Pub Date : 2016-01-26DOI: 10.1080/19336950.2015.1126010
A. Billet, Yanlin Jia, T. Jensen, Yue‐xian Hou, X. Chang, J. Riordan, J. Hanrahan
ABSTRACT The CFTR chloride channel is tightly regulated by phosphorylation at multiple serine residues. Recently it has been proposed that its activity is also regulated by tyrosine kinases, however the tyrosine phosphorylation sites remain to be identified. In this study we examined 2 candidate tyrosine residues near the boundary between the first nucleotide binding domain and the R domain, a region which is important for channel function but devoid of PKA consensus sequences. Mutating tyrosines at positions 625 and 627 dramatically reduced responses to Src or Pyk2 without altering the activation by PKA, suggesting they may contribute to CFTR regulation.
{"title":"Potential sites of CFTR activation by tyrosine kinases","authors":"A. Billet, Yanlin Jia, T. Jensen, Yue‐xian Hou, X. Chang, J. Riordan, J. Hanrahan","doi":"10.1080/19336950.2015.1126010","DOIUrl":"https://doi.org/10.1080/19336950.2015.1126010","url":null,"abstract":"ABSTRACT The CFTR chloride channel is tightly regulated by phosphorylation at multiple serine residues. Recently it has been proposed that its activity is also regulated by tyrosine kinases, however the tyrosine phosphorylation sites remain to be identified. In this study we examined 2 candidate tyrosine residues near the boundary between the first nucleotide binding domain and the R domain, a region which is important for channel function but devoid of PKA consensus sequences. Mutating tyrosines at positions 625 and 627 dramatically reduced responses to Src or Pyk2 without altering the activation by PKA, suggesting they may contribute to CFTR regulation.","PeriodicalId":9750,"journal":{"name":"Channels","volume":null,"pages":null},"PeriodicalIF":3.3,"publicationDate":"2016-01-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"75416155","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2016-01-26DOI: 10.1080/19336950.2015.1121335
M. Vornanen, M. Hassinen
ABSTRACT The zebrafish (Danio rerio) has become a popular model for human cardiac diseases and pharmacology including cardiac arrhythmias and its electrophysiological basis. Notably, the phenotype of zebrafish cardiac action potential is similar to the human cardiac action potential in that both have a long plateau phase. Also the major inward and outward current systems are qualitatively similar in zebrafish and human hearts. However, there are also significant differences in ionic current composition between human and zebrafish hearts, and the molecular basis and pharmacological properties of human and zebrafish cardiac ionic currents differ in several ways. Cardiac ionic currents may be produced by non-orthologous genes in zebrafish and humans, and paralogous gene products of some ion channels are expressed in the zebrafish heart. More research on molecular basis of cardiac ion channels, and regulation and drug sensitivity of the cardiac ionic currents are needed to enable rational use of the zebrafish heart as an electrophysiological model for the human heart.
