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17 pages, 3921 KiB

Open AccessArticle

TRPV1 Activation Promotes β-arrestin2 Interaction with the Ribosomal Biogenesis Machinery in the Nucleolus: Implications for p53 Regulation and Neurite Outgrowth

by Ahmed Hassan, Mircea Iftinca, Daniel Young, Robyn Flynn, Francina Agosti, Nasser Abdullah, Manon Defaye, Mark G. H. Scott, Antoine Dufour and Christophe Altier

Int. J. Mol. Sci. 2021, 22(5), 2280; https://doi.org/10.3390/ijms22052280 - 25 Feb 2021

Cited by 1 | Viewed by 3107

Abstract

Transient receptor potential vanilloids (TRPV1) are non-selective cation channels that sense and transduce inflammatory pain signals. We previously reported that activation of TRPV1 induced the translocation of β-arrestin2 (ARRB2) from the cytoplasm to the nucleus, raising questions about the functional role of ARRB2 [...] Read more.

Transient receptor potential vanilloids (TRPV1) are non-selective cation channels that sense and transduce inflammatory pain signals. We previously reported that activation of TRPV1 induced the translocation of β-arrestin2 (ARRB2) from the cytoplasm to the nucleus, raising questions about the functional role of ARRB2 in the nucleus. Here, we determined the ARRB2 nuclear signalosome by conducting a quantitative proteomic analysis of the nucleus-sequestered L395Q ARRB2 mutant, compared to the cytosolic wild-type ARRB2 (WT ARRB2), in a heterologous expression system. We identified clusters of proteins that localize to the nucleolus and are involved in ribosomal biogenesis. Accordingly, L395Q ARRB2 or WT ARRB2 after capsaicin treatment were found to co-localize and interact with the nucleolar marker nucleophosmin (NPM1), treacle protein (TCOF1) and RNA polymerase I (POL I). We further investigated the role of nuclear ARRB2 signaling in regulating neuroplasticity. Using neuroblastoma (neuro2a) cells and dorsal root ganglia (DRG) neurons, we found that L395Q ARRB2 mutant increased POL I activity, inhibited the tumor suppressorp53 (p53) level and caused a decrease in the outgrowth of neurites. Together, our results suggest that the activation of TRPV1 promotes the ARRB2-mediated regulation of ribosomal biogenesis in the nucleolus. The ARRB2-TCOF1-p53 checkpoint signaling pathway might be involved in regulating neurite outgrowth associated with pathological pain conditions. Full article

(This article belongs to the Special Issue Ion Channel and Ion-Related Signaling 2020)

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15 pages, 2384 KiB

Open AccessArticle

Ca²⁺-Dependent Protein Kinase 6 Enhances KAT2 Shaker Channel Activity in Arabidopsis thaliana

by Elsa Ronzier, Claire Corratgé-Faillie, Frédéric Sanchez, Christian Brière and Tou Cheu Xiong

Int. J. Mol. Sci. 2021, 22(4), 1596; https://doi.org/10.3390/ijms22041596 - 5 Feb 2021

Cited by 3 | Viewed by 2749

Abstract

Post-translational regulations of Shaker-like voltage-gated K⁺ channels were reported to be essential for rapid responses to environmental stresses in plants. In particular, it has been shown that calcium-dependent protein kinases (CPKs) regulate Shaker channels in plants. Here, the focus was on KAT2, [...] Read more.

Post-translational regulations of Shaker-like voltage-gated K⁺ channels were reported to be essential for rapid responses to environmental stresses in plants. In particular, it has been shown that calcium-dependent protein kinases (CPKs) regulate Shaker channels in plants. Here, the focus was on KAT2, a Shaker channel cloned in the model plant Arabidopsis thaliana, where is it expressed namely in the vascular tissues of leaves. After co-expression of KAT2 with AtCPK6 in Xenopuslaevis oocytes, voltage-clamp recordings demonstrated that AtCPK6 stimulates the activity of KAT2 in a calcium-dependent manner. A physical interaction between these two proteins has also been shown by Förster resonance energy transfer by fluorescence lifetime imaging (FRET-FLIM). Peptide array assays support that AtCPK6 phosphorylates KAT2 at several positions, also in a calcium-dependent manner. Finally, K⁺ fluorescence imaging in planta suggests that K⁺ distribution is impaired in kat2 knock-out mutant leaves. We propose that the AtCPK6/KAT2 couple plays a role in the homeostasis of K⁺ distribution in leaves. Full article

(This article belongs to the Special Issue Ion Channel and Ion-Related Signaling 2020)

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12 pages, 1714 KiB

Open AccessArticle

Tyrosine Phosphorylation of the K_v2.1 Channel Contributes to Injury in Brain Ischemia

by Min-Young Song, Ji Yeon Hwang, Eun Ji Bae, Saesbyeol Kim, Hye-Min Kang, Yong Jun Kim, Chan Park and Kang-Sik Park

Int. J. Mol. Sci. 2020, 21(24), 9538; https://doi.org/10.3390/ijms21249538 - 15 Dec 2020

Cited by 1 | Viewed by 2503

Abstract

In brain ischemia, oxidative stress induces neuronal apoptosis, which is mediated by increased activity of the voltage-gated K⁺ channel K_v2.1 and results in an efflux of intracellular K⁺. The molecular mechanisms underlying the regulation of K_v2.1 [...] Read more.

