura Dependente da Tensão da Membrana/antagonistas & inibidores , Canais de Potássio de Abertura Dependente da Tensão da Membrana/metabolismo , Canais de Potássio Ativados por Sódio
15.
Biochem Pharmacol ; 226: 116368, 2024 08.
Artigo em Inglês | MEDLINE | ID: mdl-38880360

RESUMO

The voltage-dependent potassium channel Kv1.3 is a promising therapeutic target for the treatment of autoimmune and chronic inflammatory disorders. Kv1.3 blockers are effective in treating multiple sclerosis (fampridine) and psoriasis (dalazatide). However, most Kv1.3 pharmacological antagonists are not specific enough, triggering potential side effects and limiting their therapeutic use. Functional Kv are oligomeric complexes in which the presence of ancillary subunits shapes their function and pharmacology. In leukocytes, Kv1.3 associates with KCNE4, which reduces the surface abundance and enhances the inactivation of the channel. This mechanism exerts profound consequences on Kv1.3-related physiological responses. Because KCNE peptides alter the pharmacology of Kv channels, we studied the effects of KCNE4 on Kv1.3 pharmacology to gain insights into pharmacological approaches. To that end, we used margatoxin, which binds the channel pore from the extracellular space, and Psora-4, which blocks the channel from the intracellular side. While KCNE4 apparently did not alter the affinity of either margatoxin or Psora-4, it slowed the inhibition kinetics of the latter in a stoichiometry-dependent manner. The results suggested changes in the Kv1.3 architecture in the presence of KCNE4. The data indicated that while the outer part of the channel mouth remains unaffected, KCNE4 disturbs the intracellular architecture of the complex. Various leukocyte types expressing different Kv1.3/KCNE4 configurations participate in the immune response. Our data provide evidence that the presence of these variable architectures, which affect both the structure of the complex and their pharmacology, should be considered when developing putative therapeutic approaches.


Assuntos
Canal de Potássio Kv1.3 , Canais de Potássio de Abertura Dependente da Tensão da Membrana , Canal de Potássio Kv1.3/antagonistas & inibidores , Canal de Potássio Kv1.3/metabolismo , Canal de Potássio Kv1.3/genética , Humanos , Canais de Potássio de Abertura Dependente da Tensão da Membrana/metabolismo , Canais de Potássio de Abertura Dependente da Tensão da Membrana/antagonistas & inibidores , Animais , Bloqueadores dos Canais de Potássio/farmacologia , Cricetulus , Células CHO , Células HEK293 , Ficusina , Venenos de Escorpião
16.
Schizophr Res ; 270: 260-272, 2024 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-38944972

RESUMO

BACKGROUND: It is known that the immune system is dysregulated in schizophrenia, having a state similar to chronic neuroinflammation. The origin of this process is unknown, but it is known that T and B lymphocytes, which are components of the adaptive immune system, play an important role in the pathogenic mechanisms of schizophrenia. METHODS: We analysed the membrane of PBMCs from patients diagnosed with schizophrenia through proteomic analysis (n = 5 schizophrenia and n = 5 control). We found the presence of the Kv1.3 voltage-gated potassium channel and its auxiliary subunit ß1 (KCNAB1) and ß2 (KCNAB2). From a sample of 90 participants, we carried out a study on lymphocytes with whole-cell patch-clamp experiments (n = 7 schizophrenia and n = 5 control), western blot (n = 40 schizophrenia and n = 40 control) and confocal microscopy to evaluate the presence and function of different channels. Kv in both cells. RESULTS: We demonstrated the overexpression of Kv1.1, Kv1.2, Kv1.3, Kv1.6, Kv4.2, Kv4.3 and Kv7.2 channels in PBMCs from patients with schizophrenia. This study represents a groundbreaking exploration, as it involves an electrophysiological analysis performed on T and B lymphocytes from patients diagnosed of schizophrenia compared to healthy participants. We observed that B lymphocytes exhibited an increase in output current along with greater peak current amplitude and voltage conductance curves among patients with schizophrenia compared with healthy controls. CONCLUSIONS: This study showed the importance of the B lymphocyte in schizophrenia. We know that the immune system is altered in schizophrenia, but the physiological mechanisms of this system are not very well known. We suggest that the B lymphocyte may be relevant in the pathophysiology of schizophrenia and that it should be investigated in more depth, opening a new field of knowledge and possibilities for new treatments combining antipsychotics and immunomodulators. The limitation is that all participants received antipsychotic medication, which may have influenced the differences observed between patients and controls. This implies that more studies need to be done where the groups can be separated according to the antipsychotic drug.


