The Nav1.7 Sodium Channel: from Molecule to Man

Total Page:16

File Type:pdf, Size:1020Kb

The Nav1.7 Sodium Channel: from Molecule to Man REVIEWS The NaV1.7 sodium channel: from molecule to man Sulayman D. Dib-Hajj1,2, Yang Yang1,2, Joel A. Black1,2 and Stephen G. Waxman1,2 Abstract | The voltage-gated sodium channel NaV1.7 is preferentially expressed in peripheral somatic and visceral sensory neurons, olfactory sensory neurons and sympathetic ganglion neurons. NaV1.7 accumulates at nerve fibre endings and amplifies small subthreshold depolarizations, poising it to act as a threshold channel that regulates excitability. Genetic and functional studies have added to the evidence that NaV1.7 is a major contributor to pain signalling in humans, and homology modelling based on crystal structures of ion channels suggests an atomic-level structural basis for the altered gating of mutant NaV1.7 that causes pain. Neuropathic pain Voltage-gated sodium channels are essential for electro- that are not seen when these channels are expressed in 11 Pain resulting from lesions or genesis in excitable cells. Nine pore-forming α‑subunits HEK 293 cells . Methods are now available that allow the diseases of the somatosensory of such channels (referred to as channels hereinafter), expression and functional profiling of sodium channels in system. 1 NaV1.1–NaV1.9, have been identified in mammals . peripheral neurons, which more closely mimic the in vivo These isoforms share a common overall structural environment of such channels12. motif (FIG. 1). They are each composed of a long poly- In humans, gain‑of‑function mutations in SCN9A, neuropathic pain peptide (1,700–2,000 amino acids) that folds into four which encodes NaV1.7, lead to severe , homologous domains (DI–DIV) that are linked by three whereas loss‑of‑function mutations in this gene lead to loops (L1–L3), with each domain having six transmem- an indifference to pain13. Studies involving animal injury brane segments (S1–S6)2. The recent determination models and functional studies of neuronal excitability fol- of the crystal structure of the homotetrameric bacte- lowing expression of human mutant NaV1.7 have provided rial sodium channel3 has provided insights into the mechanistic insights into the role of this channel in the atomic structure of mammalian sodium channels and pathophysiology of pain. Additional studies have linked 14,15 the interactions between the voltage-sensing domain NaV1.7 to other sensory modalities, including olfaction , (VSD; encompassing S1–S4) and the pore module (PM; the afferent limb of the cough reflex16 and acid sensing17. S5–S6) within each of the four homologous domains. In this Review, we discuss functional and modelling Genetic, structural and functional studies have shown studies of NaV1.7 that have yielded new insights into the that Nav1.7 regulates sensory neuron excitability and is structure–function relationship of gating mechanisms in a major contributor to several sensory modalities, and this channel and its contribution to neuronal responses 1Department of Neurology, have established the contribution of this sodium channel under normal and pathological conditions. We also and Center for Neuroscience isoform to human pain disorders (FIG. 1). explore strategies for targeting Na 1.7 in the treatment and Regeneration Research, V Yale University School of The nine sodium channel isoforms display different of pain and, finally, identify unanswered questions 1 Medicine, New Haven, kinetics and voltage-dependent properties . Their differ- regarding the role of NaV1.7 in pain signalling. Connecticut 06510, USA. ential deployment in different types of neurons endows 2Rehabilitation Research these cells with distinct firing properties. Sodium chan- Cellular and subcellular distribution Center, Veterans Affairs Connecticut Healthcare nels associate with multiple protein partners that regulate Three sodium channels — NaV1.7, NaV1.8 and NaV1.9 4–7 System, West Haven, channel trafficking and gating , allowing sodium chan- — are preferentially expressed in peripheral neurons. Connecticut 06516, USA. nel properties to be modulated in a cell-type-specific NaV1.7 expression was first detected in somatosensory Correspondence to S.G.W. manner (for examples, see REFS 8–10), highlighting the and sympathetic ganglion neurons18, and has since e‑mail: stephen.waxman@ need to study these channels within their native neu- been reported in myenteric neurons19, olfactory sen- yale.edu 14,15 16,20 doi:10.1038/nrn3404 ronal background whenever practicable. For example, sory neurons (OSNs) , visceral sensory neurons 21–23 Published online the pathogenic G616R variant of NaV1.7 displays gating and smooth myocytes . NaV1.7 is expressed in both 12 December 2012 abnormalities within dorsal root ganglion (DRG) neurons large and small diameter DRG neurons (FIG. 2), including NATURE REVIEWS | NEUROSCIENCE VOLUME 14 | JANUARY 2013 | 49 © 2013 Macmillan Publishers Limited. All rights reserved REVIEWS a DI DII DIII DIV VSD PM S3 S5 S2 S4 S1 S6 7 3 10 6 7 2 4 8 8 16 2 15 5 17 19 4 11 14 2 18 1 5 6 1213 6 3 5 6 7 9 10 C 4 1 1 L1 3 L2 L3 9 N 1 b VSD 1 Q10R 2 I136V 3 S211P 4 F216S 5 I234T 1 7 6 S241T 7 N395K 8 V400M 9 G616R 10 L823R DI PM DIV 4 3 6 11 I848T 12 G856D* 13 L858H 14 L858F 15 A863P 2 16 16 V872G 17 ΔL955 18 P1308L 19 F1449V 8 6 5 8 2 1 R996C 2 V1298F 3 V1298D 4 V1299F 5 I1461T 15 9 13 14 7 10 6 F1462V 7 T1464I 8 G1607R 9 M1627K 10 A1632E‡ 12 17 19 4 3 D623N 4 I720K 5 I739V 11 1 R185H 2 I228M 2 3 6 M932L/V991L 7 M1532I 10 18 DII DIII 5 1 § 4 R1150W Figure 1 | Domain structure of NaV1.7 and locations of characterized mutations inNature NaV1.7‑related Reviews | Neurosciencepain disorders. a | The sodium channel α‑subunit is a long polypeptide that folds into four homologous domains (DI–DIV), each of which consists of six transmembrane segments (S1–S6). The four domains are joined by three loops (L1‑L3). Within each domain, S1–S4 comprise the voltage-sensing domain (VSD; S4, depicted in green, characteristically contains positively charged arginine and lysine residues), and S5–S6 and their extracellular linker comprise the pore module (PM). The linear schematic of the full-length channel shows the locations of amino acids affected by the gain‑of‑function SCN9A mutations that are linked to inherited erythromelalgia (IEM; red symbols), paroxysmal extreme pain disorder (PEPD; grey symbols), and small-fibre neuropathy (SFN; yellow symbols). b | View of the folded NaV1.7 from the intracellular side of the membrane based on the recently determined crystal structure of a bacterial sodium channel3. The structure shows the central ion-conducting PM and four peripheral VSDs. Conformational changes in the VSDs in response to membrane depolarization are transmitted to the PMs through the S4–S5 linkers (identified by arrows through the helices). Mutations that seem distant from each other on the linear model can in fact be in close proximity to each other in the more biologically relevant folded structure. *The patient with this mutation showed symptoms common to IEM and SFN. ‡The patient carrying this mutation showed symptoms and channel properties common to both IEM and PEPD. §This substitution is encoded by a polymorphism that was present in approximately 30% of ethnically matched Caucasian individuals of European descent in a control population81. Part a is modified, with permission, from REF. 37 © (2007) Elsevier Science. 50 | JANUARY 2013 | VOLUME 14 www.nature.com/reviews/neuro © 2013 Macmillan Publishers Limited. All rights reserved REVIEWS Roles in multiple sensory modalities Box 1 | NaV1.7 contributes most of the sodium current in OSNs Pain. As stated above, NaV1.7 is expressed in both large Na 1.7 is the predominant sodium channel in olfactory sensory neurons (OSNs)14,15. 13 V and small diameter DRG neurons , including 85% of 2+ – Although an elaborate Ca -based and Cl -based signalling amplification system in the functionally identified nociceptors24. These observations, OSN cilia can boost odorant receptor potential133,134, the abundant expression of Na 1.7 V together with its properties as a threshold channel, sug- in these cells14,15 and the ability of Na 1.7 to boost weak depolarizations, support a role V gested that Na 1.7 contributes to pain signalling. The for this sodium channel in the initiation of action potential firing along the peripheral V olfactory neuraxis. Mouse and rat OSNs produce a tetrodotoxin (TTX)-sensitive recent discovery of gain‑of‑function SCN9A mutations 14,15,135 in human pain disorders solidified the status of Na 1.7 as current that is consistent with the predominant expression of NaV1.7 in these V cells. Interestingly, the hyperpolarized activation and inactivation properties of this having a central role in pain signalling13, and its involve- TTX-sensitive current are different from those recorded from NaV1.7 expressed in HEK ment in pathological pain signalling is discussed below. 293 cells33,136 or dorsal root ganglion (DRG) neurons11,35, and those in native rat DRG 137–139 14 neurons . Ahn et al. reported identical sequences of the NaV1.7 cDNA in mouse Olfaction. The initial discovery that global knockout of OSN and DRG neuron samples. Together, these data suggest that post-translational Scn9a in mice is neonatally lethal, probably because the modulation of Na 1.7 or interaction with OSN-specific channel partners may lead to 43 V newborn mice do not feed , and the subsequent discov- altered gating properties of Na 1.7 in OSNs compared with DRG neurons. V ery that humans with homozygous SCN9A-null muta- 44,45 tions are anosmic suggested that NaV1.7 is important in olfaction. Nassar et al.43 noted the absence of milk in 24 –/– functionally identified Aβ-fibres and C‑fibres .
