Kv2.1 Channel Activation and Inactivation Is Influenced by Physical Interactions of Both Syntaxin 1A and the Syntaxin 1A/Soluble<i>N</i>-Ethylmaleimide-Sensitive Factor-25 (t-SNARE) Complex with the C Terminus of the Channel

Tsuk, Sharon; Michaelevski, Izhak; Bentley, Geoffrey N.; Joho, Rolf H.; Chikvashvili, Dodo; Lotan, Ilana

doi:10.1124/mol.104.005314

Cited by 36 publications

(35 citation statements)

References 33 publications

(48 reference statements)

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“…Previous work had shown that these t-SNAREs can bind to and regulate the gating of Kv2.1. [15][16][17][18] However, since the gating of the delayed rectifier does not change with apoptosis, 2 the neurotoxin results, in concert with our trafficking experiments, are most consistent with the requirement of t-SNARE proteins for the membrane fusion required for insertion of new channels into the cell surface. This conclusion also is in accordance with the observations that overexpression of syntaxin in cell lines inhibits exocytosis 19 and Kv2.1 surface expression.…”

Section: Discussionsupporting

confidence: 79%

Apoptotic surface delivery of K+ channels

et al. 2005

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Apoptosis in cortical neurons requires efflux of cytoplasmic potassium mediated by a surge in Kv2.1 channel activity. Pharmacological blockade or molecular disruption of these channels in neurons prevents apoptotic cell death, while ectopic expression of Kv2.1 channels promotes apoptosis in non-neuronal cells. Here, we use a cysteine-containing mutant of Kv2.1 and a thiol-reactive covalent inhibitor to demonstrate that the increase in K þ current during apoptosis is due to de novo insertion of functional channels into the plasma membrane. Biotinylation experiments confirmed the delivery of additional Kv2.1 protein to the cell surface following an apoptotic stimulus. Finally, expression of botulinum neurotoxins that cleave syntaxin and synaptosome-associated protein of 25 kDa (SNAP-25) blocked upregulation of surface Kv2.1 channels in cortical neurons, suggesting that target soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins support proapoptotic delivery of K þ channels. These data indicate that trafficking of Kv2.1 channels to the plasma membrane causes the apoptotic surge in K þ current.

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Section: Discussionsupporting

confidence: 79%

Apoptotic surface delivery of K+ channels

et al. 2005

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“…35 S]Met/Cys-labeled oocytes were subjected to immunoprecipitation as described previously (Michaelevski et al, 2003;Tsuk et al, 2005). Oocytes were injected with 0.75 ng/oocyte syntaxin, 5 ng/oocyte Kv2.1, or 15 ng/oocyte Kv2.1⌬C1a mRNAs, as indicated in Figure 5c.…”

Section: Methodsmentioning

confidence: 99%

“…4a) but not to the C terminus of Kv1.1 (Fili et al, 2001;Michaelevski et al, 2003;Tsuk et al, 2005). Therefore, we examined whether the Kv2.1 syntaxin-binding peptides could be used to compete for the in vivo association of syntaxin with Kv2.1.…”

Section: Kv21-derived Syntaxin-binding Peptides Inhibit the Facilitamentioning

confidence: 99%

“…Specifically, Kv2.1 channels interact physically with syntaxin and SNAP25 in PC12 cells, oocytes, and in vitro (MacDonald et al, 2002;Leung et al, 2003;Michaelevski et al, 2003), and these interactions modulate Kv2.1 currents in HEK 293 cells, islet ␤-cells (MacDonald et al, 2002;Leung et al, 2003), and oocytes (Michaelevski et al, 2003;Tsuk et al, 2005). The Kv2.1 channel is commonly expressed in the soma and dendrites of neurons (Hwang et al, 1993;Trimmer, 1993;Rhodes et al, 1995;Du et al, 1998;Murakoshi and Trimmer, 1999), where it could influence the release of neuropeptides and neurotrophins, and in neuroendocrine cells (MacDonald et al, 2001;Yan et al, 2004;Wolf-Goldberg et al, 2006), where it is well positioned to regulate hormone release.…”

Section: Introductionmentioning

confidence: 99%

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K⁺Channel Facilitation of Exocytosis by Dynamic Interaction with Syntaxin