{"title":"Zebrafish heart as a model for human cardiac electrophysiology","authors":"M. Vornanen, M. Hassinen","doi":"10.1080/19336950.2015.1121335","DOIUrl":"https://doi.org/10.1080/19336950.2015.1121335","url":null,"abstract":"ABSTRACT The zebrafish (Danio rerio) has become a popular model for human cardiac diseases and pharmacology including cardiac arrhythmias and its electrophysiological basis. Notably, the phenotype of zebrafish cardiac action potential is similar to the human cardiac action potential in that both have a long plateau phase. Also the major inward and outward current systems are qualitatively similar in zebrafish and human hearts. However, there are also significant differences in ionic current composition between human and zebrafish hearts, and the molecular basis and pharmacological properties of human and zebrafish cardiac ionic currents differ in several ways. Cardiac ionic currents may be produced by non-orthologous genes in zebrafish and humans, and paralogous gene products of some ion channels are expressed in the zebrafish heart. More research on molecular basis of cardiac ion channels, and regulation and drug sensitivity of the cardiac ionic currents are needed to enable rational use of the zebrafish heart as an electrophysiological model for the human heart.","PeriodicalId":9750,"journal":{"name":"Channels","volume":null,"pages":null},"PeriodicalIF":3.3,"publicationDate":"2016-01-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"89350298","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2016-01-26DOI: 10.1080/19336950.2015.1119631
Delany Torres-Salazar, Aneysis D Gonzalez-Suarez, S. Amara
Excitatory Amino Acid Transporters (EAATs) are integral membrane proteins that subserve multiple functions during neurotransmission. They are secondary-active transporters that catalyze the movement of glutamate molecules along with co-transported ions across the plasma membrane of neurons and glial cells. This function is critical to maintain glutamate homeostasis, to limit the diffusion of glutamate released at synapses, and to prevent excessive increases in extracellular glutamate, which has been associated with several neurodegenerative disorders. In addition, EAATs mediate a sodiumand substratedependent anion selective conductance. This channel activity allows the transporter to serve as a glutamate sensor that regulates cell excitability and may also promote electrogenic glutamate transport by clamping the cell at negative potentials. How these 2 functions exist and communicate within the homotrimeric glutamate transporter structure remains an unanswered question. Substrate transport and anion permeation in EAATs are widely accepted to be thermodynamically uncoupled. However, the requirement for glutamate and/or sodium to activate the channel suggests that transitions involved in anion channel opening and closing (“gating”) may be structurally coupled to conformational changes involved in substrate transport. Although this idea has been entertained by several groups, little direct evidence has been provided for how the 2 processes might be linked. In a recent study published in the Journal of Biological Chemistry, we identified a point mutation that drives the carrier in a substrateand voltage-independent constitutive open channel state, and displays a substantially reduced substrate transport activity. In this mutant, substitution of a highly conserved arginine (Arg-388 in EAAT1) with a negatively charged residue decreased substrate transport to 5% of wild type, whereas the macroscopic anion current amplitude was increased six-fold. In contrast to wild-type carriers, this large anion conductance measured in cells expressing R388D/E persisted at its maximum activity in the absence of sodium and glutamate, as well as throughout the entire voltage range from ¡100 to C60 mV. These observations indicate that the mutants exist in a constitutive open channel state and suggest that some of the conformational changes required for substrate transport are tightly coupled to anion channel gating. Recently, several groups in the field have begun to examine the nature of the conformational states of the protein that facilitate anion permeation. Crystallographic data from Verdon and Boudker captured a stable intermediate conformation of at least one of the protomers, which consists of a small (~3.5 A ) inward movement of the transport domain and was termed an intermediate outward facing state (iOFS). Because in this conformation the authors observed a cavity lined by conserved hydrophobic residues, they hypothesized it may provide the anion permeation pa