Assuntos
Canais de Potássio de Abertura Dependente da Tensão da Membrana , Esquizofrenia , Humanos , Esquizofrenia/metabolismo , Esquizofrenia/imunologia , Esquizofrenia/fisiopatologia , Masculino , Feminino , Adulto , Pessoa de Meia-Idade , Canais de Potássio de Abertura Dependente da Tensão da Membrana/metabolismo , Técnicas de Patch-Clamp , Linfócitos/metabolismo , Potenciais da Membrana/fisiologia , Potenciais da Membrana/efeitos dos fármacos , Proteômica
17.
Gene ; 923: 148590, 2024 Sep 25.
Artigo em Inglês | MEDLINE | ID: mdl-38772516

RESUMO

Long noncoding RNAs (lncRNAs) are implicated in a number of regulatory functions in eukaryotic genomes. In humans, KCNQ1OT1 is a 91 kb imprinted lncRNA that inhibits multiple surrounding genes in cis. Among them, CDKN1C is closely related to KCNQ1OT1 and is involved in multiple epigenetic disorders. Here, we found that pigs also had a relatively conserved paternal allele expressing KCNQ1OT1 and had a shorter 5' end (∼27 kb) compared to human KCNQ1OT1. Knockdown of KCNQ1OT1 using antisense oligonucleotides (ASO) showed that upregulation of CDKN1C expression in pigs. However, porcine KCNQ1OT1 did not affect the DNA methylation status of the CpG islands in the promoters of KCNQ1OT1 and CDKN1C. Inhibition of DNA methyltransferase using Decitabine treatment resulted in a significant increase in both KCNQ1OT1 and CDKN1C expression, suggesting that the regulation between KCNQ1OT1 and CDKN1C may not be dependent on RNA interference. Further use of chromosome conformation capture and reverse transcription-associated trap detection in the region where CDKN1C was located revealed that KCNQ1OT1 bound to the CDKN1C promoter and affected chromosome folding. Phenotypically, inhibition of KCNQ1OT1 at the cumulus-oocyte complex promoted cumulus cell transformation, and to upregulated the expression of ALPL at the early stage of osteogenic differentiation of porcine bone marrow mesenchymal stem cells. Our results confirm that the expression of KCNQ1OT1 imprinting in pigs as well as porcine KCNQ1OT1 regulates the expression of CDKN1C through direct promoter binding and chromatin folding alteration. And this regulatory mechanism played an important role in cell differentiation.


Assuntos
Cromatina , Inibidor de Quinase Dependente de Ciclina p57 , Metilação de DNA , Impressão Genômica , Regiões Promotoras Genéticas , RNA Longo não Codificante , Animais , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Suínos , Inibidor de Quinase Dependente de Ciclina p57/genética , Inibidor de Quinase Dependente de Ciclina p57/metabolismo , Cromatina/genética , Cromatina/metabolismo , Canais de Potássio de Abertura Dependente da Tensão da Membrana/genética , Canais de Potássio de Abertura Dependente da Tensão da Membrana/metabolismo , Ilhas de CpG , Regulação da Expressão Gênica
18.
Mar Drugs ; 22(5)2024 May 13.
Artigo em Inglês | MEDLINE | ID: mdl-38786608

RESUMO

We identified a new human voltage-gated potassium channel blocker, NnK-1, in the jellyfish Nemopilema nomurai based on its genomic information. The gene sequence encoding NnK-1 contains 5408 base pairs, with five introns and six exons. The coding sequence of the NnK-1 precursor is 894 nucleotides long and encodes 297 amino acids containing five presumptive ShK-like peptides. An electrophysiological assay demonstrated that the fifth peptide, NnK-1, which was chemically synthesized, is an effective blocker of hKv1.3, hKv1.4, and hKv1.5. Multiple-sequence alignment with cnidarian Shk-like peptides, which have Kv1.3-blocking activity, revealed that three residues (3Asp, 25Lys, and 34Thr) of NnK-1, together with six cysteine residues, were conserved. Therefore, we hypothesized that these three residues are crucial for the binding of the toxin to voltage-gated potassium channels. This notion was confirmed by an electrophysiological assay with a synthetic peptide (NnK-1 mu) where these three peptides were substituted with 3Glu, 25Arg, and 34Met. In conclusion, we successfully identified and characterized a new voltage-gated potassium channel blocker in jellyfish that interacts with three different voltage-gated potassium channels. A peptide that interacts with multiple voltage-gated potassium channels has many therapeutic applications in various physiological and pathophysiological contexts.


