Modification of Kv2.1 K+ currents by the silent Kv10 subunits

Miera, E C Vega-Saenz de

doi:10.1016/j.molbrainres.2004.01.004

Cited by 18 publications

(17 citation statements)

References 43 publications

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“…The observed differences in the V 1/2 of inactivation of K v 2 channels recorded in native cells and heterologous expression systems have been well documented and are thought to be due to the presence of additional electrically silent K v subunits in native cells (23,26,41) or differences in the phosphorylation status of the channel (21,22). The silent K v subunits comprise the K v 5, K v 6, K v 8, and K v 9 families, which cannot form functional ion channels when expressed alone, but can modify the biophysical properties when coexpressed with the other pore-forming K v subunits (26,27,30,31,39,41,42). Thus, coexpression of K v 2.1 with the silent subunit K v 9.3 has been shown to slow deactivation, shift the activation V 1/2 by approximately Ϫ20 mV, and alter the inactivation V 1/2 by approximately Ϫ15 mV (26,41).…”

Section: Discussionmentioning

confidence: 99%

Contribution of K_v2.1 channels to the delayed rectifier current in freshly dispersed smooth muscle cells from rabbit urethra

Kyle

Bradley

Ohya

et al. 2011

American Journal of Physiology-Cell Physiology

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We have characterized the native voltage-dependent K(+) (K(v)) current in rabbit urethral smooth muscle cells (RUSMC) and compared its pharmacological and biophysical properties with K(v)2.1 and K(v)2.2 channels cloned from the rabbit urethra and stably expressed in human embryonic kidney (HEK)-293 cells (HEK(Kv2.1) and HEK(Kv2.2)). RUSMC were perfused with Hanks' solution at 37°C and studied using the patch-clamp technique with K(+)-rich pipette solutions. Cells were bathed in 100 nM Penitrem A (Pen A) to block large-conductance Ca(2+)-activated K(+) (BK) currents and depolarized to +40 mV for 500 ms to evoke K(v) currents. These were unaffected by margatoxin, κ-dendrotoxin, or α-dendrotoxin (100 nM, n = 3-5) but were blocked by stromatoxin-1 (ScTx, IC(50) ∼130 nM), consistent with the idea that the currents were carried through K(v)2 channels. RNA was detected for K(v)2.1, K(v)2.2, and the silent subunit K(v)9.3 in urethral smooth muscle. Immunocytochemistry showed membrane staining for both K(v)2 subtypes and K(v)9.3 in isolated RUSMC. HEK(Kv2.1) and HEK(Kv2.2) currents were blocked in a concentration-dependent manner by ScTx, with estimated IC(50) values of ∼150 nM (K(v)2.1, n = 5) and 70 nM (K(v)2.2, n = 6). The mean half-maximal voltage (V(1/2)) of inactivation of the USMC K(v) current was -56 ± 3 mV (n = 9). This was similar to the HEK(Kv2.1) current (-55 ± 3 mV, n = 13) but significantly different from the HEK(Kv2.2) currents (-30 ± 3 mV, n = 11). Action potentials (AP) evoked from RUSMC studied under current-clamp mode were unaffected by ScTx. However, when ScTx was applied in the presence of Pen A, the AP duration was significantly prolonged. Similarly, ScTx increased the amplitude of spontaneous contractions threefold, but only after Pen A application. These data suggest that K(v)2.1 channels contribute significantly to the K(v) current in RUSMC.

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Section: Discussionmentioning

confidence: 99%

Contribution of K_v2.1 channels to the delayed rectifier current in freshly dispersed smooth muscle cells from rabbit urethra

Kyle

Bradley

Ohya

et al. 2011

American Journal of Physiology-Cell Physiology

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“…Reported differences in Kv2 channels upon γ subunit coexpression include decrease in the current amplitude, changes in the voltage dependence of activation and inactivation, in the time course of activation, inactivation and deactivation and in the drug sensitivity of the heteromultimeric channels. In the case of Kv6.3, there are only a few studies (Ottschytsch et al 2002; Sano et al 2002; Vega‐Saenz de Miera, 2004) in which the modifications reported are not fully coincident, and for this reason we have carried out the comparative study (Kv2.1 versus Kv2.1/Kv6.3 currents) in transfected HEK cells. We found a decrease of the current amplitude in heteromeric Kv2.1/Kv6.3 that has been previously reported in one of the above‐mentioned studies (Vega‐Saenz de Miera, 2004) but was not explored in the others.…”