{"title":"Transport and channel functions in EAATs: the missing link","authors":"Delany Torres-Salazar, Aneysis D Gonzalez-Suarez, S. Amara","doi":"10.1080/19336950.2015.1119631","DOIUrl":"https://doi.org/10.1080/19336950.2015.1119631","url":null,"abstract":"Excitatory Amino Acid Transporters (EAATs) are integral membrane proteins that subserve multiple functions during neurotransmission. They are secondary-active transporters that catalyze the movement of glutamate molecules along with co-transported ions across the plasma membrane of neurons and glial cells. This function is critical to maintain glutamate homeostasis, to limit the diffusion of glutamate released at synapses, and to prevent excessive increases in extracellular glutamate, which has been associated with several neurodegenerative disorders. In addition, EAATs mediate a sodiumand substratedependent anion selective conductance. This channel activity allows the transporter to serve as a glutamate sensor that regulates cell excitability and may also promote electrogenic glutamate transport by clamping the cell at negative potentials. How these 2 functions exist and communicate within the homotrimeric glutamate transporter structure remains an unanswered question. Substrate transport and anion permeation in EAATs are widely accepted to be thermodynamically uncoupled. However, the requirement for glutamate and/or sodium to activate the channel suggests that transitions involved in anion channel opening and closing (“gating”) may be structurally coupled to conformational changes involved in substrate transport. Although this idea has been entertained by several groups, little direct evidence has been provided for how the 2 processes might be linked. In a recent study published in the Journal of Biological Chemistry, we identified a point mutation that drives the carrier in a substrateand voltage-independent constitutive open channel state, and displays a substantially reduced substrate transport activity. In this mutant, substitution of a highly conserved arginine (Arg-388 in EAAT1) with a negatively charged residue decreased substrate transport to 5% of wild type, whereas the macroscopic anion current amplitude was increased six-fold. In contrast to wild-type carriers, this large anion conductance measured in cells expressing R388D/E persisted at its maximum activity in the absence of sodium and glutamate, as well as throughout the entire voltage range from ¡100 to C60 mV. These observations indicate that the mutants exist in a constitutive open channel state and suggest that some of the conformational changes required for substrate transport are tightly coupled to anion channel gating. Recently, several groups in the field have begun to examine the nature of the conformational states of the protein that facilitate anion permeation. Crystallographic data from Verdon and Boudker captured a stable intermediate conformation of at least one of the protomers, which consists of a small (~3.5 A ) inward movement of the transport domain and was termed an intermediate outward facing state (iOFS). Because in this conformation the authors observed a cavity lined by conserved hydrophobic residues, they hypothesized it may provide the anion permeation pa","PeriodicalId":9750,"journal":{"name":"Channels","volume":null,"pages":null},"PeriodicalIF":3.3,"publicationDate":"2016-01-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"80435045","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2016-01-26DOI: 10.1080/19336950.2015.1125737
Stefan Pfeffer, F. Förster
During their synthesis at the ribosome, many proteins have to be either translocated across or inserted into the endoplasmic reticulum (ER) membrane by the translocon, a multi-subunit complex located in the ER membrane. The universally conserved protein-conducting channel Sec61 forms the functional core of the translocon. Accessory translocon components, most notably the stoichiometric translocon associated protein complex (TRAP) and the near-stoichiometric oligosaccharyl-transferase (OST) complex, complement Sec61 and assist in protein transport and membrane protein integration or facilitate maturation of nascent chains by covalent modifications. Early characterization of Sec61 by conductance measurements indicated that it adopts at least 2 distinct conformational states to enable protein translocation and membrane insertion while preventing extensive ion flux: a moreconductive state when bound to the ribosome and a less-conductive state upon ribosome release. Sec61 is a hetero-trimeric complex, consisting of the central Sec61a subunit and 2 much smaller peripheral subunits, Sec61b and Sec61g. X-ray crystallographic analyses of prokaryotic Sec61 homologs revealed that Sec61a consists of 2 pseudo-symmetrical Nand C-terminal halves, each comprising 5 transmembrane helices, which form the translocation channel. The two domains are connected by a short ‘hinge’ helix allowing a jaw-like motion of the Nand C-terminal halves with respect to each other. Consistent with the early characterization of the protein-conducting channel, Sec61 was found to adopt 2 functionally different conformations: a state with a lateral opening between the 2 Sec61a halves, which allows hydrophobic helices to partition into the lipid bilayer (termed the lateral gate), as well as a laterally closed state (Fig. 1). Recent mechanistic models for the interplay of the ribosome and Sec61 were derived from single particle cryo-EM structures of ribosome-bound, detergent-solubilized Sec61 in distinct functional states. They suggested that ribosome-bound Sec61 is mostly present in a closed state and opens only