In brain ischemia, oxidative stress induces neuronal apoptosis, which is mediated by increased activity of the voltage-gated K⁺ channel K_v2.1 and results in an efflux of intracellular K⁺. The molecular mechanisms underlying the regulation of K_v2.1 and its activity during brain ischemia are not yet fully understood. Here this study provides evidence that oxidant-induced apoptosis resulting from brain ischemia promotes rapid tyrosine phosphorylation of K_v2.1. When the tyrosine phosphorylation sites Y124, Y686, and Y810 on the K_v2.1 channel are mutated to non-phosphorylatable residues, PARP-1 cleavage levels decrease, indicating suppression of neuronal cell death. The tyrosine residue Y810 on K_v2.1 was a major phosphorylation site. In fact, cells mutated Y810 were more viable in our study than were wild-type cells, suggesting an important role for this site during ischemic neuronal injury. In an animal model, tyrosine phosphorylation of K_v2.1 increased after ischemic brain injury, with an observable sustained increase for at least 2 h after reperfusion. These results demonstrate that tyrosine phosphorylation of the K_v2.1 channel in the brain may play a critical role in regulating neuronal ischemia and is therefore a potential therapeutic target in patients with brain ischemia. Full article

(This article belongs to the Special Issue Ion Channel and Ion-Related Signaling 2020)

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16 pages, 2291 KiB

Open AccessArticle

Inter-Regulation of K_v4.3 and Voltage-Gated Sodium Channels Underlies Predisposition to Cardiac and Neuronal Channelopathies

by Jérôme Clatot, Nathalie Neyroud, Robert Cox, Charlotte Souil, Jing Huang, Pascale Guicheney and Charles Antzelevitch

Int. J. Mol. Sci. 2020, 21(14), 5057; https://doi.org/10.3390/ijms21145057 - 17 Jul 2020

Cited by 14 | Viewed by 3059

Abstract

Background: Genetic variants in voltage-gated sodium channels (Na_v) encoded by SCNXA genes, responsible for I_Na, and K_v4.3 channels encoded by KCND3, responsible for the transient outward current (I_to), contribute to the manifestation of both [...] Read more.

Background: Genetic variants in voltage-gated sodium channels (Na_v) encoded by SCNXA genes, responsible for I_Na, and K_v4.3 channels encoded by KCND3, responsible for the transient outward current (I_to), contribute to the manifestation of both Brugada syndrome (BrS) and spinocerebellar ataxia (SCA19/22). We examined the hypothesis that K_v4.3 and Na_v variants regulate each other’s function, thus modulating I_Na/I_to balance in cardiomyocytes and I_Na/I_(A) balance in neurons. Methods: Bicistronic and other constructs were used to express WT or variant Na_v1.5 and K_v4.3 channels in HEK293 cells. I_Na and I_to were recorded. Results: SCN5A variants associated with BrS reduced I_Na, but increased I_to. Moreover, BrS and SCA19/22 KCND3 variants associated with a gain of function of I_to, significantly reduced I_Na, whereas the SCA19/22 KCND3 variants associated with a loss of function (LOF) of I_to significantly increased I_Na. Auxiliary subunits Na_vβ1, MiRP3 and KChIP2 also modulated I_Na/I_to balance. Co-immunoprecipitation and Duolink studies suggested that the two channels interact within the intracellular compartments and biotinylation showed that LOF SCN5A variants can increase K_v4.3 cell-surface expression. Conclusion: Na_v and K_v4.3 channels modulate each other’s function via trafficking and gating mechanisms, which have important implications for improved understanding of these allelic cardiac and neuronal syndromes. Full article

(This article belongs to the Special Issue Ion Channel and Ion-Related Signaling 2020)

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12 pages, 1354 KiB

Open AccessArticle

GABA_A Receptor/STEP61 Signaling Pathway May Be Involved in Emulsified Isoflurane Anesthesia in Rats

by Xingkai Zhao, Guangjun Chang, Yan Cheng and Zhenlei Zhou

Int. J. Mol. Sci. 2020, 21(11), 4078; https://doi.org/10.3390/ijms21114078 - 7 Jun 2020

Cited by 5 | Viewed by 2297

Abstract

(1) Background: Emulsified isoflurane (EISO) is a type of intravenous anesthetic. How emulsified isoflurane works in the brain is still unclear. The aim of this study was to explore whether epigenetic mechanisms affect anesthesia and to evaluate the anesthetic effects of emulsified isoflurane [...] Read more.

(1) Background: Emulsified isoflurane (EISO) is a type of intravenous anesthetic. How emulsified isoflurane works in the brain is still unclear. The aim of this study was to explore whether epigenetic mechanisms affect anesthesia and to evaluate the anesthetic effects of emulsified isoflurane in rats. (2) Methods: Rats were randomly divided into four groups (n = 8/group): The tail vein was injected with normal saline 0.1 mL·kg⁻¹·min⁻¹ for the control (Con) group, with intralipid for the fat emulsion (FE) group, with EISO at 60 mg·kg⁻¹·min⁻¹ for the high-concentration (HD) group, and 45 mg·kg⁻¹·min⁻¹ for the low-concentration (LD) group. The consciousness state, motor function of limbs, and response to nociceptive stimulus were observed after drug administration. (3) Results: Using real-time polymerase chain reaction (PCR) to assess the promoter methylation of ion channel proteins in the cerebral cortex of rats anesthetized by EISO, we demonstrated that the change in the promoters’ methylation of the coding genes for gamma-aminobutyric acid A receptor α1 subunit (GABA_Aα1), N-methyl-D-aspartate receptor subunit 1 (NMDAR1), and mu opioid receptor 1 (OPRM1) was accompanied by the change in messenger ribonucleic acid (mRNA) and protein expression by these genes. (4) Conclusion: These data suggest that the epigenetic factors’ modulation might offer a novel approach to explore the anesthetic mechanism of EISO. Full article