Recommended publications
  • Genetic Analysis of the Shaker Gene Complex of Drosophila Melanogaster
    Copyright 0 1990 by the Genetics Society of America Genetic Analysisof the Shaker Gene Complexof Drosophila melanogaster A. Ferriis,* S. LLamazares,* J. L. de la Pornpa,* M. A. Tanouye? and0. Pongs* *Institute Cajal, CSIC, 28006 Madrid, Spain, +California Instituteof Technology, Pasadena, California 91 125, and 'Ruhr Universitat,Bochum, Federal Republic of Germany Manuscript received May 10, 1989 Accepted for publication March 3, 1990 ABSTRACT The Shaker complex (ShC) spans over 350 kb in the 16F region of the X chromosome. It can be dissected by means of aneuploids into three main sections: the maternal effect (ME), the viable (V) and the haplolethal (HL) regions. The mutational analysis of ShC shows a high density of antimorphic mutations among 12 lethal complementation groupsin addition to 14 viable alleles. The complex is the structural locus of a family of potassium channels as well as a number of functions relevant to the biology of the nervous system. The constituents of ShC seem to be linked by functional relationships in view of the similarity of the phenotypes, antimorphic nature of their mutations and the behavior in transheterozygotes. We discuss the relationship between the genetic organization of ShC and the functional coupling of potassium currents with the other functions encodedin the complex. HAKER was the first behavioral mutant detected do not appear tohave the capability of generating, by S in Drosophila melanogaster (CATSCH1944). It was themselves, gated ionic channels. named after another mutantwith a similar phenotype K+ currents are known to be themost diverse class isolated earlier in Drosophila funebris (LUERS 1936). of ionic currents in terms of kinetics, pharmacology The original phenotype was described as the trem- and sensitivity (HILLE 1984; RUDY 1988).Also, these bling of appendagesin the anesthetized fly.
    [Show full text]
  • Chemical Synthesis, Proper Folding, Nav Channel Selectivity Profile And
    toxins Article Chemical Synthesis, Proper Folding, Nav Channel Selectivity Profile and Analgesic Properties of the Spider Peptide Phlotoxin 1 1, 2,3, 4,5, 1 Sébastien Nicolas y, Claude Zoukimian y, Frank Bosmans y,Jérôme Montnach , Sylvie Diochot 6, Eva Cuypers 5, Stephan De Waard 1,Rémy Béroud 2, Dietrich Mebs 7 , David Craik 8, Didier Boturyn 3 , Michel Lazdunski 6, Jan Tytgat 5 and Michel De Waard 1,2,* 1 Institut du Thorax, Inserm UMR 1087/CNRS UMR 6291, LabEx “Ion Channels, Science & Therapeutics”, F-44007 Nantes, France; [email protected] (S.N.); [email protected] (J.M.); [email protected] (S.D.W.) 2 Smartox Biotechnology, 6 rue des Platanes, F-38120 Saint-Egrève, France; [email protected] (C.Z.); [email protected] (R.B.) 3 Department of Molecular Chemistry, Univ. Grenoble Alpes, CNRS, 570 rue de la chimie, CS 40700, 38000 Grenoble, France; [email protected] 4 Faculty of Medicine and Health Sciences, Department of Basic and Applied Medical Sciences, 9000 Gent, Belgium; [email protected] 5 Toxicology and Pharmacology, University of Leuven, Campus Gasthuisberg, P.O. Box 922, Herestraat 49, 3000 Leuven, Belgium; [email protected] (E.C.); [email protected] (J.T.) 6 Université Côte d’Azur, CNRS UMR7275, Institut de Pharmacologie Moléculaire et Cellulaire, 660 route des lucioles, 6560 Valbonne, France; [email protected] (S.D.); [email protected] (M.L.) 7 Institute of Legal Medicine, University of Frankfurt, Kennedyallee 104, 60488 Frankfurt, Germany; [email protected] 8 Institute for Molecular Bioscience, University of Queensland, Brisbane 4072, Australia; [email protected] * Correspondence: [email protected]; Tel.: +33-228-080-076 Contributed equally to this work.