transiently for integration of a nascent transmembrane helix into the membrane. However, these models were inconsistent with the earlier conductance measurements, which indicated that ribosome binding alone induces conformational changes of the native protein-conducting channel toward a more conductive state. This discrepancy illustrates the need for visualizing the conformation of ribosome-bound Sec61 in a lipid environment and in presence of all other translocon components. Cryo-electron tomography (CET) in combination with subtomogram analysis is an excellent method for studying the structures of large macromolecules in their natural environment. It is particularly attractive for studying membrane-embedded and –associated complexes, because detergent solubilization is not required, avoiding destabilization of the complex during purification. Developments in direct detector te
{"title":"Sec61: A static framework for membrane-protein insertion","authors":"Stefan Pfeffer, F. Förster","doi":"10.1080/19336950.2015.1125737","DOIUrl":"https://doi.org/10.1080/19336950.2015.1125737","url":null,"abstract":"During their synthesis at the ribosome, many proteins have to be either translocated across or inserted into the endoplasmic reticulum (ER) membrane by the translocon, a multi-subunit complex located in the ER membrane. The universally conserved protein-conducting channel Sec61 forms the functional core of the translocon. Accessory translocon components, most notably the stoichiometric translocon associated protein complex (TRAP) and the near-stoichiometric oligosaccharyl-transferase (OST) complex, complement Sec61 and assist in protein transport and membrane protein integration or facilitate maturation of nascent chains by covalent modifications. Early characterization of Sec61 by conductance measurements indicated that it adopts at least 2 distinct conformational states to enable protein translocation and membrane insertion while preventing extensive ion flux: a moreconductive state when bound to the ribosome and a less-conductive state upon ribosome release. Sec61 is a hetero-trimeric complex, consisting of the central Sec61a subunit and 2 much smaller peripheral subunits, Sec61b and Sec61g. X-ray crystallographic analyses of prokaryotic Sec61 homologs revealed that Sec61a consists of 2 pseudo-symmetrical Nand C-terminal halves, each comprising 5 transmembrane helices, which form the translocation channel. The two domains are connected by a short ‘hinge’ helix allowing a jaw-like motion of the Nand C-terminal halves with respect to each other. Consistent with the early characterization of the protein-conducting channel, Sec61 was found to adopt 2 functionally different conformations: a state with a lateral opening between the 2 Sec61a halves, which allows hydrophobic helices to partition into the lipid bilayer (termed the lateral gate), as well as a laterally closed state (Fig. 1). Recent mechanistic models for the interplay of the ribosome and Sec61 were derived from single particle cryo-EM structures of ribosome-bound, detergent-solubilized Sec61 in distinct functional states. They suggested that ribosome-bound Sec61 is mostly present in a closed state and opens only transiently for integration of a nascent transmembrane helix into the membrane. However, these models were inconsistent with the earlier conductance measurements, which indicated that ribosome binding alone induces conformational changes of the native protein-conducting channel toward a more conductive state. This discrepancy illustrates the need for visualizing the conformation of ribosome-bound Sec61 in a lipid environment and in presence of all other translocon components. Cryo-electron tomography (CET) in combination with subtomogram analysis is an excellent method for studying the structures of large macromolecules in their natural environment. It is particularly attractive for studying membrane-embedded and –associated complexes, because detergent solubilization is not required, avoiding destabilization of the complex during purification. Developments in direct detector te","PeriodicalId":9750,"journal":{"name":"Channels","volume":null,"pages":null},"PeriodicalIF":3.3,"publicationDate":"2016-01-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"83759250","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2016-01-26DOI: 10.1080/19336950.2015.1120391
U. Hansen, O. Rauh, I. Schroeder
abstract The calculation of flux equations or current-voltage relationships in reaction kinetic models with a high number of states can be very cumbersome. Here, a recipe based on an arrow scheme is presented, which yields a straightforward access to the minimum form of the flux equations and the occupation probability of the involved states in cyclic and linear reaction schemes. This is extremely simple for cyclic schemes without branches. If branches are involved, the effort of setting up the equations is a little bit higher. However, also here a straightforward recipe making use of so-called reserve factors is provided for implementing the branches into the cyclic scheme, thus enabling also a simple treatment of such cases.