(This article belongs to the Special Issue Ion Channel and Ion-Related Signaling 2020)

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18 pages, 3229 KiB

Open AccessArticle

Interactive Actions of Aldosterone and Insulin on Epithelial Na⁺ Channel Trafficking

by Rie Marunaka and Yoshinori Marunaka

Int. J. Mol. Sci. 2020, 21(10), 3407; https://doi.org/10.3390/ijms21103407 - 12 May 2020

Cited by 5 | Viewed by 2794

Abstract

Epithelial Na⁺ channel (ENaC) participates in renal epithelial Na⁺ reabsorption, controlling blood pressure. Aldosterone and insulin elevate blood pressure by increasing the ENaC-mediated Na⁺ reabsorption. However, little information is available on the interactive action of aldosterone and insulin on the [...] Read more.

Epithelial Na⁺ channel (ENaC) participates in renal epithelial Na⁺ reabsorption, controlling blood pressure. Aldosterone and insulin elevate blood pressure by increasing the ENaC-mediated Na⁺ reabsorption. However, little information is available on the interactive action of aldosterone and insulin on the ENaC-mediated Na⁺ reabsorption. In the present study, we tried to clarify if insulin would modify the aldosterone action on the ENaC-mediated Na⁺ reabsorption from a viewpoint of intracellular ENaC trafficking. We measured the ENaC-mediated Na⁺ transport as short-circuit currents using a four-state mathematical ENaC trafficking model in renal A6 epithelial cells with or without aldosterone treatment under the insulin-stimulated and -unstimulated conditions. We found that: (A) under the insulin-stimulated condition, aldosterone treatment (1 µM for 20 h) significantly elevated the ENaC insertion rate to the apical membrane (

k_{I}

) 3.3-fold and the ENaC recycling rate (

k_{R}

) 2.0-fold, but diminished the ENaC degradation rate (

k_{D}

) 0.7-fold without any significant effect on the ENaC endocytotic rate (

k_{E}

); (B) under the insulin-unstimulated condition, aldosterone treatment decreased

k_{E}

0.5-fold and increased

k_{R}

1.4-fold, without any significant effect on

k_{I}

or

k_{D}

. Thus, the present study indicates that: (1) insulin masks the well-known inhibitory action of aldosterone on the ENaC endocytotic rate; (2) insulin induces a stimulatory action of aldosterone on ENaC apical insertion and an inhibitory action of aldosterone on ENaC degradation; (3) insulin enhances the aldosterone action on ENaC recycling; (4) insulin has a more effective action on diminution of ENaC endocytosis than aldosterone. Full article

(This article belongs to the Special Issue Ion Channel and Ion-Related Signaling 2020)

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16 pages, 4505 KiB

Open AccessArticle

Possible Contribution of Inflammation-Associated Hypoxia to Increased K_2P5.1 K⁺ Channel Expression in CD4⁺ T Cells of the Mouse Model for Inflammatory Bowel Disease

by Kyoko Endo, Hiroaki Kito, Ryo Tanaka, Junko Kajikuri, Satoshi Tanaka, Elghareeb E. Elboray, Takayoshi Suzuki and Susumu Ohya

Int. J. Mol. Sci. 2020, 21(1), 38; https://doi.org/10.3390/ijms21010038 - 19 Dec 2019

Cited by 7 | Viewed by 3866

Abstract

Previous studies have reported the up-regulation of the two-pore domain K⁺ channel K_2P5.1 in the CD4⁺ T cells of patients with multiple sclerosis (MS) and rheumatoid arthritis (RA), as well as in a mouse model of inflammatory bowel disease [...] Read more.

Previous studies have reported the up-regulation of the two-pore domain K⁺ channel K_2P5.1 in the CD4⁺ T cells of patients with multiple sclerosis (MS) and rheumatoid arthritis (RA), as well as in a mouse model of inflammatory bowel disease (IBD). However, the mechanisms underlying this up-regulation remain unclear. Inflammation-associated hypoxia is involved in the pathogenesis of autoimmune diseases, such as IBD, MS, and RA, and T cells are exposed to a hypoxic environment during their recruitment from inflamed tissues to secondary lymphoid tissues. We herein investigated whether inflammation-associated hypoxia is attributable to the increased expression and activity of K_2P5.1 in the splenic CD4⁺ T cells of chemically-induced IBD model mice. Significant increases in hypoxia-inducible factor (HIF)-1α transcripts and proteins were found in the splenic CD4⁺ T cells of the IBD model. In the activated splenic CD4⁺ T cells, hypoxia (1.5% O₂) increased K_2P5.1 expression and activity, whereas a treatment with the HIF inhibitor FM19G11 but not the selective HIF-2 inhibitor exerted the opposite effect. Hypoxia-exposed K_2P5.1 up-regulation was also detected in stimulated thymocytes and the mouse T-cell line. The class III histone deacetylase sirtuin-1 (SIRT1) is a downstream molecule of HIF-1α signaling. We examined the effects of the SIRT1 inhibitor NCO-01 on K_2P5.1 transcription in activated CD4⁺ T cells, and we found no significant effects on the K_2P5.1 transcription. No acute compensatory responses of K_2P3.1–K_2P5.1 up-regulation were found in the CD4⁺ T cells of the IBD model and the hypoxia-exposed T cells. Collectively, these results suggest a mechanism for K_2P5.1 up-regulation via HIF-1 in the CD4⁺ T cells of the IBD model. Full article