    [Show full text]
  • Characterization of Functional Residues for Catalysis and Kinetics of Aminoacylhistidine Dipeptidase from Vibrio Alginolyticus
    國 立 交 通 大 學 生物科技研究所 碩士論文 溶藻弧菌胺醯組胺酸雙胜肽酶之生化特性分析 及其功能性胺基酸之研究 Characterization of Functional Residues for Catalysis and Kinetics of Aminoacylhistidine Dipeptidase from Vibrio alginolyticus 研究生: 陳怡親 指導教授: 吳東昆 博士 中華民國九十六年七月 Characterization of Functional Residues for Catalysis and Kinetics of Aminoacylhistidine Dipeptidase from Vibrio alginolyticus 研究生:陳怡親 Student: Yi-Chin Chen 指導教授:吳東昆 博士 Advisor: Dr. Tung-Kung Wu 國 立 交 通 大 學 生物科技研究所 碩士論文 A Thesis Submitted to Department of Biological Science and Technology College of Science National Chiao Tung University in partial Fulfillment of the Requirements for the Degree of Master in Biological Science and Technology July, 2007 Hsinchu, Taiwan, Republic of China 中華民國九十六年七月 溶藻弧菌胺醯組胺酸雙胜肽酶之生化特性分析及其功能性胺基酸 之研究 研究生:陳怡親 指導教授:吳東昆 博士 國立交通大學 生物科技研究所碩士班 摘要 胺醯組胺酸雙胜肽酶(PepD, EC 3.4.13.3)為胜肽酶家族M20 中的一員。胜肽酶家族M20 中的酵素皆屬於金屬雙胜肽酶,而經由研究發現可被應用於抗菌、癌症的臨床治療及神 經傳導物質的調控等方面。過去對於細菌中胺醯組胺酸雙胜肽酶的研究很少,只針對其 序列和部分生化特性進行探討,並無其生理角色或活性區胺基酸相關之研究。本論文將 溶藻弧菌pepD基因殖入pET-28a(+)質體中,表現出N端帶有His-tag之重組蛋白,並利用 Ni-NTA親和層析管柱純化之。純化出的蛋白質對於特定的Xaa-His雙胜肽(例如: L-carnosine)具有水解的能力,但無水解三胜肽之活性。經酵素動力學研究,溶藻弧菌 -1 PepD蛋白對雙胜肽L-carnosine之Km與kcat值分別為 0.36 mM與 8.6 min 。經由序列分析 預測溶藻弧菌PepD蛋白上胺基酸位置His80、Asp82、Asp119、Glu149、Glu150、Asp173 及His461 為活性區胺基酸。其中將所預測影響金屬鍵結之胺基酸Asp119 以及扮演催化 角色之胺基酸Glu149 分別進行飽和定點突變,發現大部分之突變蛋白皆失去或降低原 有之活性。此外,以同屬M20 胜肽酶家族之PepV蛋白結晶結構為模板做出溶藻弧菌PepD 蛋白之同源模擬,顯示出相同之活性區胺基酸。因此,根據本論文實驗結果將首次提出 胺醯組胺酸雙胜肽酶活性區胺基酸之分佈情形與其可能扮演之功能。 I Characterization of Functional Residues for Catalysis and Kinetics of Aminoacylhistidine Dipeptidase from Vibrio alginolyticus Student : Yi-Chin Chen Advisor : Dr. Tung-Kung Wu Institute of Biological Science and Technology National Chiao Tung University Abstract Proteins of the aminoacylase-1/metallopeptidase 20 (Acyl/M20) family were characterized to contain a zinc-binding domain at their active site. Aminoacylhistidine dipeptidase (PepD, EC 3.4.13.3) is a member of peptidase family M20 which catalyzes the cleavage and release of N-terminal amino acid, usually are neutral or hydrophobic residue, from Xaa-His peptide or polypeptide.