{"title":"A simple recipe for setting up the flux equations of cyclic and linear reaction schemes of ion transport with a high number of states: The arrow scheme","authors":"U. Hansen, O. Rauh, I. Schroeder","doi":"10.1080/19336950.2015.1120391","DOIUrl":"https://doi.org/10.1080/19336950.2015.1120391","url":null,"abstract":"abstract The calculation of flux equations or current-voltage relationships in reaction kinetic models with a high number of states can be very cumbersome. Here, a recipe based on an arrow scheme is presented, which yields a straightforward access to the minimum form of the flux equations and the occupation probability of the involved states in cyclic and linear reaction schemes. This is extremely simple for cyclic schemes without branches. If branches are involved, the effort of setting up the equations is a little bit higher. However, also here a straightforward recipe making use of so-called reserve factors is provided for implementing the branches into the cyclic scheme, thus enabling also a simple treatment of such cases.","PeriodicalId":9750,"journal":{"name":"Channels","volume":null,"pages":null},"PeriodicalIF":3.3,"publicationDate":"2016-01-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"80778012","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2016-01-26DOI: 10.1080/19336950.2015.1125278
M. Awayda
The epithelial NaC Channel (ENaC) is rate limiting to NaC entry across many epithelia. Its activity modifies transepithelial NaC transport and by extension salt and water absorption. It has long been known that the channel’s conductance increases, albeit non-linearly, as the NaC concentration increases. However, this channel is unique in that it is also inhibited by elevating the concentration of the transported ion leading to inhibition of permeability. This effect occurs at 2 different time domains stemming from at least 2 distinct mechanisms. The initial acute time course occurs by interaction of the sodium ion with the channel or channel associated proteins and occurs within seconds. The prolonged time course spans minutes to days. This has been recognized early on and termed feedback inhibition. Feedback inhibition is ubiquitously observed for ENaC in many native and heterologous systems and has been reported to involve the ubiquitin ligase protein Nedd4-2, and protein kinase C. The physiological significance of feedback inhibition has been demonstrated by Palmer and colleagues; however, an exact link to the channel has remained missing. In the 8(5) issue of Channels Patel et al. examined the mechanism of feedback inhibition of ENaC by [NaC]i . They subdivide feedback inhibition into one with a 1–2 hour time course and one with an overnight (>8 h) time course. In the early phase, inhibition was not accompanied by detectable effects on subunit trafficking or plasma membrane density but was dependent on the presence of intact channel subunit C-termini. By utilizing a truncated C-terminus b subunit they demonstrated that the sensitivity to increased [NaC]i was rightward shifted or reduced. This demonstrates for the first time that feedback inhibition may affect individual channel activity or open probability (Po), but interestingly, in a manner that depends on the presence of intact intracellular Ctermini. The broader implications are that Po cannot be simply assigned to a single amino acid but is rather the collective activity of numerous extra, intra and transmembrane domains. The second implication is that prolonged increases of [NaC]i may involve downstream modification or signaling with the intracellular C-termini. The second phase of feedback inhibition was observed at periods >8 h. This longer phase was dependent on internalization of gENaC possibly accompanied by reduced cleavage of this subunit indicating reduced channel activity by reducing membrane protein density and by reducing subunit cleavagea process that leads to channel activation. Their results indicate the importance of the C-termini in this phase and present a continuum of events whereby early feedback inhibition likely occurs by inactivating membrane resident channels, while prolonged inactivation occurs by reduced endogenous and presumably intracellular cleavage of subunits accompanied by enhanced internalization from the plasma membrane. The physiological significance of
