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1.
Simon Perathoner Jacob M. Daane Ulrike Henrion Guiscard Seebohm Charles W. Higdon Stephen L. Johnson Christiane Nüsslein-Volhard Matthew P. Harris 《PLoS genetics》2014,10(1)
The scaling relationship between the size of an appendage or organ and that of the body as a whole is tightly regulated during animal development. If a structure grows at a different rate than the rest of the body, this process is termed allometric growth. The zebrafish another longfin (alf) mutant shows allometric growth resulting in proportionally enlarged fins and barbels. We took advantage of this mutant to study the regulation of size in vertebrates. Here, we show that alf mutants carry gain-of-function mutations in kcnk5b, a gene encoding a two-pore domain potassium (K+) channel. Electrophysiological analysis in Xenopus oocytes reveals that these mutations cause an increase in K+ conductance of the channel and lead to hyperpolarization of the cell. Further, somatic transgenesis experiments indicate that kcnk5b acts locally within the mesenchyme of fins and barbels to specify appendage size. Finally, we show that the channel requires the ability to conduct K+ ions to increase the size of these structures. Our results provide evidence for a role of bioelectric signaling through K+ channels in the regulation of allometric scaling and coordination of growth in the zebrafish. 相似文献
2.
Vladimir Avdonin Erwin F. Shibata Toshinori Hoshi 《The Journal of general physiology》1997,109(2):169-180
Dihydropyridines (DHPs) are well known for their effects on L-type voltage-dependent Ca2+ channels. However, these drugs also affect other voltage-dependent ion channels, including Shaker K+ channels. We examined the effects of DHPs on the Shaker K+ channels expressed in Xenopus oocytes. Intracellular applications of
DHPs quickly and reversibly induced apparent inactivation in the Shaker K+ mutant channels with disrupted
N- and C-type inactivation. We found that DHPs interact with the open state of the channel as evidenced by the
decreased mean open time. The DHPs effects are voltage-dependent, becoming more effective with hyperpolarization. A model which involves binding of two DHP molecules to the channel is consistent with the results obtained in our experiments. 相似文献
3.
Effects of Light on the Membrane Potential of Protoplasts from Samanea saman Pulvini : Involvement of K Channels and the H -ATPase 总被引:1,自引:0,他引:1
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Rhythmic light-sensitive movements of the leaflets of Samanea saman depend upon ion fluxes across the plasma membrane of extensor and flexor cells in opposing regions of the leaf-movement organ (pulvinus). We have isolated protoplasts from the extensor and flexor regions of S. saman pulvini and have examined the effects of brief 30-second exposures to white, blue, or red light on the relative membrane potential using the fluorescent dye, 3,3′-dipropylthiadicarbocyanine iodide. White and blue light induced transient membrane hyperpolarization of both extensor and flexor protoplasts; red light had no effect. Following white or blue light-induced hyperpolarization, the addition of 200 millimolar K+ resulted in a rapid depolarization of extensor, but not of flexor protoplasts. In contrast, addition of K+ following red light or in darkness resulted in a rapid depolarization of flexor, but not of extensor protoplasts. In both flexor and extensor protoplasts, depolarization was completely inhibited by tetraethylammonium, implicating channel-mediated movement of K+ ions. These results suggest that K+ channels are closed in extensor plasma membranes and open in flexor plasma membranes in darkness and that white and blue light, but not red light, close the channels in flexor plasma membranes and open them in extensor plasma membranes. Vanadate treatment inhibited hyperpolarization in response to blue or white light, but did not affect K+ -induced depolarization. This suggests that white or blue light-induced hyperpolarization results from activation of the H+ -ATPase, but this hyperpolarization is not the sole factor controlling the opening of K+ channels. 相似文献
4.
