The KCNQ1 (Kv7.1) COOH Terminus, a Multitiered Scaffold for Subunit Assembly and Protein Interaction

Wiener, Reuven; Haitin, Yoni; Shamgar, Liora; Fernández‐Alonso, M. Carmen; Martos, Ariadna; Chomsky-Hecht, Orna; Rivas, Germán; Attali, Bernard; Hirsch, Joel A.

doi:10.1074/jbc.m707541200

Cited by 127 publications

(175 citation statements)

References 44 publications

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“…Our results from the experiments on C terminus-and N terminus-truncated VSOP suggest that the presence of the cytoplasmic domains is important for the dimerization of VSOP. In fact, the C terminus sequence exhibits a coiled-coil structure that is known to mediate subunit interactions in many examples of ion channel assembly (16)(17)(18). However, because the C terminusand N terminus-truncated VSOP construct still conduct protons, the dimerization does not seem to be necessary for VSOP to function as a proton channel.…”

Section: Discussionmentioning

confidence: 99%

Multimeric nature of voltage-gated proton channels

Koch

Kurokawa

Okochi

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

185

375

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Voltage-gated potassium channels are comprised of four subunits, and each subunit has a pore domain and a voltage-sensing domain (VSD). The four pore domains assemble to form one single central pore, and the four individual VSDs control the gate of the pore. Recently, a family of voltage-gated proton channels, such as HV or voltage sensor only protein (VSOP), was discovered that contain a single VSD but no pore domain. It has been assumed that VSOP channels are monomeric and contain a single VSD that functions as both the VSD and the pore domain. It remains unclear, however, how a protein that contains only a VSD and no pore domain can conduct ions. Using fluorescence measurements and immunoprecipitation techniques, we show here that VSOP channels are expressed as multimeric channels. Further, FRET experiments on constructs with covalently linked subunits show that VSOP channels are dimers. Truncation of the cytoplasmic regions of VSOP reduced the dimerization, suggesting that the dimerization is caused mainly by cytoplasmic protein-protein interactions. However, these N terminus-and C terminus-deleted channels displayed large proton currents. Therefore, we conclude that even though VSOP channels are expressed mainly as dimers in the cell membrane, single VSOP subunits could function independently as proton channels.dimer ͉ FRET ͉ Hv ͉ voltage sensor ͉ voltage sensor only protein V oltage-gated proton channels (H V channels) have been found in many mammalian cells, including skeletal muscle, lungs, microglia, and blood (1). H V channels have also been shown to play a crucial role in the immune system: H V channels in macrophages are involved in the pathway for the generation of reactive oxygen species, which is critical to the process of phagocytosis and the destruction of foreign pathogens (1). H V channels are activated at depolarized voltages, and their activation can be blocked by Zn 2ϩ (2, 3).Recently, a family of voltage-gated proton channels, called H V or voltage sensor only protein (VSOP) channels, was cloned. VSOP channels were found to have four transmembrane domains, and these domains are homologous to the four transmembrane domains of the voltage-sensing domain (VSD) of voltage-gated potassium channels (2, 3). Voltage-gated potassium channels are comprised of four subunits, each of which has a pore domain and a VSD domain. The four pore domains come together to form one single central pore, and the four individual VSDs control the gate of the pore (4). It is not clear, however, how VSOP channels containing only a VSD and no pore domain can conduct ions. Recently, it was shown that the VSD in Na ϩ and K ϩ channels could function as a proton or cation pore independently of the centrally located pore (5-8). It was therefore suggested that a VSOP channel functions as a single, independent VSD and that this VSD makes up the entire proton channel (1, 9, 10). In the present work, we tested the hypothesis that Hv channels are expressed as monomers. We did this by examining whether HA-tagged mouse VSOP (mVSO...

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Section: Discussionmentioning

confidence: 99%

Multimeric nature of voltage-gated proton channels

Koch

Kurokawa

Okochi

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

185

375

View full text Add to dashboard Cite

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“…It should also be taken into consideration that many tetrameric K ϩ channels are devoid of so called "assembly domain." On the other hand, the isolated C-terminal "assembly domain" of KCNQ forms tetramers spontaneously in vitro (Howard et al, 2007;Wehling et al, 2007;Wiener et al, 2008). It has been suggested that monomers, dimers, and tetramers of AMPAR are in a dynamic equilibrium, and that the pore determines the balance between these intermediates (Greger et al, 2003).…”

Section: Discussionmentioning

confidence: 99%

A Pore Residue of the KCNQ3 Potassium M-Channel Subunit Controls Surface Expression