{"title":"Brakes and gas-regulation of ENaC by sodium","authors":"M. Awayda","doi":"10.1080/19336950.2015.1125278","DOIUrl":"https://doi.org/10.1080/19336950.2015.1125278","url":null,"abstract":"The epithelial NaC Channel (ENaC) is rate limiting to NaC entry across many epithelia. Its activity modifies transepithelial NaC transport and by extension salt and water absorption. It has long been known that the channel’s conductance increases, albeit non-linearly, as the NaC concentration increases. However, this channel is unique in that it is also inhibited by elevating the concentration of the transported ion leading to inhibition of permeability. This effect occurs at 2 different time domains stemming from at least 2 distinct mechanisms. The initial acute time course occurs by interaction of the sodium ion with the channel or channel associated proteins and occurs within seconds. The prolonged time course spans minutes to days. This has been recognized early on and termed feedback inhibition. Feedback inhibition is ubiquitously observed for ENaC in many native and heterologous systems and has been reported to involve the ubiquitin ligase protein Nedd4-2, and protein kinase C. The physiological significance of feedback inhibition has been demonstrated by Palmer and colleagues; however, an exact link to the channel has remained missing. In the 8(5) issue of Channels Patel et al. examined the mechanism of feedback inhibition of ENaC by [NaC]i . They subdivide feedback inhibition into one with a 1–2 hour time course and one with an overnight (>8 h) time course. In the early phase, inhibition was not accompanied by detectable effects on subunit trafficking or plasma membrane density but was dependent on the presence of intact channel subunit C-termini. By utilizing a truncated C-terminus b subunit they demonstrated that the sensitivity to increased [NaC]i was rightward shifted or reduced. This demonstrates for the first time that feedback inhibition may affect individual channel activity or open probability (Po), but interestingly, in a manner that depends on the presence of intact intracellular Ctermini. The broader implications are that Po cannot be simply assigned to a single amino acid but is rather the collective activity of numerous extra, intra and transmembrane domains. The second implication is that prolonged increases of [NaC]i may involve downstream modification or signaling with the intracellular C-termini. The second phase of feedback inhibition was observed at periods >8 h. This longer phase was dependent on internalization of gENaC possibly accompanied by reduced cleavage of this subunit indicating reduced channel activity by reducing membrane protein density and by reducing subunit cleavagea process that leads to channel activation. Their results indicate the importance of the C-termini in this phase and present a continuum of events whereby early feedback inhibition likely occurs by inactivating membrane resident channels, while prolonged inactivation occurs by reduced endogenous and presumably intracellular cleavage of subunits accompanied by enhanced internalization from the plasma membrane. The physiological significance of","PeriodicalId":9750,"journal":{"name":"Channels","volume":null,"pages":null},"PeriodicalIF":3.3,"publicationDate":"2016-01-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"88358878","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2016-01-26DOI: 10.1080/19336950.2015.1120392
L. Motin, T. Durek, David John Adams
ABSTRACT Nine different voltage-gated sodium channel isoforms are responsible for inducing and propagating action potentials in the mammalian nervous system. The Nav1.7 channel isoform plays an important role in conducting nociceptive signals. Specific mutations of this isoform may impair gating behavior of the channel resulting in several pain syndromes. In addition to channel mutations, similar or opposite changes in gating may be produced by spider and scorpion toxins binding to different parts of the voltage-gated sodium channel. In the present study, we analyzed the effects of the α-scorpion toxin OD1 and 2 synthetic toxin analogs on the gating properties of the Nav1.7 sodium channel. All toxins potently inhibited channel inactivation, however, both toxin analogs showed substantially increased potency by more than one order of magnitude when compared with that of wild-type OD1. The decay phase of the whole-cell Na+ current was substantially slower in the presence of toxins than in their absence. Single-channel recordings in the presence of the toxins revealed that Na+ current inactivation slowed due to prolonged flickering of the channel between open and closed states. Our findings support the voltage-sensor trapping model of α-scorpion toxin action, in which the toxin prevents a conformational change in the domain IV voltage sensor that normally leads to fast channel inactivation.