Jose Luis De La Vega-Beltran Claudia Sánchez-Cárdenas Darío Krapf Enrique O. Hernandez-González Eva Wertheimer Claudia L. Trevi?o Pablo E. Visconti Alberto Darszon 《The Journal of biological chemistry》2012,287(53):44384-44393
Mammalian sperm are unable to fertilize the egg immediately after ejaculation; they acquire this capacity during migration in the female reproductive tract. This maturational process is called capacitation and in mouse sperm it involves a plasma membrane reorganization, extensive changes in the state of protein phosphorylation, increases in intracellular pH (pHi) and Ca2+ ([Ca2+]i), and the appearance of hyperactivated motility. In addition, mouse sperm capacitation is associated with the hyperpolarization of the cell membrane potential. However, the functional role of this process is not known. In this work, to dissect the role of this membrane potential change, hyperpolarization was induced in noncapacitated sperm using either the ENaC inhibitor amiloride, the CFTR agonist genistein or the K+ ionophore valinomycin. In this experimental setting, other capacitation-associated processes such as activation of a cAMP-dependent pathway and the consequent increase in protein tyrosine phosphorylation were not observed. However, hyperpolarization was sufficient to prepare sperm for the acrosome reaction induced either by depolarization with high K+ or by addition of solubilized zona pellucida (sZP). Moreover, K+ and sZP were also able to increase [Ca2+]i in non-capacitated sperm treated with these hyperpolarizing agents but not in untreated cells. On the other hand, in conditions that support capacitation-associated processes blocking hyperpolarization by adding valinomycin and increasing K+ concentrations inhibited the agonist-induced acrosome reaction as well as the increase in [Ca2+]i. Altogether, these results suggest that sperm hyperpolarization by itself is key to enabling mice sperm to undergo the acrosome reaction. 相似文献
5.
The activation of Ca2+-dependent K+ channel by propranolol or by ascorbate-phenazine methosulphate stimulates Na+-dependent transport of α-aminoisobutyric acid. This stimulation arises from a membrane hyperpolarization due to the specific increase of membrane K+ conductance. The same treatment does not modify the Na+-independent uptake of the norbornane amino acid. 相似文献
6.
Summary The fluorescent anionic dye, bisoxonol, and flow cytometry have been used to monitor changes in the membrane potential of rat thymocytes exposed to the B subunit of cholera toxin. The B subunit induced a rapid hyperpolarization, which was due to activation of a Ca2+-sensitive K+ channel. Reduction of extracellular Ca2+ to <1 m by the addition of [ethylenebis(oxyethylenenitrilo)]tetraacetic acid immediately abolished the hyperpolarization caused by the B subunit. Cells treated with quinine and tetraethylammonium lost their ability to respond to the B subunit, whereas 4-aminopyridine did not have any effect. Thus, calcium-sensitive and not voltage-gated K+ channels appeared to be responsible for the hyperpolarization. The results of ion substitution experiments indicated that extracellular Na+ was not essential for changes in membrane potential. Further studies with ouabain, amiloride and furosemide demonstrated that electrogenic Na+/K+ ATPase, Na+/H+ antiporter and Na+/K+/Cl– cotransporter, respectively, were not involved in the hyperpolarization process induced by the B subunit. Thus, crosslinking of several molecules of ganglioside GM1 on the cell surface of rat thymocytes by the pentavalent B subunit of cholera toxin modulated plasma membrane permeability to K+ by triggering the opening of Ca2+-sensitive K+ channels. A role for gangliosides in regulating ion permeability would have important implications for the function of gangliosides in various cellular phenomena. 相似文献
7.
We have investigated the effects of hyperpolarization and depolarization, and the presence of K+ and/or Ca2+, on 22Na+ influx into corn (Zea mays L.) root segments. In freshly excised root tissue which is injured, Na+ influx is unaffected by hyperpolarization with fusicoccin, or depolarization with uncoupler (protonophore), or by addition of K+. However, added Ca2+ suppresses Na+ influx by 60%. In washed tissue which has recovered, Na+ influx is doubled over that of freshly excised tissue, and the influx is increased by fusicoccin and suppressed by uncoupler. This energy-linked component of Na+ influx is completely eliminated by low concentrations of K+, leaving the same level and kind of Na+ influx seen in freshly excised roots. The K+-sensitive energy linkage appears to be by the carrier for active K+ influx. Calcium is equally inhibitory to Na+ influx in washed as in fresh tissue. Other divalent cations are only slightly less effective. Net Na+ uptake was about 25% of 22Na+ influx, but proportionately the response to K+ and Ca2+ was about the same.