Section: Discussionmentioning

confidence: 99%

“…In the case of Kv6.3, there are only a few studies (Ottschytsch et al 2002; Sano et al 2002; Vega‐Saenz de Miera, 2004) in which the modifications reported are not fully coincident, and for this reason we have carried out the comparative study (Kv2.1 versus Kv2.1/Kv6.3 currents) in transfected HEK cells. We found a decrease of the current amplitude in heteromeric Kv2.1/Kv6.3 that has been previously reported in one of the above‐mentioned studies (Vega‐Saenz de Miera, 2004) but was not explored in the others. Regarding the kinetic changes, we saw a significant slowdown of activation (in agreement with the data of Vega‐Saenz de Miera but opposite to the report of Ottschytsch et al ) and deactivation (reported by Sano et al 2002), but found to be not significant in the work of Ottschytsch et al Finally, we saw a leftward shift in the steady‐state activation curve in the presence of Kv6.3 channels as reported by Ottschytsch et al but not by Sano et al It is likely that the differences in the expression systems used (oocytes, L929 and Ltk cells) as well as in the ratio of Kv2.1: Kv6.3 (Vega‐Saenz de Miera, 2004) contribute to the observed discrepancies.…”

Section: Discussionmentioning

confidence: 99%

De novo expression of Kv6.3 contributes to changes in vascular smooth muscle cell excitability in a hypertensive mice strain

Moreno‐Domínguez

Cidad

Miguel-Velado

et al. 2009

The Journal of Physiology

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Essential hypertension involves a gradual and sustained increase in total peripheral resistance, reflecting an increased vascular tone. This change associates with a depolarization of vascular myocytes, and relies on a change in the expression profile of voltage-dependent ion channels (mainly Ca 2+ and K + channels) that promotes arterial contraction. However, changes in expression and/or modulation of voltage-dependent K + channels (Kv channels) are poorly defined, due to their large molecular diversity and their vascular bed-specific expression. Here we endeavor to characterize the molecular and functional expression of Kv channels in vascular smooth muscle cells (VSMCs) and their regulation in essential hypertension, by using VSMCs from resistance (mesenteric) or conduit (aortic) arteries obtained from a hypertensive inbred mice strain, BPH, and the corresponding normotensive strain, BPN. Real-time PCR reveals a differential distribution of Kv channel subunits in the different vascular beds as well as arterial bed-specific changes under hypertension. In mesenteric arteries, the most conspicuous change was the de novo expression of Kv6.3 (Kcng3) mRNA in hypertensive animals. The functional relevance of this change was studied by using patch-clamp techniques. VSMCs from BPH arteries were more depolarized than BPN ones, and showed significantly larger capacitance values. Moreover, Kv current density in BPH VSMCs is decreased mainly due to the diminished contribution of the Kv2 component. The kinetic and pharmacological profile of Kv2 currents suggests that the expression of Kv6.3 could contribute to the natural development of hypertension.

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“…Most silent subunits affect the Kv2.1 inactivation and current density markedly, whereas the effects on the Kv2.1 activation are more limited (13,(15)(16)(17)(18)(19)(20)(21)(22)(23)(24)(25)(26)(27). This is also the case for Kv6.4: WT Kv6.4 subunits do not affect the activation properties markedly but shift the voltage dependence of inactivation by Ϫ40 mV in a hyperpolarized direction and cause a severe down-regulation of the Kv2.1 current density (13) (in the original nomenclature Kv6.3).…”

Section: Discussionmentioning

confidence: 99%

“…These latter Kv subfamilies are designated silent ␣-subunits because they fail to express functional channels in a homotetrameric configuration (13)(14)(15)(16)(17) and are retained in the endoplasmic reticulum (ER). This retention is rescued by the selective co-assembly with members of the Kv2 subfamily in which the silent Kv subunits modulate the Kv2 currents by reducing the current amplitude, altering the inactivation and deactivation kinetics and shifting the voltage dependence of inactivation toward more hyperpolarized potentials (13,(15)(16)(17)(18)(19)(20)(21)(22)(23)(24)(25)(26)(27).…”

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confidence: 99%

Conserved Negative Charges in the N-terminal Tetramerization Domain Mediate Efficient Assembly of Kv2.1 and Kv2.1/Kv6.4 Channels

