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CONTRIBUTORS
HelenaM.Abelaira
LaboratoryofNeurosciences,GraduatePrograminHealthSciences,HealthSciencesUnit,
UniversityofSouthernSantaCatarina,Criciuma,SantaCatarina,Brazil
RashmiK.Ambasta
MolecularNeuroscienceandFunctionalGenomicsLaboratory,DelhiTechnological
University(FormerlyDCE),Delhi,India
AdelaBanciu
DepartmentofAnatomy,AnimalPhysiologyandBiophysics,FacultyofBiology,University
ofBucharest,Bucharest,Romania
DanielDumitruBanciu
DepartmentofAnatomy,AnimalPhysiologyandBiophysics,FacultyofBiology,University
ofBucharest,Bucharest,Romania
XuChen
CollegeofLifeSciences,ShaanxiNormalUniversity,Xi’an,Shaanxi,PRChina
JinkeCheng
DepartmentofBiochemistryandMolecularCellBiology,ShanghaiJiaoTongUniversity
SchoolofMedicine,Shanghai,PRChina
ChantelleFourie
DepartmentofPhysiology,CentreforBrainResearch,UniversityofAuckland,Auckland,
NewZealand
RomanV.Frolov
DivisionofBiophysics,DepartmentofPhysics,UniversityofOulu,OulunYliopisto,
Finland
Yan-LinFu
DepartmentofPhysiologyandBiophysics,CaseWesternReserveUniversitySchoolof
Medicine,Cleveland,Ohio,USA
LucyGoodman
DepartmentofPhysiology,CentreforBrainResearch,UniversityofAuckland,Auckland,
NewZealand
ZuleideM.Igna´cio
LaboratoryofNeurosciences,GraduatePrograminHealthSciences,HealthSciencesUnit,
UniversityofSouthernSantaCatarina,Criciuma,SantaCatarina,Brazil
NirajKumarJha
MolecularNeuroscienceandFunctionalGenomicsLaboratory,DelhiTechnological
University(FormerlyDCE),Delhi,India
ix
x Contributors
SaurabhKumarJha
MolecularNeuroscienceandFunctionalGenomicsLaboratory,DelhiTechnological
University(FormerlyDCE),Delhi,India
DhirajKumar
MolecularNeuroscienceandFunctionalGenomicsLaboratory,DelhiTechnological
University(FormerlyDCE),Delhi,India
PravirKumar
MolecularNeuroscienceandFunctionalGenomicsLaboratory,DelhiTechnological
University(FormerlyDCE),Delhi,India,andDepartmentofNeurology,Adjunctfaculty,
TuftsUniversitySchoolofMedicine,Boston,Massachusetts,USA
KevinLee
DepartmentofPhysiology,CentreforBrainResearch,UniversityofAuckland,Auckland,
NewZealand
BeulahLeitch
DepartmentofAnatomy,UniversityofOtago,Dunedin,NewZealand
JohannaM.Montgomery
DepartmentofPhysiology,CentreforBrainResearch,UniversityofAuckland,Auckland,
NewZealand
Ting-WeiMu
DepartmentofPhysiologyandBiophysics,CaseWesternReserveUniversitySchoolof
Medicine,Cleveland,Ohio,USA
YitaoQi
CollegeofLifeSciences,ShaanxiNormalUniversity,Xi’an,Shaanxi,PRChina
Joa˜oQuevedo
LaboratoryofNeurosciences,GraduatePrograminHealthSciences,HealthSciencesUnit,
UniversityofSouthernSantaCatarina,Criciuma,SantaCatarina,Brazil;Centerfor
TranslationalPsychiatry;CenterofExcellenceonMoodDisorders,Departmentof
PsychiatryandBehavioralSciences,MedicalSchool,andNeuroscienceGraduateProgram,
GraduateSchoolofBiomedicalSciences,TheUniversityofTexasHealthScienceCenterat
Houston,Houston,Texas,USA
BeatriceMihaelaRadu
DepartmentofNeurologicalandMovementSciences,SectionofAnatomyandHistology,
UniversityofVerona,Verona,Italy,andDepartmentofAnatomy,AnimalPhysiologyand
Biophysics,FacultyofBiology,UniversityofBucharest,Bucharest,Romania