The constancy of K+-insensitive Na+ influx under conditions known to hyperpolarize and depolarize suggests that if Na+ transport is by means of a voltage-sensitive channel, the rise or fall of channel resistance must be proportional to the rise or fall in potential difference. The alternative is a passive electroneutral exchange of 22Na+ for endogenous Na+. The data suggest that an inwardly directed Na+ current is largely offset by an efflux current, giving both a small net uptake and isotopic exchange.
相似文献8.
A. Lepple-Wienhues S. Berweck M. Böhmig C.P. Leo B. Meyling C. Garbe M. Wiederholt 《The Journal of membrane biology》1996,151(2):149-157
K+ channels, membrane voltage, and intracellular free Ca2+ are involved in regulating proliferation in a human melanoma cell line (SK MEL 28). Using patch-clamp techniques, we found
an inwardly rectifying K+ channel and a calcium-activated K+ channel. The inwardly rectifying K+ channel was calcium independent, insensitive to charybdotoxin, and carried the major part of the whole-cell current. The
K+ channel blockers quinidine, tetraethylammonium chloride and Ba2+ and elevated extracellular K+ caused a dose-dependent membrane depolarization. This depolarization was correlated to an inhibition of cell proliferation.
Charybdotoxin affected neither membrane voltage nor proliferation. Basic fibroblast growth factor and fetal calf serum induced
a transient peak in intracellular Ca2+ followed by a long-lasting Ca2+ influx. Depolarization by voltage clamp decreased and hyperpolarization increased intracellular Ca2+, illustrating a transmembrane flux of Ca2+ following its electrochemical gradient. We conclude that K+ channel blockers inhibit cell-cycle progression by membrane depolarization. This in turn reduces the driving force for the
influx of Ca2+, a messenger in the mitogenic signal cascade of human melanoma cells.
Received: 9 May 1995/Revised: 30 January 1996 相似文献
9.
10.
Jonathan Berrout Mykola Mamenko Oleg L. Zaika Lihe Chen Wenzheng Zang Oleh Pochynyuk Roger G. O'Neil 《PloS one》2014,9(4)
The Ca2+-activated, maxi-K (BK) K+ channel, with low Ca2+-binding affinity, is expressed in the distal tubule of the nephron and contributes to flow-dependent K+ secretion. In the present study we demonstrate that the Ca2+-activated, SK3 (KCa2.3) K+ channel, with high Ca2+-binding affinity, is also expressed in the mouse kidney (RT-PCR, immunoblots). Immunohistochemical evaluations using tubule specific markers demonstrate significant expression of SK3 in the distal tubule and the entire collecting duct system, including the connecting tubule (CNT) and cortical collecting duct (CCD). In CNT and CCD, main sites for K+ secretion, the highest levels of expression were along the apical (luminal) cell membranes, including for both principal cells (PCs) and intercalated cells (ICs), posturing the channel for Ca2+-dependent K+ secretion. Fluorescent assessment of cell membrane potential in native, split-opened CCD, demonstrated that selective activation of the Ca2+-permeable TRPV4 channel, thereby inducing Ca2+ influx and elevating intracellular Ca2+ levels, activated both the SK3 channel and the BK channel leading to hyperpolarization of the cell membrane. The hyperpolarization response was decreased to a similar extent by either inhibition of SK3 channel with the selective SK antagonist, apamin, or by inhibition of the BK channel with the selective antagonist, iberiotoxin (IbTX). Addition of both inhibitors produced a further depolarization, indicating cooperative effects of the two channels on Vm. It is concluded that SK3 is functionally expressed in the distal nephron and collecting ducts where induction of TRPV4-mediated Ca2+ influx, leading to elevated intracellular Ca2+ levels, activates this high Ca2+-affinity K+ channel. Further, with sites of expression localized to the apical cell membrane, especially in the CNT and CCD, SK3 is poised to be a key pathway for Ca2+-dependent regulation of membrane potential and K+ secretion. 相似文献
11.