    [Show full text]
  • Chapter 25 Mechanisms of Action of Antiepileptic Drugs
    Chapter 25 Mechanisms of action of antiepileptic drugs GRAEME J. SILLS Department of Molecular and Clinical Pharmacology, University of Liverpool _________________________________________________________________________ Introduction The serendipitous discovery of the anticonvulsant properties of phenobarbital in 1912 marked the foundation of the modern pharmacotherapy of epilepsy. The subsequent 70 years saw the introduction of phenytoin, ethosuximide, carbamazepine, sodium valproate and a range of benzodiazepines. Collectively, these compounds have come to be regarded as the ‘established’ antiepileptic drugs (AEDs). A concerted period of development of drugs for epilepsy throughout the 1980s and 1990s has resulted (to date) in 16 new agents being licensed as add-on treatment for difficult-to-control adult and/or paediatric epilepsy, with some becoming available as monotherapy for newly diagnosed patients. Together, these have become known as the ‘modern’ AEDs. Throughout this period of unprecedented drug development, there have also been considerable advances in our understanding of how antiepileptic agents exert their effects at the cellular level. AEDs are neither preventive nor curative and are employed solely as a means of controlling symptoms (i.e. suppression of seizures). Recurrent seizure activity is the manifestation of an intermittent and excessive hyperexcitability of the nervous system and, while the pharmacological minutiae of currently marketed AEDs remain to be completely unravelled, these agents essentially redress the balance between neuronal excitation and inhibition. Three major classes of mechanism are recognised: modulation of voltage-gated ion channels; enhancement of gamma-aminobutyric acid (GABA)-mediated inhibitory neurotransmission; and attenuation of glutamate-mediated excitatory neurotransmission. The principal pharmacological targets of currently available AEDs are highlighted in Table 1 and discussed further below.
    [Show full text]
  • Expression Profiling of Ion Channel Genes Predicts Clinical Outcome in Breast Cancer
    UCSF UC San Francisco Previously Published Works Title Expression profiling of ion channel genes predicts clinical outcome in breast cancer Permalink https://escholarship.org/uc/item/1zq9j4nw Journal Molecular Cancer, 12(1) ISSN 1476-4598 Authors Ko, Jae-Hong Ko, Eun A Gu, Wanjun et al. Publication Date 2013-09-22 DOI http://dx.doi.org/10.1186/1476-4598-12-106 Peer reviewed eScholarship.org Powered by the California Digital Library University of California Ko et al. Molecular Cancer 2013, 12:106 http://www.molecular-cancer.com/content/12/1/106 RESEARCH Open Access Expression profiling of ion channel genes predicts clinical outcome in breast cancer Jae-Hong Ko1, Eun A Ko2, Wanjun Gu3, Inja Lim1, Hyoweon Bang1* and Tong Zhou4,5* Abstract Background: Ion channels play a critical role in a wide variety of biological processes, including the development of human cancer. However, the overall impact of ion channels on tumorigenicity in breast cancer remains controversial. Methods: We conduct microarray meta-analysis on 280 ion channel genes. We identify candidate ion channels that are implicated in breast cancer based on gene expression profiling. We test the relationship between the expression of ion channel genes and p53 mutation status, ER status, and histological tumor grade in the discovery cohort. A molecular signature consisting of ion channel genes (IC30) is identified by Spearman’s rank correlation test conducted between tumor grade and gene expression. A risk scoring system is developed based on IC30. We test the prognostic power of IC30 in the discovery and seven validation cohorts by both Cox proportional hazard regression and log-rank test.