Bocksteins

Labro

Mayeur

et al. 2009

Journal of Biological Chemistry

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Voltage-gated potassium (Kv) channels are transmembrane tetramers of individual ␣-subunits. Eight different Shaker-related Kv subfamilies have been identified in which the tetramerization domain T1, located on the intracellular N terminus, facilitates and controls the assembly of both homoand heterotetrameric channels. Only the Kv2 ␣-subunits are able to form heterotetramers with members of the silent Kv subfamilies (Kv5, Kv6, Kv8, and Kv9). The T1 domain contains two subdomains, A and B box, which presumably determine subfamily specificity by preventing incompatible subunits to assemble. In contrast, little is known about the involvement of the A/B linker sequence. Both Kv2 and silent Kv subfamilies contain a fully conserved and negatively charged sequence (CDD) in this linker that is lacking in the other subfamilies. Neutralizing these aspartates in Kv2.1 by mutating them to alanines did not affect the gating properties, but reduced the current density moderately. However, charge reversal arginine substitutions strongly reduced the current density of these homotetrameric mutant Kv2.1 channels and immunocytochemistry confirmed the reduced expression at the plasma membrane. Förster resonance energy transfer measurements using confocal microscopy showed that the latter was not due to impaired trafficking, but to a failure to assemble the tetramer. This was further confirmed with co-immunoprecipitation experiments. The corresponding arginine substitution in Kv6.4 prevented its heterotetrameric interaction with Kv2.1. These results indicate that these aspartates (especially the first one) in the A/B box linker of the T1 domain are required for efficient assembly of both homotetrameric Kv2.1 and heterotetrameric Kv2.1/silent Kv6.4 channels.Shaker-related channel subfamilies (Kv1-Kv6 and Kv8 -Kv9) have been identified based on the degree of sequence homology (1). Fully assembled Kv channels are composed of four ␣-subunits arranged around a central pore. Each ␣-subunit consists of six transmembrane segments S1-S6 with a cytoplasmic N and C terminus. The N terminus contains the T1 domain, a tetramerization domain that facilitates the assembly of ␣-subunits into functional channels. The presence of a T1 domain is not absolutely required for channel assembly because subunits without a T1 domain could also assemble into a functional tetramer, although less efficiently (2-4). However, the T1 domain not only promotes but also restricts the formation of possible homo-and heterotetramers by preventing incompatible subunits from assembling (5-7). When four compatible T1 domains assemble, they are arranged with the same 4-fold symmetry as the transmembrane segments, forming a hanging gondola structure (8).The T1 domain contains two subdomains, designated A and B box, which are both highly conserved within the different Kv subfamilies (9). Several residues in both the A and B boxes form the contact interface between neighboring subunits and are commonly assumed to be the key determinants for (subfamily specific) channel tetramer...

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Modification of Kv2.1 K+ currents by the silent Kv10 subunits

Cited by 18 publications

References 43 publications

Contribution of K_v2.1 channels to the delayed rectifier current in freshly dispersed smooth muscle cells from rabbit urethra

Contribution of K_v2.1 channels to the delayed rectifier current in freshly dispersed smooth muscle cells from rabbit urethra

De novo expression of Kv6.3 contributes to changes in vascular smooth muscle cell excitability in a hypertensive mice strain

Conserved Negative Charges in the N-terminal Tetramerization Domain Mediate Efficient Assembly of Kv2.1 and Kv2.1/Kv6.4 Channels

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Modification of Kv2.1 K+ currents by the silent Kv10 subunits

Cited by 18 publications

References 43 publications

Contribution of Kv2.1 channels to the delayed rectifier current in freshly dispersed smooth muscle cells from rabbit urethra

Contribution of Kv2.1 channels to the delayed rectifier current in freshly dispersed smooth muscle cells from rabbit urethra

De novo expression of Kv6.3 contributes to changes in vascular smooth muscle cell excitability in a hypertensive mice strain

Conserved Negative Charges in the N-terminal Tetramerization Domain Mediate Efficient Assembly of Kv2.1 and Kv2.1/Kv6.4 Channels

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Contribution of K_v2.1 channels to the delayed rectifier current in freshly dispersed smooth muscle cells from rabbit urethra

Contribution of K_v2.1 channels to the delayed rectifier current in freshly dispersed smooth muscle cells from rabbit urethra