(This article belongs to the Special Issue Ion Channel and Ion-Related Signaling 2019)

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15 pages, 4579 KiB

Open AccessArticle

Windup of Nociceptive Flexion Reflex Depends on Synaptic and Intrinsic Properties of Dorsal Horn Neurons in Adult Rats

by Franck Aby, Rabia Bouali-Benazzouz, Marc Landry and Pascal Fossat

Int. J. Mol. Sci. 2019, 20(24), 6146; https://doi.org/10.3390/ijms20246146 - 5 Dec 2019

Cited by 5 | Viewed by 3637

Abstract

Windup, a progressive increase in spinal response to repetitive stimulations of nociceptive peripheral fibers, is a useful model to study central sensitization to pain. Windup is expressed by neurons in both the dorsal and ventral horn of the spinal cord. In juvenile rats, [...] Read more.

Windup, a progressive increase in spinal response to repetitive stimulations of nociceptive peripheral fibers, is a useful model to study central sensitization to pain. Windup is expressed by neurons in both the dorsal and ventral horn of the spinal cord. In juvenile rats, it has been demonstrated both in vivo and in vitro that windup depends on calcium-dependent intrinsic properties and their modulation by synaptic components. However, the involvement of these two components in the adults remains controversial. In the present study, by means of electromyographic and extracellular recordings, we show that windup in adults, in vivo, depends on a synaptic balance between excitatory N-methyl-D-aspartate (NMDA) receptors and inhibitory glycinergic receptors. We also demonstrate the involvement of L-type calcium channels in both the dorsal and ventral horn of the spinal cord. These results indicate that windup in adults is similar to juvenile rats and that windup properties are the same regardless of the spinal network, i.e., sensory or motor. Full article

(This article belongs to the Special Issue Ion Channel and Ion-Related Signaling 2019)

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12 pages, 1951 KiB

Open AccessArticle

RBM20 Regulates CaV1.2 Surface Expression by Promoting Exon 9* Inclusion of CACNA1C in Neonatal Rat Cardiomyocytes

by Akihito Morinaga, Jumpei Ito, Tomoaki Niimi and Andrés D. Maturana

Int. J. Mol. Sci. 2019, 20(22), 5591; https://doi.org/10.3390/ijms20225591 - 8 Nov 2019

Cited by 6 | Viewed by 3280

Abstract

The CACNA1C gene encodes for the CaV1.2 protein, which is the pore subunit of cardiac l-type voltage-gated calcium (Ca²⁺) channels (l-channels). Through alternative splicing, CACNA1C encodes for various CaV1.2 isoforms with different electrophysiological properties. Splice variants of CaV1.2 [...] Read more.

The CACNA1C gene encodes for the CaV1.2 protein, which is the pore subunit of cardiac l-type voltage-gated calcium (Ca²⁺) channels (l-channels). Through alternative splicing, CACNA1C encodes for various CaV1.2 isoforms with different electrophysiological properties. Splice variants of CaV1.2 are differentially expressed during heart development or pathologies. The molecular mechanisms of CACNA1C alternative splicing still remain incompletely understood. RNA sequencing analysis has suggested that CACNA1C is a potential target of the splicing factor RNA-binding protein motif 20 (RBM20). Here, we aimed at elucidating the role of RBM20 in the regulation of CACNA1C alternative splicing. We found that in neonatal rat cardiomyocytes (NRCMs), RBM20 overexpression promoted the inclusion of CACNA1C’s exon 9*, whereas the skipping of exon 9* occurred upon RBM20 siRNA knockdown. The splicing of other known alternative exons was not altered by RBM20. RNA immunoprecipitation suggested that RBM20 binds to introns flanking exon 9*. Functionally, in NRCMs, RBM20 overexpression decreased l-type Ca²⁺ currents, whereas RBM20 siRNA knockdown increased l-type Ca²⁺ currents. Finally, we found that RBM20 overexpression reduced CaV1.2 membrane surface expression in NRCMs. Taken together, our results suggest that RBM20 specifically regulates the inclusion of exon 9* in CACNA1C mRNA, resulting in reduced cell-surface membrane expression of l-channels in cardiomyocytes. Full article

(This article belongs to the Special Issue Ion Channel and Ion-Related Signaling 2019)

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18 pages, 2965 KiB

Open AccessArticle

Single-Channel Properties of the ROMK-Pore-Forming Subunit of the Mitochondrial ATP-Sensitive Potassium Channel

by Michał Laskowski, Bartłomiej Augustynek, Piotr Bednarczyk, Monika Żochowska, Justyna Kalisz, Brian O’Rourke, Adam Szewczyk and Bogusz Kulawiak

Int. J. Mol. Sci. 2019, 20(21), 5323; https://doi.org/10.3390/ijms20215323 - 25 Oct 2019

Cited by 32 | Viewed by 4784

Abstract

An increased flux of potassium ions into the mitochondrial matrix through the ATP-sensitive potassium channel (mitoK_ATP) has been shown to provide protection against ischemia-reperfusion injury. Recently, it was proposed that the mitochondrial-targeted isoform of the renal outer medullary potassium channel (ROMK) [...] Read more.