MihaiRadu
DepartmentofNeurologicalandMovementSciences,SectionofAnatomyandHistology,
UniversityofVerona,Verona,Italy,andDepartmentofLifeandEnvironmentalPhysics,
‘HoriaHulubei’NationalInstituteforPhysicsandNuclearEngineering,Magurele,Romania
GislaineZ.R(cid:1)eus
LaboratoryofNeurosciences,GraduatePrograminHealthSciences,HealthSciencesUnit,
UniversityofSouthernSantaCatarina,Criciuma,SantaCatarina,Brazil
Contributors xi
AnaLu´ciaS.Rodrigues
LaboratoryofNeurobiologyofDepression,DepartmentofBiochemistry,Centerof
BiologicalSciences,FederalUniversityofSantaCatarina,Floriano´polis,SantaCatarina,
Brazil
SusanSchenk
SchoolofPsychology,VictoriaUniversity,Wellington,NewZealand
GeraldSeifert
InstituteofCellularNeurosciences,MedicalFaculty,UniversityofBonn,Bonn,Germany
ChristianSteinha¨user
InstituteofCellularNeurosciences,MedicalFaculty,UniversityofBonn,Bonn,Germany
StephanieE.Titus
CenterforTranslationalPsychiatry,DepartmentofPsychiatryandBehavioralSciences,
MedicalSchool,TheUniversityofTexasHealthScienceCenteratHouston,Houston,
Texas,USA
TalitaTuon
LaboratoryofNeurosciences,GraduatePrograminHealthSciences,HealthSciencesUnit,
UniversityofSouthernSantaCatarina,Criciuma,SantaCatarina,Brazil
Ya-JuanWang
CenterforProteomicsandBioinformaticsandDepartmentofEpidemiologyand
Biostatistics,CaseWesternReserveUniversitySchoolofMedicine,Cleveland,Ohio,USA
MattiWeckstro€mw
DivisionofBiophysics,DepartmentofPhysics,UniversityofOulu,OulunYliopisto,
Finland
JohannesWeller
InstituteofCellularNeurosciences,MedicalFaculty,UniversityofBonn,Bonn,Germany
HongmeiWu
CollegeofLifeSciences,ShaanxiNormalUniversity,Xi’an,Shaanxi,PRChina
w MattiWeckstro€mhasdied.
PREFACE
Ion channels are pore-forming membrane proteins expressed in almost all
cell types. These proteins trigger electrical signaling throughout the body
bygatingtheflowofionsacrossthecellmembrane.Twocharacteristicfea-
tures of ion channels distinguish them from other types of ion transporter
proteins.First,thisistheveryhighrateofiontransportthroughthechannel
comparedtoothertransporterproteins(often106ionspersecondorgreater)
andsecond,ionspassthroughchannelsdowntheirelectrochemicalgradient
without the participation of metabolic energy.
The sequencing of the human genome has identified more than 400
putativeionchannels.However,onlyafractionofthesetheoreticallyiden-
tifiedchannelshavebeenclonedandfunctionallycharacterized.Thewide-
spread tissue distribution of ion channels, along with the multiple
physiological consequences of their opening and closing, makes targeting
of ion channels very promising targets for development of therapeutics.
The potential validation of ion channels as drug targets provides an enor-
mous market opportunity for their reemergence as key targets in drug dis-
covery. However, to realize the great potential of this target class, an
understanding of the validation of these targets as well as development of
suitable screening technologies that reflect the complexity of ion channel
structure and function remains key drivers for exploitation of this
opportunity.