The effect of adrenaline on the K+-activated hyperpolarization of frog skeletal muscle fibres was studied. The amplitude of K+-activated hyperpolarization, which was produced when the external K+ concentration was changed from 0 to 2 mM, was markedly increased in the presence of adrenaline. In the presence of ouabain (1 × 10?5 M), which completely and reversibly eliminated the K+-activated hyperpolarization, adrenaline caused no significant changes in both the membrane potential and conductance under the condition where the K+-activated hyperpolarization was supposed to be produced. These results suggested that adrenaline accelerated the electrogenic Na+ pump which produced the K+-activated hyperpolarization. 相似文献
12.
J.R. Clay 《The Journal of membrane biology》1996,153(3):195-201
An increase in extracellular potassium ion concentration, K
o
, significantly slows the potassium channel deactivation rate in squid giant axons, as previously shown. Surprisingly, the
effect does not occur in all preparations which, coupled with the voltage independence of this result in preparations in which
it does occur, suggests that it is mediated at a site outside of the electric field of the channel, and that this site is
accessible to potassium ions in some preparations, but not in others. In other words, the effect does not appear to be related
to occupancy of the channel by potassium ions. This conclusion is supported by a four-barrier, three-binding site model of
single file diffusion through the channel in which one site, at most, is unoccupied by a potassium ion (single-vacancy model).
The model is consistent with current-voltage relations with various levels of K
o
, and, by definition, with multiple occupancy by K+. The model predicts that occupancy of any given site is essentially independent of K
o
(or K
i
). The effects of extracellular Rb+ and Cs+ on gating are strongly voltage dependent, and they were observed in all preparations investigated. Consequently, the mechanism
underlying these results would appear to be different from that which underlies the effect of K+ on gating. In particular, the effect of Rb+ on gating is reduced by strong hyperpolarization, which in the context of the occupancy hypothesis, is consistent with the
voltage dependence of the current-voltage relation in the presence of Rb+. The primary, novel, finding in this study is that the effects of Cs+ are counterintuitive in this regard. Specifically, the slowing of channel deactivation rate by Cs+ is also reduced by hyperpolarization, similar to the Rb+ results, whereas blockade is enhanced, which is seemingly inconsistent with the concept that occupancy of the channel by
Cs+ underlies the effect of this ion on gating. This result is further elucidated by barrier modeling of the current-voltage
relation in the presence of Cs+.
Received: 19 December 1995/Revised: 10 June 1996 相似文献
13.
《Comparative biochemistry and physiology. A, Comparative physiology》1993,104(1):149-156
- 1.1. Potassium-selective channels were analysed in growth cones of cultured leech Retzius cells.
- 2.2. In the cell-attached mode and at physiological bath and pipette solution little channel activity was observed at resting membrane potential. The channel open probability (po) increased with cell depolarization, and the slope conductance of the single K+ channel current was about 60 pS.
- 3.3. With symmetrical high KCl solution on both sides of the excised membrane patch three K+ -selective channels could be discriminated. Two channels exhibited a linear current-voltage relation of about 18 pS and 106 pS, respectively.
- 4.4. The most frequently observed K+ channel showed a non-linear current-voltage relation and po increased with increasing free cytoplasmic Ca2+ and during cell hyperpolarization.
14.
V.M. Baizabal-Aguirre S. Clemens N. Uozumi J.I. Schroeder 《The Journal of membrane biology》1999,167(2):119-125
The Arabidopsis thaliana cDNA, KAT1 encodes a hyperpolarization-activated K+ (K+
in
) channel. In the present study, we identify and characterize dominant negative point mutations that suppress K+
in
channel function. Effects of two mutations located in the H5 region of KAT1, at positions 256 (T256R) and 262 (G262K), were
studied. The co-expression of either T256R or G262K mutants with KAT1 produced an inhibition of K+ currents upon membrane hyperpolarization. The magnitude of this inhibition was dependent upon the molar ratio of cRNA for
wild-type to mutant channel subunits injected. Inhibition of KAT1 currents by the co-expression of T256R or G262K did not
greatly affect the ion selectivity of residual currents for Rb+, Na+, Li+, or Cs+. When T256R or G262K were co-expressed with a different K+ channel, AKT2, an inhibition of the channel currents was also observed. Voltage-dependent Cs+ block experiments with co-expressed wild type, KAT1 and AKT2, channels further indicated that KAT1 and AKT2 formed heteromultimers.