An increased flux of potassium ions into the mitochondrial matrix through the ATP-sensitive potassium channel (mitoK_ATP) has been shown to provide protection against ischemia-reperfusion injury. Recently, it was proposed that the mitochondrial-targeted isoform of the renal outer medullary potassium channel (ROMK) protein creates a pore-forming subunit of mitoK_ATP in heart mitochondria. Our research focuses on the properties of mitoK_ATP from heart-derived H9c2 cells. For the first time, we detected single-channel activity and describe the pharmacology of mitoK_ATP in the H9c2 heart-derived cells. The patch-clamping of mitoplasts from wild type (WT) and cells overexpressing ROMK2 revealed the existence of a potassium channel that exhibits the same basic properties previously attributed to mitoK_ATP. ROMK2 overexpression resulted in a significant increase of mitoK_ATP activity. The conductance of both channels in symmetric 150/150 mM KCl was around 97 ± 2 pS in WT cells and 94 ± 3 pS in cells overexpressing ROMK2. The channels were inhibited by 5-hydroxydecanoic acid (a mitoK_ATP inhibitor) and by Tertiapin Q (an inhibitor of both the ROMK-type channels and mitoK_ATP). Additionally, mitoK_ATP from cells overexpressing ROMK2 were inhibited by ATP/Mg²⁺ and activated by diazoxide. We used an assay based on proteinase K to examine the topology of the channel in the inner mitochondrial membrane and found that both termini of the protein localized to the mitochondrial matrix. We conclude that the observed activity of the channel formed by the ROMK protein corresponds to the electrophysiological and pharmacological properties of mitoK_ATP. Full article

(This article belongs to the Special Issue Ion Channel and Ion-Related Signaling 2019)

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19 pages, 2798 KiB

Open AccessArticle

Functional Consequences of the SCN5A-p.Y1977N Mutation within the PY Ubiquitylation Motif: Discrepancy between HEK293 Cells and Transgenic Mice

by Simona Casini, Maxime Albesa, Zizun Wang, Vincent Portero, Daniela Ross-Kaschitza, Jean-Sébastien Rougier, Gerard A. Marchal, Wendy K. Chung, Connie R. Bezzina, Hugues Abriel and Carol Ann Remme

Int. J. Mol. Sci. 2019, 20(20), 5033; https://doi.org/10.3390/ijms20205033 - 11 Oct 2019

Cited by 9 | Viewed by 4150

Abstract

Dysfunction of the cardiac sodium channel Nav1.5 (encoded by the SCN5A gene) is associated with arrhythmias and sudden cardiac death. SCN5A mutations associated with long QT syndrome type 3 (LQT3) lead to enhanced late sodium current and consequent action potential (AP) prolongation. Internalization [...] Read more.

Dysfunction of the cardiac sodium channel Nav1.5 (encoded by the SCN5A gene) is associated with arrhythmias and sudden cardiac death. SCN5A mutations associated with long QT syndrome type 3 (LQT3) lead to enhanced late sodium current and consequent action potential (AP) prolongation. Internalization and degradation of Na_v1.5 is regulated by ubiquitylation, a post-translational mechanism that involves binding of the ubiquitin ligase Nedd4-2 to a proline-proline-serine-tyrosine sequence of Na_v1.5, designated the PY-motif. We investigated the biophysical properties of the LQT3-associated SCN5A-p.Y1977N mutation located in the Na_v1.5 PY-motif, both in HEK293 cells as well as in newly generated mice harboring the mouse homolog mutation Scn5a-p.Y1981N. We found that in HEK293 cells, the SCN5A-p.Y1977N mutation abolished the interaction between Na_v1.5 and Nedd4-2, suppressed PY-motif-dependent ubiquitylation of Na_v1.5, and consequently abrogated Nedd4-2 induced sodium current (I_Na) decrease. Nevertheless, homozygous mice harboring the Scn5a-p.Y1981N mutation showed no electrophysiological alterations nor changes in AP or (late) I_Na properties, questioning the in vivo relevance of the PY-motif. Our findings suggest the presence of compensatory mechanisms, with additional, as yet unknown, factors likely required to reduce the “ubiquitylation reserve” of Na_v1.5. Future identification of such modulatory factors may identify potential triggers for arrhythmias and sudden cardiac death in the setting of LQT3 mutations. Full article

(This article belongs to the Special Issue Ion Channel and Ion-Related Signaling 2019)

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20 pages, 3535 KiB

Open AccessArticle

PIEZO1 and TRPV4, which Are Distinct Mechano-Sensors in the Osteoblastic MC3T3-E1 Cells, Modify Cell-Proliferation

by Maki Yoneda, Hiroka Suzuki, Noriyuki Hatano, Sayumi Nakano, Yukiko Muraki, Ken Miyazawa, Shigemi Goto and Katsuhiko Muraki

Int. J. Mol. Sci. 2019, 20(19), 4960; https://doi.org/10.3390/ijms20194960 - 8 Oct 2019

Cited by 65 | Viewed by 7220

Abstract

Mechanical-loading and unloading can modify osteoblast functioning. Ca²⁺ signaling is one of the earliest events in osteoblasts to induce a mechanical stimulus, thereby demonstrating the importance of the underlying mechanical sensors for the sensation. Here, we examined the mechano-sensitive channels PIEZO1 and [...] Read more.