    [Show full text]
  • Current, TTX-Resistant Na؉ Current, and Ca2؉ Current in the Action Potentials of Nociceptive Sensory Neurons
    The Journal of Neuroscience, December 1, 2002, 22(23):10277–10290 Roles of Tetrodotoxin (TTX)-Sensitive Na؉ Current, TTX-Resistant Na؉ Current, and Ca2؉ Current in the Action Potentials of Nociceptive Sensory Neurons Nathaniel T. Blair and Bruce P. Bean Department of Neurobiology, Harvard Medical School, Boston, Massachusetts 20114 Nociceptive sensory neurons are unusual in expressing carry the largest inward charge during the upstroke of the voltage-gated inward currents carried by sodium channels re- nociceptor action potential (ϳ58%), with TTX-sensitive sodium sistant to block by tetrodotoxin (TTX) as well as currents carried channels also contributing significantly (ϳ40%), especially near by conventional TTX-sensitive sodium channels and voltage- threshold, and high voltage-activated calcium currents much dependent calcium channels. To examine how currents carried less (ϳ2%). Action potentials had a prominent shoulder during by each of these helps to shape the action potential in small- the falling phase, characteristic of nociceptive neurons. TTX- diameter dorsal root ganglion cell bodies, we voltage clamped resistant sodium channels did not inactivate completely during cells by using the action potential recorded from each cell as the action potential and carried the majority (58%) of inward the command voltage. Using intracellular solutions of physio- current flowing during the shoulder, with high voltage-activated logical ionic composition, we isolated individual components of calcium current also contributing significantly (39%).
    [Show full text]
  • Mapping Protein Interactions of Sodium Channel Nav1.7 Using 2 Epitope-Tagged Gene Targeted Mice
    bioRxiv preprint doi: https://doi.org/10.1101/118497; this version posted March 20, 2017. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. 1 Mapping protein interactions of sodium channel NaV1.7 using 2 epitope-tagged gene targeted mice 3 Alexandros H. Kanellopoulos1†, Jennifer Koenig1, Honglei Huang2, Martina Pyrski3, 4 Queensta Millet1, Stephane Lolignier1, Toru Morohashi1, Samuel J. Gossage1, Maude 5 Jay1, John Linley1, Georgios Baskozos4, Benedikt Kessler2, James J. Cox1, Frank 6 Zufall3, John N. Wood1* and Jing Zhao1†** 7 1Molecular Nociception Group, WIBR, University College London, Gower Street, 8 London WC1E 6BT, UK. 9 2Mass Spectrometry Laboratory, Target Discovery Institute, University of Oxford, The 10 Old Road Campus, Oxford, OX3 7FZ, UK. 11 3Center for Integrative Physiology and Molecular Medicine, Saarland University, 12 Kirrbergerstrasse, Bldg. 48, 66421 Homburg, Germany. 13 4Division of Bioscience, University College London, Gower Street, London WC1E 6BT, UK.14 15 †These authors contributed equally to directing this work. 16 *Correspondence author. Tel: +44 207 6796 954; E-mail: [email protected] 17 **Corresponding author: Tel: +44 207 6790 959; E-mail: [email protected] 1 bioRxiv preprint doi: https://doi.org/10.1101/118497; this version posted March 20, 2017. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. 18 Abstract 19 The voltage-gated sodium channel NaV1.7 plays a critical role in pain pathways. 20 Besides action potential propagation, NaV1.7 regulates neurotransmitter release, 21 integrates depolarizing inputs over long periods and regulates transcription.
    [Show full text]
  • Allosteric Features of KCNQ1 Gating Revealed by Alanine Scanning Mutagenesis
    View metadata, citation and similar papers at core.ac.uk brought to you by CORE provided by Elsevier - Publisher Connector Biophysical Journal Volume 100 February 2011 885–894 885 Allosteric Features of KCNQ1 Gating Revealed by Alanine Scanning Mutagenesis Li-Juan Ma, Iris Ohmert, and Vitya Vardanyan* Institut fu¨r Neurale Signalverarbeitung, Zentrum fu¨r Molekulare Neurobiologie, Universita¨t Hamburg, Hamburg, Germany ABSTRACT Controlled opening and closing of an ion-selective pathway in response to changes of membrane potential is a fundamental feature of voltage-gated ion channels. In recent decades, various details of this process have been revealed with unprecedented precision based on studies of prototypic potassium channels. Though current scientific efforts are focused more on a thorough description of voltage-sensor movement, much less is known about the similarities and differences of the gating mechanisms among potassium channels. Here, we describe the peculiarities of the KCNQ1 gating process in parallel comparison to Shaker. We applied alanine scanning mutagenesis to the S4-S5 linker and pore region and followed the regular- ities of gating perturbations in KCNQ1. We found a fractional constitutive conductance for wild-type KCNQ1. This component increased significantly in mutants with considerably leftward-shifted steady-state activation curves. In contrast to Shaker,no correlation between V1/2 and Z parameters was observed for the voltage-dependent fraction of KCNQ1. Our experimental find- ings are explained by a simple allosteric gating scheme with voltage-driven and voltage-independent transitions. Allosteric features are discussed in the context of extreme gating adaptability of KCNQ1 upon interaction with KCNE b-subunits.