Assuntos
Peptídeos , Bloqueadores dos Canais de Potássio , Canais de Potássio de Abertura Dependente da Tensão da Membrana , Cifozoários , Animais , Humanos , Bloqueadores dos Canais de Potássio/farmacologia , Bloqueadores dos Canais de Potássio/química , Canais de Potássio de Abertura Dependente da Tensão da Membrana/antagonistas & inibidores , Peptídeos/farmacologia , Peptídeos/química , Sequência de Aminoácidos , Venenos de Cnidários/farmacologia , Venenos de Cnidários/química , Alinhamento de Sequência
19.
Int J Mol Sci ; 25(9)2024 Apr 24.
Artigo em Inglês | MEDLINE | ID: mdl-38731872

RESUMO

Numerous studies suggest the involvement of adenosine-5'-triphosphate (ATP) and similar nucleotides in the pathophysiology of asthma. Androgens, such as testosterone (TES), are proposed to alleviate asthma symptoms in young men. ATP and uridine-5'-triphosphate (UTP) relax the airway smooth muscle (ASM) via purinergic P2Y2 and P2Y4 receptors and K+ channel opening. We previously demonstrated that TES increased the expression of voltage-dependent K+ (KV) channels in ASM. This study investigates how TES may potentiate ASM relaxation induced by ATP and UTP. Tracheal tissues treated with or without TES (control group) from young male guinea pigs were used. In organ baths, tracheas exposed to TES (40 nM for 48 h) showed enhanced ATP- and UTP-evoked relaxation. Tetraethylammonium, a K+ channel blocker, annulled this effect. Patch-clamp experiments in tracheal myocytes showed that TES also increased ATP- and UTP-induced K+ currents, and this effect was abolished with flutamide (an androgen receptor antagonist). KV channels were involved in this phenomenon, which was demonstrated by inhibition with 4-aminopyridine. RB2 (an antagonist of almost all P2Y receptors except for P2Y2), as well as N-ethylmaleimide and SQ 22,536 (inhibitors of G proteins and adenylyl cyclase, respectively), attenuated the enhancement of the K+ currents induced by TES. Immunofluorescence and immunohistochemistry studies revealed that TES did not modify the expression of P2Y4 receptors or COX-1 and COX-2, while we have demonstrated that this androgen augmented the expression of KV1.2 and KV1.5 channels in ASM. Thus, TES leads to the upregulation of P2Y4 signaling and KV channels in guinea pig ASM, enhancing ATP and UTP relaxation responses, which likely limits the severity of bronchospasm in young males.


Assuntos
Trifosfato de Adenosina , Adenilil Ciclases , Relaxamento Muscular , Músculo Liso , Testosterona , Traqueia , Uridina Trifosfato , Animais , Uridina Trifosfato/farmacologia , Uridina Trifosfato/metabolismo , Cobaias , Relaxamento Muscular/efeitos dos fármacos , Masculino , Trifosfato de Adenosina/metabolismo , Traqueia/metabolismo , Traqueia/efeitos dos fármacos , Testosterona/farmacologia , Testosterona/metabolismo , Adenilil Ciclases/metabolismo , Músculo Liso/metabolismo , Músculo Liso/efeitos dos fármacos , Canais de Potássio de Abertura Dependente da Tensão da Membrana/metabolismo , Transdução de Sinais/efeitos dos fármacos , Receptores Purinérgicos P2/metabolismo
20.
J Med Chem ; 67(11): 9124-9149, 2024 Jun 13.
Artigo em Inglês | MEDLINE | ID: mdl-38782404

RESUMO

Gain-of-function (GoF) variants in KCNT1 channels cause severe, drug-resistant forms of epilepsy. Quinidine is a known KCNT1 blocker, but its clinical use is limited due to severe drawbacks. To identify novel KCNT1 blockers, a homology model of human KCNT1 was built and used to screen an in-house library of compounds. Among the 20 molecules selected, five (CPK4, 13, 16, 18, and 20) showed strong KCNT1-blocking ability in an in vitro fluorescence-based assay. Patch-clamp experiments confirmed a higher KCNT1-blocking potency of these compounds when compared to quinidine, and their selectivity for KCNT1 over hERG and Kv7.2 channels. Among identified molecules, CPK20 displayed the highest metabolic stability; this compound also blocked KCNT2 currents, although with a lower potency, and counteracted GoF effects prompted by 2 recurrent epilepsy-causing KCNT1 variants (G288S and A934T). The present results provide solid rational basis for future design of novel compounds to counteract KCNT1-related neurological disorders.


Assuntos
Epilepsia , Humanos , Epilepsia/tratamento farmacológico , Epilepsia/metabolismo , Canais de Potássio de Abertura Dependente da Tensão da Membrana/metabolismo , Canais de Potássio de Abertura Dependente da Tensão da Membrana/antagonistas & inibidores , Bloqueadores dos Canais de Potássio/farmacologia , Bloqueadores dos Canais de Potássio/síntese química , Bloqueadores dos Canais de Potássio/química , Animais , Proteínas do Tecido Nervoso/metabolismo , Proteínas do Tecido Nervoso/antagonistas & inibidores , Proteínas do Tecido Nervoso/genética , Relação Estrutura-Atividade , Células HEK293 , Simulação por Computador , Canais de Potássio Ativados por Sódio
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