Singer-Lahat¹,

Sheinin²,

Chikvashvili³

et al. 2007

J. Neurosci.

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Kv channels inhibit release indirectly by hyperpolarizing membrane potential, but the significance of Kv channel interaction with the secretory apparatus is not known. The Kv2.1 channel is commonly expressed in the soma and dendrites of neurons, where it could influence the release of neuropeptides and neurotrophins, and in neuroendocrine cells, where it could influence hormone release. Here we show that Kv2.1 channels increase dense-core vesicle (DCV)-mediated release after elevation of cytoplasmic Ca 2ϩ . This facilitation occurs even after disruption of pore function and cannot be explained by changes in membrane potential and cytoplasmic Ca 2ϩ . However, triggering release increases channel binding to syntaxin, a secretory apparatus protein. Disrupting this interaction with competing peptides or by deleting the syntaxin association domain of the channel at the C terminus blocks facilitation of release. Thus, direct association of Kv2.1 with syntaxin promotes exocytosis. The dual functioning of the Kv channel to influence release, through its pore to hyperpolarize the membrane potential and through its C-terminal association with syntaxin to directly facilitate release, reinforces the requirements for repetitive firing for exocytosis of DCVs in neuroendocrine cells and in dendrites.

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“…Recently, syntaxin 1A (Syx), a plasma membrane protein component of the exocytotic SNARE complex, shown to interact physically and functionally with different voltage-gated K ϩ channels (Fili et al, 2001;Leung et al, 2003Leung et al, , 2007Michaelevski et al, 2003Michaelevski et al, , 2007Tsuk et al, 2005;Yamakawa et al, 2007;Regev et al, 2009), was suggested to be another blocker of M-channels (Regev et al, 2009). …”

Section: Introductionmentioning

confidence: 99%

Regulation of Neuronal M-Channel Gating in an Isoform-Specific Manner: Functional Interplay between Calmodulin and Syntaxin 1A

Etzioni¹,

Siloni²,

Chikvashvilli³

et al. 2011

J. Neurosci.

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Whereas neuronal M-type Kϩ channels composed of KCNQ2 and KCNQ3 subunits regulate firing properties of neurons, presynaptic KCNQ2 subunits were demonstrated to regulate neurotransmitter release by directly influencing presynaptic function. Two interaction partners of M-channels, syntaxin 1A and calmodulin, are known to act presynaptically, syntaxin serving as a major protein component of the membrane fusion machinery and calmodulin serving as regulator of several processes related to neurotransmitter release. Notably, both partners specifically modulate KCNQ2 but not KCNQ3 subunits, suggesting selective presynaptic targeting to directly regulate exocytosis without interference in neuronal firing properties. Here, having first demonstrated in Xenopus oocytes, using analysis of single-channel biophysics, that both modulators downregulate the open probability of KCNQ2 but not KCNQ3 homomers, we sought to resolve the channel structural determinants that confer the isoform-specific gating downregulation and to get insights into the molecular events underlying this mechanism. We show, using optical, biochemical, electrophysiological, and molecular biology analyses, the existence of constitutive interactions between the N and C termini in homomeric KCNQ2 and KCNQ3 channels in living cells. Furthermore, rearrangement in the relative orientation of the KCNQ2 termini that accompanies reduction in single-channel open probability is induced by both regulators, strongly suggesting that closer N-C termini proximity underlies gating downregulation. Different structural determinants, identified at the N and C termini of KCNQ3, prevent the effects by syntaxin 1A and calmodulin, respectively. Moreover, we show that the syntaxin 1A and calmodulin effects can be additive or blocked at different concentration ranges of calmodulin, bearing physiological significance with regard to presynaptic exocytosis.

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Kv2.1 Channel Activation and Inactivation Is Influenced by Physical Interactions of Both Syntaxin 1A and the Syntaxin 1A/SolubleN-Ethylmaleimide-Sensitive Factor-25 (t-SNARE) Complex with the C Terminus of the Channel

Cited by 36 publications

References 33 publications

Apoptotic surface delivery of K+ channels

Apoptotic surface delivery of K+ channels

K⁺Channel Facilitation of Exocytosis by Dynamic Interaction with Syntaxin

Regulation of Neuronal M-Channel Gating in an Isoform-Specific Manner: Functional Interplay between Calmodulin and Syntaxin 1A

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Kv2.1 Channel Activation and Inactivation Is Influenced by Physical Interactions of Both Syntaxin 1A and the Syntaxin 1A/SolubleN-Ethylmaleimide-Sensitive Factor-25 (t-SNARE) Complex with the C Terminus of the Channel

Cited by 36 publications

References 33 publications

Apoptotic surface delivery of K+ channels

Apoptotic surface delivery of K+ channels

K+Channel Facilitation of Exocytosis by Dynamic Interaction with Syntaxin

Regulation of Neuronal M-Channel Gating in an Isoform-Specific Manner: Functional Interplay between Calmodulin and Syntaxin 1A

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K⁺Channel Facilitation of Exocytosis by Dynamic Interaction with Syntaxin