These data show that AKT2 and KAT1 are able to co-assemble and suggest that suppression of channel function can be pursued
in vivo by the expression of the dominant negative K
+
in
channel mutants described here.
Received: 2 July 1998/Revised: 23 October 1998 相似文献
15.
Robert L. Nakamura 《Molecular membrane biology》2013,30(5-7):293-308
Kat1 is a highly selective inward-rectifying K+ channel that opens for extended periods under conditions of extreme hyperpolarization. Over 200 point mutants in the pore region of the Kat1 K+ channel were generated and examined in the yeast Saccharomyces cerevisiae and Xenopus oocytes to assess the effect of the mutations on ion selectivity. Substitutions at the tyrosine of the signature sequence G-Y-G resulted in the most significant alterations in ion selectivity, consistent with its role in the selectivity filter. However, greater than 80% of the mutations throughout the greater pore region also conferred a defect in selectivity demonstrating that the entire pore of Kat1 contributes to the ion selectivity of this channel. Surprisingly, we identified a novel class of mutant channel that conferred enhanced selectivity of K+ over Na+. Mutants of this class frequently displayed sensitivity to the competing ion Cs+. This finding has led us to speculate that the Kat1 channel pore has evolved to balance not only K+/Na+ selectivity, but selectivity over Cs+, and possibly a wide spectrum of potential competing ions. 相似文献
16.
Leon Pape 《生物化学与生物物理学报:生物膜》1982,686(2):225-232
Changes in fluorescence intensity of thiodicarbocyanine, DiS-C3(5), were correlated with direct microelectrode potential measurements in red blood cells from Amphiuma means and applied qualitatively to evaluate the effects of extracellular Ca2+, K+ and pH on the membrane potential of human red cells. Increasing extracellular [Ca2+] from 1.8 to 15 mM causes a K+-dependent hyperpolarization and decrease in fluorescence intensity in Amphiuma red cells. Both the hyperpolarization and fluorescence change disappear when the temperature is raised from 17 to 37°C. No change in fluorescence intensity is observed in human red cells with comparable increase in extracellular Ca2+ in the temperature range 5–37°C. Increasing the extracellular pH, however, causes human red cells to respond to an increase in extracellular Ca2+ with a significant but temporary loss in fluorescence intensity. This effect is blocked by EGTA, quinine or by increasing extracellular [K+], indicating that at elevated extracellular pH, human erythrocytes respond to an increase in extracellular Ca2+ with an opening of K+ channels and associated hyperpolarization of the plasma membrane. 相似文献
17.
《Biochimica et Biophysica Acta (BBA)/Molecular Cell Research》2019,1866(9):1376-1388
Cargo receptors in the endoplasmic reticulum (ER) recognize and help membrane and soluble proteins along the secretory pathway to reach their location and functional site. We characterized physiological properties of Saccharomyces cerevisiae strains lacking the ERV14 gene, which encodes a cargo receptor part of COPII-coated vesicles that cycles between the ER and Golgi membranes. The lack of Erv14 resulted in larger cell volume, plasma-membrane hyperpolarization, and intracellular pH decrease. Cells lacking ERV14 exhibited increased sensitivity to toxic cationic drugs and decreased ability to grow on low K+. We found no change in the localization of plasma membrane H+-ATPase Pma1, Na+, K+-ATPase Ena1 and K+ importer Trk2 or vacuolar K+-Cl− co-transporter Vhc1 in the absence of Erv14. However, Erv14 influenced the targeting of two K+-specific plasma-membrane transport systems, Tok1 (K+ channel) and Trk1 (K+ importer), that were retained in the ER in erv14Δ cells. The lack of Erv14 resulted in growth phenotypes related to a diminished amount of Trk1 and Tok1 proteins. We confirmed that Rb+ whole-cell uptake via Trk1 is not efficient in cells lacking Erv14. ScErv14 helped to target Trk1 homologues from other yeast species to the S. cerevisiae plasma membrane. The direct interaction between Erv14 and Tok1 or Trk1 was confirmed by co-immunoprecipitation and by a mating-based Split Ubiquitin System. In summary, our results identify Tok1 and Trk1 to be new cargoes for Erv14 and show this receptor to be an important player participating in the maintenance of several physiological parameters of yeast cells. 相似文献
18.