    [Show full text]
  • Engineering Biosynthetic Excitable Tissues from Unexcitable Cells for Electrophysiological and Cell Therapy Studies
    ARTICLE Received 11 Nov 2010 | Accepted 5 Apr 2011 | Published 10 May 2011 DOI: 10.1038/ncomms1302 Engineering biosynthetic excitable tissues from unexcitable cells for electrophysiological and cell therapy studies Robert D. Kirkton1 & Nenad Bursac1 Patch-clamp recordings in single-cell expression systems have been traditionally used to study the function of ion channels. However, this experimental setting does not enable assessment of tissue-level function such as action potential (AP) conduction. Here we introduce a biosynthetic system that permits studies of both channel activity in single cells and electrical conduction in multicellular networks. We convert unexcitable somatic cells into an autonomous source of electrically excitable and conducting cells by stably expressing only three membrane channels. The specific roles that these expressed channels have on AP shape and conduction are revealed by different pharmacological and pacing protocols. Furthermore, we demonstrate that biosynthetic excitable cells and tissues can repair large conduction defects within primary 2- and 3-dimensional cardiac cell cultures. This approach enables novel studies of ion channel function in a reproducible tissue-level setting and may stimulate the development of new cell-based therapies for excitable tissue repair. 1 Department of Biomedical Engineering, Duke University, Durham, North Carolina 27708, USA. Correspondence and requests for materials should be addressed to N.B. (email: [email protected]). NatURE COMMUNicatiONS | 2:300 | DOI: 10.1038/ncomms1302 | www.nature.com/naturecommunications © 2011 Macmillan Publishers Limited. All rights reserved. ARTICLE NatUre cOMMUNicatiONS | DOI: 10.1038/ncomms1302 ll cells express ion channels in their membranes, but cells a b with a significantly polarized membrane that can undergo e 0 a transient all-or-none membrane depolarization (action A 1 potential, AP) are classified as ‘excitable cells’ .
    [Show full text]
  • A Mycobacterium Avium Subsp. Paratuberculosis Predicted Serine Protease Is Associated with Acid Stress and Intraphagosomal Survival
    University of Nebraska - Lincoln DigitalCommons@University of Nebraska - Lincoln U.S. Department of Agriculture: Agricultural Publications from USDA-ARS / UNL Faculty Research Service, Lincoln, Nebraska 2016 A Mycobacterium avium subsp. paratuberculosis Predicted Serine Protease Is Associated with Acid Stress and Intraphagosomal Survival Abirami Kugadas University of Minnesota, Boston Elise A. Lamont University of Minnesota, Saint Paul John Bannantine USDA ARS National Animal Disease Center, [email protected] Fernanda Miyagaki Shoyama Michigan State University, [email protected] Evan Brenner USDA ARS National Animal Disease Center See next page for additional authors Follow this and additional works at: https://digitalcommons.unl.edu/usdaarsfacpub Part of the Agriculture Commons Kugadas, Abirami; Lamont, Elise A.; Bannantine, John; Shoyama, Fernanda Miyagaki; Brenner, Evan; Janagama, Harish K.; and Sreevatsan, Srinand, "A Mycobacterium avium subsp. paratuberculosis Predicted Serine Protease Is Associated with Acid Stress and Intraphagosomal Survival" (2016). Publications from USDA-ARS / UNL Faculty. 2391. https://digitalcommons.unl.edu/usdaarsfacpub/2391 This Article is brought to you for free and open access by the U.S. Department of Agriculture: Agricultural Research Service, Lincoln, Nebraska at DigitalCommons@University of Nebraska - Lincoln. It has been accepted for inclusion in Publications from USDA-ARS / UNL Faculty by an authorized administrator of DigitalCommons@University of Nebraska - Lincoln. Authors Abirami Kugadas, Elise A. Lamont, John Bannantine, Fernanda Miyagaki Shoyama, Evan Brenner, Harish K. Janagama, and Srinand Sreevatsan This article is available at DigitalCommons@University of Nebraska - Lincoln: https://digitalcommons.unl.edu/ usdaarsfacpub/2391 ORIGINAL RESEARCH published: 22 August 2016 doi: 10.3389/fcimb.2016.00085 A Mycobacterium avium subsp. paratuberculosis Predicted Serine Protease Is Associated with Acid Stress and Intraphagosomal Survival Abirami Kugadas 1†, Elise A.