{"title":"Modulation of human Nav1.7 channel gating by synthetic α-scorpion toxin OD1 and its analogs","authors":"L. Motin, T. Durek, David John Adams","doi":"10.1080/19336950.2015.1120392","DOIUrl":"https://doi.org/10.1080/19336950.2015.1120392","url":null,"abstract":"ABSTRACT Nine different voltage-gated sodium channel isoforms are responsible for inducing and propagating action potentials in the mammalian nervous system. The Nav1.7 channel isoform plays an important role in conducting nociceptive signals. Specific mutations of this isoform may impair gating behavior of the channel resulting in several pain syndromes. In addition to channel mutations, similar or opposite changes in gating may be produced by spider and scorpion toxins binding to different parts of the voltage-gated sodium channel. In the present study, we analyzed the effects of the α-scorpion toxin OD1 and 2 synthetic toxin analogs on the gating properties of the Nav1.7 sodium channel. All toxins potently inhibited channel inactivation, however, both toxin analogs showed substantially increased potency by more than one order of magnitude when compared with that of wild-type OD1. The decay phase of the whole-cell Na+ current was substantially slower in the presence of toxins than in their absence. Single-channel recordings in the presence of the toxins revealed that Na+ current inactivation slowed due to prolonged flickering of the channel between open and closed states. Our findings support the voltage-sensor trapping model of α-scorpion toxin action, in which the toxin prevents a conformational change in the domain IV voltage sensor that normally leads to fast channel inactivation.","PeriodicalId":9750,"journal":{"name":"Channels","volume":null,"pages":null},"PeriodicalIF":3.3,"publicationDate":"2016-01-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"79213401","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2016-01-21DOI: 10.1080/19336950.2016.1141842
L. Islas
Abstract Voltage-gated potassium channels or Kv's are membrane proteins with fundamental physiological roles. They are composed of 2 main functional protein domains, the pore domain, which regulates ion permeation, and the voltage-sensing domain, which is in charge of sensing voltage and undergoing a conformational change that is later transduced into pore opening. The voltage-sensing domain or VSD is a highly conserved structural motif found in all voltage-gated ion channels and can also exist as an independent feature, giving rise to voltage sensitive enzymes and also sustaining proton fluxes in proton-permeable channels. In spite of the structural conservation of VSDs in potassium channels, there are several differences in the details of VSD function found across variants of Kvs. These differences are mainly reflected in variations in the electrostatic energy needed to open different potassium channels. In turn, the differences in detailed VSD functioning among voltage-gated potassium channels might have physiological consequences that have not been explored and which might reflect evolutionary adaptations to the different roles played by Kv channels in cell physiology.
{"title":"Functional diversity of potassium channel voltage-sensing domains","authors":"L. Islas","doi":"10.1080/19336950.2016.1141842","DOIUrl":"https://doi.org/10.1080/19336950.2016.1141842","url":null,"abstract":"Abstract Voltage-gated potassium channels or Kv's are membrane proteins with fundamental physiological roles. They are composed of 2 main functional protein domains, the pore domain, which regulates ion permeation, and the voltage-sensing domain, which is in charge of sensing voltage and undergoing a conformational change that is later transduced into pore opening. The voltage-sensing domain or VSD is a highly conserved structural motif found in all voltage-gated ion channels and can also exist as an independent feature, giving rise to voltage sensitive enzymes and also sustaining proton fluxes in proton-permeable channels. In spite of the structural conservation of VSDs in potassium channels, there are several differences in the details of VSD function found across variants of Kvs. These differences are mainly reflected in variations in the electrostatic energy needed to open different potassium channels. In turn, the differences in detailed VSD functioning among voltage-gated potassium channels might have physiological consequences that have not been explored and which might reflect evolutionary adaptations to the different roles played by Kv channels in cell physiology.","PeriodicalId":9750,"journal":{"name":"Channels","volume":null,"pages":null},"PeriodicalIF":3.3,"publicationDate":"2016-01-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"74690557","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2016-01-13DOI: 10.1080/19336950.2015.1124311
Dong-Il Kim, B. Suh
ABSTRACT Voltage-gated calcium (CaV) channels are responsible for Ca2+ influx in excitable cells. As one of the auxiliary subunits, the CaV β subunit plays a pivotal role in the membrane expression and receptor modulation of CaV channels. In particular, the subcellular localization of the β subunit is critical for determining the biophysical properties of CaV channels. Recently, we showed that the β2e isotype is tethered to the plasma membrane. Such a feature of β2e is due to the reversible electrostatic interaction with anionic membrane phospholipids. Here, we further explored the membrane interaction property of β2e by comparing it with that of myristoylated alanine-rich C kinase substrate (MARCKS). First, the charge neutralization of the inner leaf of the plasma membrane induced the translocation of both β2e and MARCKS to the cytosol, while the transient depletion of poly-phosphoinositides (poly-PIs) by translocatable pseudojanin (PJ) systems induced the cytosolic translocation of β2e but not MARCKS. Second, the activation of protein kinase C (PKC) induced the translocation of MARCKS but not β2e. We also found that after the cytosolic translocation of MARCKS by receptor activation, depletion of poly-PIs slowed the recovery of MARCKS to the plasma membrane. Together, our data demonstrate that both β2e and MARCKS bind to the membrane through electrostatic interaction but with different binding affinity, and thus, they are differentially regulated by enzymatic degradation of membrane PIs.