Inspiteofsomeimportantdrugstargetingionchannelswhicharetoday
inclinicaluse,asaclass,ionchannelsremainunderexploitedindrugdiscov-
ery. Furthermore, many existing drugsare poorly selective with significant
toxicities or suboptimal efficacy. This thematic volume of the Advances in
Protein Chemistry and Structural Biology is dedicated to ion channels as ther-
apeutic targets and more specifically as promising treatment targets in neu-
rological and psychiatric disorders. Chapter 1 in this volume summarizes
current advances about the protein biogenesis process of the Cys-loop
receptors. Operating on individual biogenesis steps influences the receptor
cell surface level; thus, manipulating the proteostasis network components
canregulatethefunctionofthereceptors,representinganemergingthera-
peutic strategy for corresponding channelopathies. Chapter 2 proposes for
the first time a novel conceptual framework binding together transient
receptor potential (TRP) channels, voltage-gated sodium channels (Nav),
xiii
xiv Preface
and voltage-gated calcium channels (Cav). Authors propose a “flow-
excitation model” that takes into account the inputs mediated by TRP
andothersimilarchannels,theoutputsinvariablyprovidedbyCavchannels,
and the regenerative transmission of signals in the neural networks, for
which Nav channels are responsible. This framework is used to examine
the function, structure, and pharmacology of these channel classes both at
cellular and whole-body physiological level. Building on that basis, the
pathologies arising from the direct or indirect malfunction of the channels
are discussed. The numerous pharmacological interventions affecting these
channelsarealsodescribed.Partofthosearewell-establishedtreatments,like
treatment of hypertension or some forms of epilepsy, but many others are
deeply problematic due to poor drug specificity, ion channel diversity,
andwidespreadexpressionofthechannelsintissuesotherthanthoseactually
targeted.
Chapter3reviewsthepotentialroleofionchannelsinmembranephys-
iologyandbrainhomeostasiswhereionchannelsandtheirassociatedfactors
havebeencharacterizedwiththeirfunctionalconsequencesinneurological
diseases.Furthermore,mechanisticroleofperturbedionchannelsidentified
in various neurodegenerative disorders is discussed. Finally, ion channel
modulators have been investigated for their therapeutic intervention in
treating common neurodegenerative disorders. Chapter 4 is dedicated to
acid-sensingionchannels(ASICs)whichareimportantpharmacologicaltar-
gets being involved in a variety of pathophysiological processes affecting
both the peripheral nervous system (e.g., peripheral pain, diabetic neurop-
athy) and the central nervous system (e.g., stroke, epilepsy, migraine, anx-
iety,fear,depression,neurodegenerativediseases).Thisreviewdiscussesthe
roleplayedbyASICsindifferentpathologiesandthepharmacologicalagents
actingonASICsthatmightrepresentpromisingdrugs.Perspectivesandlim-
itations in the use of ASICs antagonists and modulators as pharmaceutical
agents are also discussed.
Chapter 5 focuses on the glutamatergic system and its associated recep-
torsthatareimplicatedinthepathophysiologyofmajordepressivedisorder.
The N-methyl-D-aspartate (NMDA), a glutamate receptor, is a binding
and/ormodulationsiteforbothclassicalantidepressantsandnewfast-acting
antidepressants.Thus,thisreviewpresentsevidencesdescribingtheeffectof
antidepressants that modulate NMDA receptors and the mechanisms that
contributetotheantidepressantresponse.Chapter6continuesontheglut-
amatergic system. Glutamate is the major neurotransmitter that mediates
Preface xv
excitatory synaptic transmission in the brain through activation of
alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionate(AMPA)recep-
tors.Thesereceptorshavethereforebeenidentifiedasatargetforthedevel-
opment of therapeutic treatments for neurological disorders including
epilepsy, neurodegenerative diseases, autism, and drug addiction. Their
therapeuticpotentialhassincedeclinedduetoinconsistentresultsinclinical
trials. However, recent advances in basic biomedical research significantly
contribute to our knowledge of AMPA receptor structure, binding sites,
and interactions with auxiliary proteins. In particular, the large complex
ofpostsynapticproteinsthatinteractwithAMPAreceptorsubunitshasbeen
shown to control AMPA receptor insertion, location, pharmacology, syn-
aptic transmission, and plasticity. Thus, these proteins are now being con-
sideredasalternativetherapeutictargetsitesformodulatingAMPAreceptors
in neurological disorders.