Gómez-Posada¹,

Etxeberría²,

Roura-Ferrer³

et al. 2010

Journal of Neuroscience

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KCNQ2 (Kv7.2) and KCNQ3 (Kv7.3) are the principal subunits underlying the potassium M-current, which exerts a strong control on neuronal excitability. KCNQ3 subunits coassemble with KCNQ2 to form functional heteromeric channels that are specifically transported to the axonal initial segment and nodes of Ranvier. In contrast, there is no evidence for functional homomeric KCNQ3 channels in neurons, and it appears that these are inefficiently trafficked to the plasma membrane. Among eukaryotic potassium channels, the KCNQ3 subunit is unusual because it has an alanine in place of a threonine at the pore inner vestibule, three residues upstream of the GYG signature sequence of the selectivity filter. This residue is critical for the potentiation of the current after heteromerization, but the mechanism is unknown. We report that the presence of this uncommon residue at position 315 has a strong impact on the stability of the homotetramers and on channel trafficking. Wild-type KCNQ3 expressed alone is retained within the endoplasmic reticulum, and this mechanism is overcome by the substitution of threonine for Ala315. KCNQ3 subunits require assembly with KCNQ2 to exit this compartment, whereas KCNQ3-A315T is no longer dependent on KCNQ2 to form channels that are efficiently trafficked to the plasma membrane. The presence of this alanine, therefore, plays an important role in regulating the subunit composition of functional M-channels expressed at the surface of neurons.

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“…Biochemical, spectroscopic, and crystallographic analyses have recently provided the structure model for the C terminus of KCNQ1, which consists of four ␣-helices, helices A, B, C, and D, and an unstructured loop between helices A and B (31). Helices A and B provide binding sites for CaM, and the Cys 445 , the target of S-nitrosylation resides in the midst of the unstructured loop between helix A and B.…”

Section: Discussionmentioning

confidence: 99%

Redox- and Calmodulin-dependent S-Nitrosylation of the KCNQ1 Channel

Asada

Kurokawa

Furukawa

2009

Journal of Biological Chemistry

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S-Nitrosylation is a nitric oxide (NO) 2 -induced post-translational modification in which a cysteinyl thiol (R-SH) is converted to a nitrosothiol (1-3) and acts as a regulatory mechanism of various classes of proteins, including ion channels, such as the skeletal muscle type ryanodine receptor (ryanodine receptor type 1) channel (4, 5), the N-methyl-D-aspartate receptor channel (6, 7), the cardiac L-type Ca 2ϩ channel (8), and the cardiac Na ϩ channel (9). We have previously reported that NO derived from endothelial NO synthase activates ion currents through the cardiac slowly activating delayed rectifier potassium channel (I Ks ) composed of the pore-forming ␣-subunit KCNQ1 and the auxiliary ␤-subunit KCNE1. The NO-dependent regulation of the I Ks channel plays a pivotal role in regulation of cardiac membrane potential by intracellular Ca 2ϩ (10) and by sex hormones (11-13). Because the NO-dependent I Ks activation was inhibited by an inhibitor of soluble guanylate cyclase, 1H-(1,2,4)oxadiazolo(4,3-a)quinoxlin-1-1 (ODQ), with only a limited magnitude but was robustly inhibited by a thiol-alkylating reagent, N-ethylmaleimide, and reversed by a reducing reagent, dithiothreitol, soluble guanylate cyclase-independent, the protein S-nitrosylation mechanism is posited to be mainly involved (14). However, the following issues remain to be addressed: (i) Is the I Ks channel S-nitrosylated? (ii) If so, then what is the target of S-nitrosylation between the ␣-subunit KCNQ1 and the ␤-subunit KCNE1? (iii) Among multiple Cys residues, which Cys is a target of S-nitrosylation? and (iv) How does NO specifically recognize the target Cys? In the present study, we used the biotin-switch assay and functional patch clamp experiment to answer these questions. Our data show that KCNQ1 is a target of S-nitrosylation, and the presence of a redox motif contributes to making the Cys at 445 in the C terminus of KCNQ1 a preferential target of S-nitrosylation.

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The KCNQ1 (Kv7.1) COOH Terminus, a Multitiered Scaffold for Subunit Assembly and Protein Interaction

Cited by 127 publications

References 44 publications

Multimeric nature of voltage-gated proton channels

Multimeric nature of voltage-gated proton channels

A Pore Residue of the KCNQ3 Potassium M-Channel Subunit Controls Surface Expression

Redox- and Calmodulin-dependent S-Nitrosylation of the KCNQ1 Channel

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