Samantha Hover Barnabas King Bradley Hall Eleni-Anna Loundras Hussah Taqi Janet Daly Mark Dallas Chris Peers Esther Schnettler Clive McKimmie Alain Kohl John N. Barr Jamel Mankouri 《The Journal of biological chemistry》2016,291(7):3411-3422
Bunyaviruses are considered to be emerging pathogens facilitated by the segmented nature of their genome that allows reassortment between different species to generate novel viruses with altered pathogenicity. Bunyaviruses are transmitted via a diverse range of arthropod vectors, as well as rodents, and have established a global disease range with massive importance in healthcare, animal welfare, and economics. There are no vaccines or anti-viral therapies available to treat human bunyavirus infections and so development of new anti-viral strategies is urgently required. Bunyamwera virus (BUNV; genus Orthobunyavirus) is the model bunyavirus, sharing aspects of its molecular and cellular biology with all Bunyaviridae family members. Here, we show for the first time that BUNV activates and requires cellular potassium (K+) channels to infect cells. Time of addition assays using K+ channel modulating agents demonstrated that K+ channel function is critical to events shortly after virus entry but prior to viral RNA synthesis/replication. A similar K+ channel dependence was identified for other bunyaviruses namely Schmallenberg virus (Orthobunyavirus) as well as the more distantly related Hazara virus (Nairovirus). Using a rational pharmacological screening regimen, two-pore domain K+ channels (K2P) were identified as the K+ channel family mediating BUNV K+ channel dependence. As several K2P channel modulators are currently in clinical use, our work suggests they may represent a new and safe drug class for the treatment of potentially lethal bunyavirus disease. 相似文献
19.
In leech P neurons the inhibition of the Na+-K+ pump by ouabain or omission of bath K+ leaves the membrane potential unaffected for a prolonged period or even induces a marked membrane hyperpolarization, although
the concentration gradients for K+ and Na+ are attenuated substantially. As shown previously, this stabilization of the membrane potential is caused by an increase
in the K+ conductance of the plasma membrane, which compensates for the reduction of the K+ gradient. The data presented here strongly suggest that the increased K+ conductance is due to Na+-activated K+ (KNa) channels. Specifically, an increase in the cytosolic Na+ concentration ([Na+]i) was paralleled by a membrane hyperpolarization, a decrease in the input resistance (Rin) of the cells, and by the occurrence of an outwardly directed membrane current. The relationship between Rin and [Na+]i followed a simple model in which the Rin decrease was attributed to K+ channels that are activated by the binding of three Na+ ions, with half-maximal activation at [Na+]i between 45 and 70 mM. At maximum channel activation, Rin was reduced by more than 90%, suggesting a significant contribution of the KNa channels to the physiological functioning of the cells, although evidence for such a contribution is still lacking. Injection
experiments showed that the KNa channels in leech P neurons are also activated by Li+. 相似文献
20.
A plant hyperpolarization-activating K+ channel, KAT1, is highly selective for K+ over Na+ and is little affected by external Na+, which is crucial to take up K+ effectively in a Na+-containing environment. It has been shown that a mutation at the location (Thr256) preceding the selectivity signature sequence
dramatically enhanced the sensitivity of the KAT1 channel to external Na+. We report here electrophysiological experiments for the mechanism of action of external Na+ on KAT1 channels. The Thr256 residue was substituted with either glutamine (Q) or glutamate (E). The wild-type channel was
insensitive to external Na+. However, the activity of both mutant channels was significantly depressed by Na+ with apparent dissociation constants of 6.7 mm and 11.3 mm for T256Q and T256E, respectively. The instantaneous current-voltage relationships revealed distinct blocking mechanisms
for these mutants. For T256Q a typical voltage-dependent fast blocking was shown. On the other hand, the blocking for the
T256E mutant was voltage-independent at low Na+ concentrations and became voltage-dependent at higher concentrations. At extreme hyperpolarization the blocking was relieved
significantly. These data strongly suggest that the mutation at the end of the pore helix rearranged the selectivity filter
and allows Na+ to penetrate into the pore.
Received: 16 October 2000/Revised: 20 February 2001 相似文献