Chapter7isanexperimentalexampleoftheroleoftheintercellulargap
junction inwardly rectifying K+ (Kir) channels and two-pore domain K+
(K P)channelsinbrainhomeostasismaintainedbyastrocytes.Authorscom-
2
bined functional and molecular analyses to clarify how low pH affects K+
channel function in astrocytes freshly isolated from the developing mouse
hippocampus. No evidence has been found for the presence of ASIC and
transient receptor potential vanilloid receptors in hippocampal astrocytes.
However, the assembly of astrocytic K+ channels allows tolerating short,
transientacidification,andglialKir4.1andK Pchannelscanbeconsidered
2
promisingnewtargetsinbraindiseasesaccompaniedbypHshifts.Chapter8
in this volume discusses the ion channels modification by small ubiquitin-
like modifier (SUMO) proteins and their role in neurological channel-
opathies, especially the determinants of the channels’ regulation. SUMO
proteinscovalentlyconjugatelysineresiduesinalargenumberoftargetpro-
teinsandmodifytheirfunctions.SUMOmodification(SUMOylation)has
emerged as an important regulatory mechanism for protein stability, func-
tion, subcellular localization, and protein–protein interactions. It is until
recently that the physiological impacts of SUMOylation on the regulation
ofneuronalK+channelshavebeeninvestigated.Itisnowclearthatthision
channel modification is a key determinant in the function of K+ channels,
andSUMOylationisimplicatedinawiderangeofchannelopathies,includ-
ing epilepsy and sudden death.
Nonetheless, ion channels remain a relatively underexploited family of
proteinsfortherapeuticinterventions.Anumberofrecentadvancesinboth
xvi Preface
technologyandbiomedicalknowledgesuggestthattheseproteinsareprom-
ising targetsfor futuretherapeutic development.Therefore,the aimofthis
volumeistopromotefurtherresearchinthestructure,function,andregu-
lation of different families of ion channels which would result in designing
new efficient targeted drugs with significantly fewer adverse effects.
DR. ROSSEN DONEV
Biomed Consult Ltd
United Kingdom
CHAPTER ONE
Proteostasis Maintenance of
Cys-Loop Receptors
Yan-Lin Fu*, Ya-Juan Wang†, Ting-Wei Mu*,1
*DepartmentofPhysiologyandBiophysics,CaseWesternReserveUniversitySchoolofMedicine,Cleveland,
Ohio,USA
†CenterforProteomicsandBioinformaticsandDepartmentofEpidemiologyandBiostatistics,CaseWestern
ReserveUniversitySchoolofMedicine,Cleveland,Ohio,USA
1Correspondingauthor:e-mailaddress:[email protected]
Contents
1. Introduction 2
2. Folding,Assembly,andDegradationofCys-LoopReceptorsintheER 5
2.1 FoldingandAssemblyofCys-LoopReceptors 5
2.2 ERADoftheCys-LoopReceptors 8
3. TraffickingofCys-LoopReceptorsfromERtoGolgiandtoPlasmaMembrane 10
4. ProteinQualityControlofCys-LoopReceptorsonthePlasmaMembrane 11
4.1 Clustering 11
4.2 Endocytosis 12
5. OtherRegulationsofCys-LoopReceptors 13
5.1 LipidInvolvementinTraffickingandClustering 13
5.2 PhosphorylationSignalingintheBiogenesisoftheReceptors 14
6. DiseaseandTherapy 15
References 16
Abstract
The Cys-loop receptors play prominent roles in the nervous system. They include γ-
aminobutyricacidtypeAreceptors,nicotinicacetylcholinereceptors,5-hydroxytrypta-
minetype-3receptors,andglycinereceptors.Proteostasisrepresentsanoptimalstateof
the cellular proteome in normal physiology. The proteostasis network regulates the
folding,assembly,degradation,andtraffickingoftheCys-loopreceptors,ensuringtheir
efficientfunctionalcellsurfaceexpressions.Here,wesummarizecurrentadvancesabout
theproteinbiogenesisprocessoftheCys-loopreceptors.Becauseoperatingonindivid-
ual biogenesis steps influences the receptor cell surface level, manipulating the
proteostasis network components can regulate the function of the receptors, rep-
resentinganemergingtherapeuticstrategyforcorrespondingchannelopathies.