    [Show full text]
  • Cholesterol Modulates the Recruitment of Kv1.5 Channels from Rab11-Associated Recycling Endosome in Native Atrial Myocytes
    Cholesterol modulates the recruitment of Kv1.5 channels from Rab11-associated recycling endosome in native atrial myocytes Elise Balsea,b, Saïd El-Haoua,b, Gilles Dillaniana,b, Aure´ lien Dauphinc, Jodene Eldstromd, David Fedidad, Alain Coulombea,b, and Ste´ phane N. Hatema,b,1 aInstitut National de la Sante´et de la Recherche Me´dicale, Unite´Mixte de Recherche Scientifique-956, 75013 Paris, France; bUniversite´Pierre et Marie Curie, Paris-6, Unite´Mixte de Recherche Scientifique-956, 75013 Paris, France; cPlate-forme imagerie cellulaire IFR14, 75013 Paris, France; and dDepartment of Anesthesiology, Pharmacology and Therapeutics, University of British Columbia, Vancouver, BC, Canada V6T 1Z3 Edited by Lily Y. Jan, University of California, San Francisco, CA, and approved June 19, 2009 (received for review March 17, 2009) Cholesterol is an important determinant of cardiac electrical proper- important role in the regulation of expression of KCNQ1/KCNE1, ties. However, underlying mechanisms are still poorly understood. pacemaker HCN channels and Kv1.5 channels. This process in- Here, we examine the hypothesis that cholesterol modulates the volves several Rab-GTPases (8–10). Rab-GTPases regulate the turnover of voltage-gated potassium channels based on previous trafficking of vesicles between plasma membrane and intracellular observations showing that depletion of membrane cholesterol in- compartments by regulating sorting, tethering and docking of creases the atrial repolarizing current IKur. Whole-cell currents and trafficking vesicles. Rab4, associated with the early endosome (EE), single-channel activity were recorded in rat adult atrial myocytes mediates the fast recycling process while Rab11, linked to the (AAM) or after transduction with hKv1.5-EGFP.
    [Show full text]
  • Disease-Related Cellular Protein Networks Differentially Affected
    www.nature.com/scientificreports OPEN Disease‑related cellular protein networks diferentially afected under diferent EGFR mutations in lung adenocarcinoma Toshihide Nishimura1,8*, Haruhiko Nakamura1,2,8, Ayako Yachie3,8, Takeshi Hase3,8, Kiyonaga Fujii1,8, Hirotaka Koizumi4, Saeko Naruki4, Masayuki Takagi4, Yukiko Matsuoka3, Naoki Furuya5, Harubumi Kato6,7 & Hisashi Saji2 It is unclear how epidermal growth factor receptor EGFR major driver mutations (L858R or Ex19del) afect downstream molecular networks and pathways. This study aimed to provide information on the infuences of these mutations. The study assessed 36 protein expression profles of lung adenocarcinoma (Ex19del, nine; L858R, nine; no Ex19del/L858R, 18). Weighted gene co-expression network analysis together with analysis of variance-based screening identifed 13 co-expressed modules and their eigen proteins. Pathway enrichment analysis for the Ex19del mutation demonstrated involvement of SUMOylation, epithelial and mesenchymal transition, ERK/mitogen- activated protein kinase signalling via phosphorylation and Hippo signalling. Additionally, analysis for the L858R mutation identifed various pathways related to cancer cell survival and death. With regard to the Ex19del mutation, ROCK, RPS6KA1, ARF1, IL2RA and several ErbB pathways were upregulated, whereas AURK and GSKIP were downregulated. With regard to the L858R mutation, RB1, TSC22D3 and DOCK1 were downregulated, whereas various networks, including VEGFA, were moderately upregulated. In all mutation types, CD80/CD86 (B7), MHC, CIITA and IFGN were activated, whereas CD37 and SAFB were inhibited. Costimulatory immune-checkpoint pathways by B7/CD28 were mainly activated, whereas those by PD-1/PD-L1 were inhibited. Our fndings may help identify potential therapeutic targets and develop therapeutic strategies to improve patient outcomes.
    [Show full text]