{"title":"Differential interaction of β2e with phosphoinositides: A comparative study between β2e and MARCKS","authors":"Dong-Il Kim, B. Suh","doi":"10.1080/19336950.2015.1124311","DOIUrl":"https://doi.org/10.1080/19336950.2015.1124311","url":null,"abstract":"ABSTRACT Voltage-gated calcium (CaV) channels are responsible for Ca2+ influx in excitable cells. As one of the auxiliary subunits, the CaV β subunit plays a pivotal role in the membrane expression and receptor modulation of CaV channels. In particular, the subcellular localization of the β subunit is critical for determining the biophysical properties of CaV channels. Recently, we showed that the β2e isotype is tethered to the plasma membrane. Such a feature of β2e is due to the reversible electrostatic interaction with anionic membrane phospholipids. Here, we further explored the membrane interaction property of β2e by comparing it with that of myristoylated alanine-rich C kinase substrate (MARCKS). First, the charge neutralization of the inner leaf of the plasma membrane induced the translocation of both β2e and MARCKS to the cytosol, while the transient depletion of poly-phosphoinositides (poly-PIs) by translocatable pseudojanin (PJ) systems induced the cytosolic translocation of β2e but not MARCKS. Second, the activation of protein kinase C (PKC) induced the translocation of MARCKS but not β2e. We also found that after the cytosolic translocation of MARCKS by receptor activation, depletion of poly-PIs slowed the recovery of MARCKS to the plasma membrane. Together, our data demonstrate that both β2e and MARCKS bind to the membrane through electrostatic interaction but with different binding affinity, and thus, they are differentially regulated by enzymatic degradation of membrane PIs.","PeriodicalId":9750,"journal":{"name":"Channels","volume":null,"pages":null},"PeriodicalIF":3.3,"publicationDate":"2016-01-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"82177488","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2016-01-13DOI: 10.1080/19336950.2016.1140956
A. Iuso, D. Križaj
Anthony Iuso and David Kri zaj Department of Ophthalmology & Visual Sciences, Moran Eye Institute, University of Utah School of Medicine, Salt Lake City, UT, USA; Interdepartmental Program in Neuroscience, University of Utah School of Medicine, Salt Lake City, UT, USA; Department of Neurobiology & Anatomy, University of Utah School of Medicine, Salt Lake City, UT, USA; Department of Bioengineering, University of Utah School of Medicine, Salt Lake City, UT, USA
Anthony Iuso和David Kri zaj美国犹他州盐湖城犹他大学医学院莫兰眼科研究所眼科与视觉科学系;美国犹他大学医学院神经科学跨院系项目，美国犹他州盐湖城;美国犹他大学医学院神经生物与解剖学系，美国犹他州盐湖城;美国犹他州盐湖城犹他大学医学院生物工程系
{"title":"TRPV4-AQP4 interactions ‘turbocharge’ astroglial sensitivity to small osmotic gradients","authors":"A. Iuso, D. Križaj","doi":"10.1080/19336950.2016.1140956","DOIUrl":"https://doi.org/10.1080/19336950.2016.1140956","url":null,"abstract":"Anthony Iuso and David Kri zaj Department of Ophthalmology & Visual Sciences, Moran Eye Institute, University of Utah School of Medicine, Salt Lake City, UT, USA; Interdepartmental Program in Neuroscience, University of Utah School of Medicine, Salt Lake City, UT, USA; Department of Neurobiology & Anatomy, University of Utah School of Medicine, Salt Lake City, UT, USA; Department of Bioengineering, University of Utah School of Medicine, Salt Lake City, UT, USA","PeriodicalId":9750,"journal":{"name":"Channels","volume":null,"pages":null},"PeriodicalIF":3.3,"publicationDate":"2016-01-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"87672594","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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