Mechanical-loading and unloading can modify osteoblast functioning. Ca²⁺ signaling is one of the earliest events in osteoblasts to induce a mechanical stimulus, thereby demonstrating the importance of the underlying mechanical sensors for the sensation. Here, we examined the mechano-sensitive channels PIEZO1 and TRPV4 were involved in the process of mechano-sensation in the osteoblastic MC3T3-E1 cells. The analysis of mRNA expression revealed a high expression of Piezo1 and Trpv4 in these cells. We also found that a PIEZO1 agonist, Yoda1, induced Ca²⁺ response and activated cationic currents in these cells. Ca²⁺ response was elicited when mechanical stimulation (MS), with shear stress, was induced by fluid flow in the MC3T3-E1 cells. Gene knockdown of Piezo1 in the MC3T3-E1 cells, by transfection with siPiezo1, inhibited the Yoda1-induced response, but failed to inhibit the MS-induced response. When MC3T3-E1 cells were transfected with siTrpv4, the MS-induced response was abolished and Yoda1 response was attenuated. Moreover, the MS-induced response was inhibited by a TRPV4 antagonist HC-067047 (HC). Yoda1 response was also inhibited by HC in MC3T3-E1 cells and HEK cells, expressing both PIEZO1 and TRPV4. Meanwhile, the activation of PIEZO1 and TRPV4 reduced the proliferation of MC3T3-E1, which was reversed by knockdown of PIEZO1, and TRPV4, respectively. In conclusion, TRPV4 and PIEZO1 are distinct mechano-sensors in the MC3T3-E1 cells. However, PIEZO1 and TRPV4 modify the proliferation of these cells, implying that PIEZO1 and TRPV4 may be functional in the osteoblastic mechano-transduction. Notably, it is also found that Yoda1 can induce TRPV4-dependent Ca²⁺ response, when both PIEZO1 and TRPV4 are highly expressed. Full article

(This article belongs to the Special Issue Ion Channel and Ion-Related Signaling 2019)

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14 pages, 2247 KiB

Open AccessArticle

Castration Induces Down-Regulation of A-Type K⁺ Channel in Rat Vas Deferens Smooth Muscle

by Susumu Ohya, Katsunori Ito, Noriyuki Hatano, Akitoshi Ohno, Katsuhiko Muraki and Yuji Imaizumi

Int. J. Mol. Sci. 2019, 20(17), 4073; https://doi.org/10.3390/ijms20174073 - 21 Aug 2019

Cited by 3 | Viewed by 3193

Abstract

A-type K⁺ channels contribute to regulating the propagation and frequency of action potentials in smooth muscle cells (SMCs). The present study (i) identified the molecular components of A-type K⁺ channels in rat vas deferens SMs (VDSMs) and (ii) showed the long-term, [...] Read more.

A-type K⁺ channels contribute to regulating the propagation and frequency of action potentials in smooth muscle cells (SMCs). The present study (i) identified the molecular components of A-type K⁺ channels in rat vas deferens SMs (VDSMs) and (ii) showed the long-term, genomic effects of testosterone on their expression in VDSMs. Transcripts of the A-type K⁺ channel α subunit, Kv4.3L and its regulatory β subunits, KChIP3, NCS1, and DPP6-S were predominantly expressed in rat VDSMs over the other related subtypes (Kv4.2, KChIP1, KChIP2, KChIP4, and DPP10). A-type K⁺ current (I_A) density in VDSM cells (VDSMCs) was decreased by castration without changes in I_A kinetics, and decreased I_A density was compensated for by an oral treatment with 17α-methyltestosterone (MET). Correspondingly, in the VDSMs of castrated rats, Kv4.3L and KChIP3 were down-regulated at both the transcript and protein expression levels. Changes in Kv4.3L and KChIP3 expression levels were compensated for by the treatment with MET. These results suggest that testosterone level changes in testosterone disorders and growth processes control the functional expression of A-type K⁺ channels in VDSMCs. Full article

(This article belongs to the Special Issue Ion Channel and Ion-Related Signaling 2019)

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15 pages, 4594 KiB

Open AccessArticle

Formation Mechanism of Ion Channel in Channelrhodopsin-2: Molecular Dynamics Simulation and Steering Molecular Dynamics Simulations

by Ting Yang, Wenying Zhang, Jie Cheng, Yanhong Nie, Qi Xin, Shuai Yuan and Yusheng Dou

Int. J. Mol. Sci. 2019, 20(15), 3780; https://doi.org/10.3390/ijms20153780 - 2 Aug 2019

Cited by 12 | Viewed by 4760

Abstract

Channelrhodopsin-2 (ChR2) is a light-activated and non-selective cationic channel protein that can be easily expressed in specific neurons to control neuronal activity by light. Although ChR2 has been extensively used as an optogenetic tool in neuroscience research, the molecular mechanism of cation channel [...] Read more.