AdvancesinProteinChemistryandStructuralBiology,Volume103 #2016ElsevierInc. 1
ISSN1876-1623 Allrightsreserved.
http://dx.doi.org/10.1016/bs.apcsb.2015.11.002
2 Yan-LinFuetal.
1. INTRODUCTION
The Cys-loop receptors, belonging to ligand-gated channels family,
are activated by neurotransmitters, allowing ion flux through neuronal cell
membrane to maintain the neuronal activity of central nervous system
(CNS; Lester, Dibas, Dahan, Leite, & Dougherty, 2004). They include
γ-aminobutyric acid type A receptors (GABA Rs), nicotinic acetylcholine
A
receptors (nAChRs), 5-hydroxytryptamine type-3 receptors (5-HT Rs),
3
andglycinereceptors(GlyRs).AstheCys-loopreceptorsarecomposedoffive
homomeric or heteromeric subunits, they are also calledpentameric ligand-
gated ion channels. The bacterial GLIC and ELIC and the Caenorhabditis
elegansGluClarealsointhissuperfamily.
TheCys-loopreceptorshaveprominentrolesinthenervoussystem.As
themoststudiedmember,nAChRsarecationchannels,permeabletoNa+,
K+, and Ca2+ upon activation. They are responsible for synaptic transmis-
sionintheCNS,inautonomicganglias,intheadrenalgland,andatneuro-
muscular junctions and other peripheral synapses. The receptors are
involved in diseases such as Alzheimer’s disease (AD), bipolar disease, and
myasthenia gravis. nAChRs located at different locations are composed of
different sets of subunit subtypes. α1, β1, γ, and δ subunits or α1, β1, δ,
and ε subunits form muscle-type nAChRs at a 2:1:1:1 ratio, whereas
α2–α10 and β2–β4 subunits compose the most neuronal-type receptors
with (α4)3(β2)2, (α4)2(β2)3, or (α7)5 subtypes predominantly found in
CNS and α3β4 subtypes in autonomic ganglion and adrenal gland (Gotti
et al., 2009; Hogg, Raggenbass, & Bertrand, 2003; Mazzaferro et al.,
2014; Palma, Bertrand, Binzoni, & Bertrand, 1996; Wu, Cheng, Jiang,
Melcher, & Xu, 2015; Xiao & Kellar, 2004).
5-HT Rs, the only inotropic receptor in serotonin receptor family, are
3
alsocationchannelspermeabletoNa+,K+,andCa2+uponactivation.They
are widely located at postsynaptic sites in hippocampus, cortex, substantia
nigra, and brain stem. They also exist in the presynaptic GABAergic nerve
terminalsintheamygdalaandCA1regionofthehippocampus,presynaptic
glutamatergicsynapses,glialcellmembranesinthemedialnucleusofthesol-
itary tract where they play a major role in regulating the release of neuro-
transmitters such as GABA, dopamine, glutamate (Connolly, 2008). They
areinvolvedinmanyclinicaldiseasessuchasdrugaddiction,cognitivefunc-
tion, schizophrenia, and satiety control. Its antagonists are used to treat
postinfectious irritable bowel syndrome and severe diarrhea-predominant