Channelrhodopsin-2 (ChR2) is a light-activated and non-selective cationic channel protein that can be easily expressed in specific neurons to control neuronal activity by light. Although ChR2 has been extensively used as an optogenetic tool in neuroscience research, the molecular mechanism of cation channel formation following retinal photoisomerization in ChR2 is not well understood. In this paper, studies of the closed and opened state ChR2 structures are presented. The formation of the cationic channel is elucidated in atomic detail using molecular dynamics simulations on the all-trans-retinal (ChR2-trans) configuration of ChR2 and its isomerization products, 13-cis-retinal (ChR2-cis) configuration, respectively. Photoisomerization of the retinal-chromophore causes the destruction of interactions among the crucial residues (e.g., E90, E82, N258, and R268) around the channel and the extended H-bond network mediated by numerous water molecules, which opens the pore. Steering molecular dynamics (SMD) simulations show that the electrostatic interactions at the binding sites in intracellular gate (ICG) and central gate (CG) can influence the transmembrane transport of Na⁺ in ChR2-cis obviously. Potential of mean force (PMF) constructed by SMD and umbrella sampling also found the existing energy wells at these two binding sites during the transportation of Na⁺. These wells partly hinder the penetration of Na⁺ into cytoplasm through the ion channel. This investigation provides a theoretical insight on the formation mechanism of ion channels and the mechanism of ion permeation. Full article

(This article belongs to the Special Issue Ion Channel and Ion-Related Signaling 2019)

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14 pages, 2102 KiB

Open AccessArticle

Extracellular l-arginine Enhances Relaxations Induced by Opening of Calcium-Activated SKCa Channels in Porcine Retinal Arteriole

by Ulf Simonsen, Anna K. Winther, Aida Oliván-Viguera, Simon Comerma-Steffensen, Ralf Köhler and Toke Bek

Int. J. Mol. Sci. 2019, 20(8), 2032; https://doi.org/10.3390/ijms20082032 - 25 Apr 2019

Cited by 1 | Viewed by 4158

Abstract

We investigated whether the substrate for nitric oxide (NO) production, extracellular l-arginine, contributes to relaxations induced by activating small (SKCa) conductance Ca²⁺-activated potassium channels. In endothelial cells, acetylcholine increased ³H-l-arginine uptake, while blocking the SKCa and the [...] Read more.

We investigated whether the substrate for nitric oxide (NO) production, extracellular l-arginine, contributes to relaxations induced by activating small (SKCa) conductance Ca²⁺-activated potassium channels. In endothelial cells, acetylcholine increased ³H-l-arginine uptake, while blocking the SKCa and the intermediate (IKCa) conductance Ca²⁺-activated potassium channels reduced l-arginine uptake. A blocker of the y+ transporter system, l-lysine also blocked ³H-l-arginine uptake. Immunostaining showed co-localization of endothelial NO synthase (eNOS), SKCa3, and the cationic amino acid transporter (CAT-1) protein of the y+ transporter system in the endothelium. An opener of SKCa channels, cyclohexyl-[2-(3,5-dimethyl-pyrazol-1-yl)-6-methyl-pyrimidin-4-yl]-amine (CyPPA) induced large currents in endothelial cells, and concentration-dependently relaxed porcine retinal arterioles. In the presence of l-arginine, concentration-response curves for CyPPA were leftward shifted, an effect unaltered in the presence of low sodium, but blocked by l-lysine in the retinal arterioles. Our findings suggest that SKCa channel activity regulates l-arginine uptake through the y+ transporter system, and we propose that in vasculature affected by endothelial dysfunction, l-arginine administration requires the targeting of additional mechanisms such as SKCa channels to restore endothelium-dependent vasodilatation. Full article

(This article belongs to the Special Issue Ion Channel and Ion-Related Signaling 2019)

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Review

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17 pages, 2184 KiB

Open AccessReview

The Ryanodine Receptor as a Sensor for Intracellular Environments in Muscles

by Takuya Kobayashi, Nagomi Kurebayashi and Takashi Murayama

Int. J. Mol. Sci. 2021, 22(19), 10795; https://doi.org/10.3390/ijms221910795 - 6 Oct 2021

Cited by 8 | Viewed by 3980

Abstract

The ryanodine receptor (RyR) is a Ca²⁺ release channel in the sarcoplasmic reticulum of skeletal and cardiac muscles and plays a key role in excitation–contraction coupling. The activity of the RyR is regulated by the changes in the level of many intracellular [...] Read more.

The ryanodine receptor (RyR) is a Ca²⁺ release channel in the sarcoplasmic reticulum of skeletal and cardiac muscles and plays a key role in excitation–contraction coupling. The activity of the RyR is regulated by the changes in the level of many intracellular factors, such as divalent cations (Ca²⁺ and Mg²⁺), nucleotides, associated proteins, and reactive oxygen species. Since these intracellular factors change depending on the condition of the muscle, e.g., exercise, fatigue, or disease states, the RyR channel activity will be altered accordingly. In this review, we describe how the RyR channel is regulated under various conditions and discuss the possibility that the RyR acts as a sensor for changes in the intracellular environments in muscles. Full article

(This article belongs to the Special Issue Ion Channel and Ion-Related Signaling 2020)

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12 pages, 1357 KiB

Open AccessReview

Regulation of Neutrophil Functions by Hv1/VSOP Voltage-Gated Proton Channels

by Yoshifumi Okochi and Yasushi Okamura

Int. J. Mol. Sci. 2021, 22(5), 2620; https://doi.org/10.3390/ijms22052620 - 5 Mar 2021

Cited by 7 | Viewed by 3054

Abstract

The voltage-gated proton channel, Hv1, also termed VSOP, was discovered in 2006. It has long been suggested that proton transport through voltage-gated proton channels regulate reactive oxygen species (ROS) production in phagocytes by counteracting the charge imbalance caused by the activation of NADPH [...] Read more.

The voltage-gated proton channel, Hv1, also termed VSOP, was discovered in 2006. It has long been suggested that proton transport through voltage-gated proton channels regulate reactive oxygen species (ROS) production in phagocytes by counteracting the charge imbalance caused by the activation of NADPH oxidase. Discovery of Hv1/VSOP not only confirmed this process in phagocytes, but also led to the elucidation of novel functions in phagocytes. The compensation of charge by Hv1/VSOP sustains ROS production and is also crucial for promoting Ca²⁺ influx at the plasma membrane. In addition, proton extrusion into neutrophil phagosomes by Hv1/VSOP is necessary to maintain neutral phagosomal pH for the effective killing of bacteria. Contrary to the function of Hv1/VSOP as a positive regulator for ROS generation, it has been revealed that Hv1/VSOP also acts to inhibit ROS production in neutrophils. Hv1/VSOP inhibits hypochlorous acid production by regulating degranulation, leading to reduced inflammation upon fungal infection, and suppresses the activation of extracellular signal-regulated kinase (ERK) signaling by inhibiting ROS production. Thus, Hv1/VSOP is a two-way player regulating ROS production. Here, we review the functions of Hv1/VSOP in neutrophils and discuss future perspectives. Full article

(This article belongs to the Special Issue Ion Channel and Ion-Related Signaling 2020)

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22 pages, 363 KiB

Open AccessReview

Potassium Channels and Their Potential Roles in Substance Use Disorders

by Michael T. McCoy, Subramaniam Jayanthi and Jean Lud Cadet

Int. J. Mol. Sci. 2021, 22(3), 1249; https://doi.org/10.3390/ijms22031249 - 27 Jan 2021

Cited by 15 | Viewed by 5098

Abstract

Substance use disorders (SUDs) are ubiquitous throughout the world. However, much remains to be done to develop pharmacotherapies that are very efficacious because the focus has been mostly on using dopaminergic agents or opioid agonists. Herein we discuss the potential of using potassium [...] Read more.

Substance use disorders (SUDs) are ubiquitous throughout the world. However, much remains to be done to develop pharmacotherapies that are very efficacious because the focus has been mostly on using dopaminergic agents or opioid agonists. Herein we discuss the potential of using potassium channel activators in SUD treatment because evidence has accumulated to support a role of these channels in the effects of rewarding drugs. Potassium channels regulate neuronal action potential via effects on threshold, burst firing, and firing frequency. They are located in brain regions identified as important for the behavioral responses to rewarding drugs. In addition, their expression profiles are influenced by administration of rewarding substances. Genetic studies have also implicated variants in genes that encode potassium channels. Importantly, administration of potassium agonists have been shown to reduce alcohol intake and to augment the behavioral effects of opioid drugs. Potassium channel expression is also increased in animals with reduced intake of methamphetamine. Together, these results support the idea of further investing in studies that focus on elucidating the role of potassium channels as targets for therapeutic interventions against SUDs. Full article

(This article belongs to the Special Issue Ion Channel and Ion-Related Signaling 2020)

21 pages, 5209 KiB

Open AccessReview

The Therapeutic Potential of Neuronal K-Cl Co-Transporter KCC2 in Huntington’s Disease and Its Comorbidities

by Katie Andrews, Sunday Solomon Josiah and Jinwei Zhang

Int. J. Mol. Sci. 2020, 21(23), 9142; https://doi.org/10.3390/ijms21239142 - 30 Nov 2020

Cited by 8 | Viewed by 5822

Abstract

Intracellular chloride levels in the brain are regulated primarily through the opposing effects of two cation-chloride co-transporters (CCCs), namely K⁺-Cl⁻ co-transporter-2 (KCC2) and Na⁺-K⁺-Cl⁻ co-transporter-1 (NKCC1). These CCCs are differentially expressed throughout the course of [...] Read more.

Intracellular chloride levels in the brain are regulated primarily through the opposing effects of two cation-chloride co-transporters (CCCs), namely K⁺-Cl⁻ co-transporter-2 (KCC2) and Na⁺-K⁺-Cl⁻ co-transporter-1 (NKCC1). These CCCs are differentially expressed throughout the course of development, thereby determining the excitatory-to-inhibitory γ-aminobutyric acid (GABA) switch. GABAergic excitation (depolarisation) is important in controlling the healthy development of the nervous system; as the brain matures, GABAergic inhibition (hyperpolarisation) prevails. This developmental switch in excitability is important, as uncontrolled regulation of neuronal excitability can have implications for health. Huntington’s disease (HD) is an example of a genetic disorder whereby the expression levels of KCC2 are abnormal due to mutant protein interactions. Although HD is primarily considered a motor disease, many other clinical manifestations exist; these often present in advance of any movement abnormalities. Cognitive change, in addition to sleep disorders, is prevalent in the HD population; the effect of uncontrolled KCC2 function on cognition and sleep has also been explored. Several mechanisms by which KCC2 expression is reduced have been proposed recently, thereby suggesting extensive investigation of KCC2 as a possible therapeutic target for the development of pharmacological compounds that can effectively treat HD co-morbidities. Hence, this review summarizes the role of KCC2 in the healthy and HD brain, and highlights recent advances that attest to KCC2 as a strong research and therapeutic target candidate. Full article

(This article belongs to the Special Issue Ion Channel and Ion-Related Signaling 2020)

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