Molecular Localization of the Inhibitory Arachidonic Acid Binding Site to the Pore of hIK1

Hamilton, Kirk L.; Syme, Colin A.; Devor, Daniel C.

doi:10.1074/jbc.m212959200

Cited by 54 publications

(68 citation statements)

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“…Although we cannot formally rule out that a depolarizing surface charge effect on KAT1 is offset by a competing effect of some other form of regulation, it is interesting to note that KCNQ1 and KCNQ2 also display isoform-selective responses to 4␤PMA (Nakajo and Kubo, 2005). It is also interesting to note that some actions of PA are mediated via specific coupling of the lipids to polybasic clusters in target proteins (Limatola et al, 1994;Frank et al, 1999;Grange et al, 2000;Sergeant et al, 2001;Nakajo and Kubo, 2005), as has been described for 4,5-PIP2 (Zhang et al, 1999;Shyng et al, 2000), whereas regulation of several voltage-gated channels by AA and its metabolites appears to involve lipid interaction with membrane-buried elements of the pore (Hamilton et al, 2003;Oliver et al, 2004).…”

Section: Discussionmentioning

confidence: 99%

HCN Pacemaker Channel Activation Is Controlled by Acidic Lipids Downstream of Diacylglycerol Kinase and Phospholipase A2

Fogle¹,

Lyashchenko²,

Turbendian³

et al. 2007

J. Neurosci.

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Section: Discussionmentioning

confidence: 99%

HCN Pacemaker Channel Activation Is Controlled by Acidic Lipids Downstream of Diacylglycerol Kinase and Phospholipase A2

Fogle¹,

Lyashchenko²,

Turbendian³

et al. 2007

J. Neurosci.

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“…In addition, we have demonstrated that ATP activates hIK1 via cAMP-dependent protein kinase and that this activation relies upon a COOH-terminal domain overlapping the Ca 2ϩ -dependent, calmodulin-binding domain (12,13). We have also demonstrated that hIK1 is inhibited by arachidonic acid (4), recently mapping the inhibitory arachidonic acid binding site to the pore of hIK1 (14). Furthermore the regulation of hIK1 by protein kinase C has been defined at the molecular level (15).…”

mentioning

confidence: 84%

Role of the NH2 Terminus in the Assembly and Trafficking of the Intermediate Conductance Ca2+-activated K+ Channel hIK1

Jones

Hamilton

Papworth

et al. 2004

Journal of Biological Chemistry

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The role of the NH 2 -terminal leucine zipper and dileucine motifs of hIK1 in the assembly, trafficking, and function of the channel was investigated using cell surface immunoprecipitation, co-immunoprecipitation (Co-IP), immunoblot, and whole-cell patch clamp techniques. Mutation of the NH 2 -terminal leucine zipper at amino acid positions 18 and 25 (L18A/L25A) resulted in a complete loss of steady-state protein expression, cell surface expression, and whole-cell current density. Inhibition of proteasomal degradation with lactacystin restored L18A/L25A protein expression, although this channel was not expressed at the cell surface as assessed by cell surface immunoprecipitation and wholecell patch clamp. In contrast, inhibitors of lysosomal degradation (leupeptin/pepstatin) and endocytosis (chloroquine) had little effect on L18A/L25A protein expression or localization. Further studies confirmed the rapid degradation of this channel, having a time constant of 19.0 ؎ 1.3 min compared with 3.2 ؎ 0.8 h for wild type hIK1. Co-expression studies demonstrated that the L18A/L25A channel associates with wild type channel, thereby attenuating its expression at the cell surface. Co-IP studies confirmed this association. However, L18A/L25A channels failed to form homotetrameric channels, as assessed by Co-IP, suggesting the NH 2 terminus plays a role in tetrameric channel assembly. As with the leucine zipper, mutation of the dileucine motif to alanines, L18A/L19A, resulted in a near complete loss in steady-state protein expression with the protein being similarly targeted to the proteasome for degradation. In contrast to our results on the leucine zipper, however, both chloroquine and growing the cells at the permissive temperature of 27°C restored expression of L18A/L19A at the cell surface, suggesting that the defect in the channel trafficking is the result of a subtle folding error. In conclusion, we demonstrate that the NH 2 terminus of hIK1 contains overlapping leucine zipper and dileucine motifs essential for channel assembly and trafficking to the plasma membrane.The KCNN gene family is composed of four members, including the small conductance Ca 2ϩ -activated K ϩ channels (SK1, SK2, and SK3) 1 and the intermediate conductance Ca 2ϩ -activated K ϩ channel (IK1 or SK4). While the IK1 and SK channels share roughly 40% homology, their expression patterns and pharmacology are widely disparate, consistent with their unique physiological functions (1). The human IK1 channel (hIK1) is widely expressed, being found in salivary gland, colon, bladder, stomach, lung, smooth muscle, red blood cells, T-cells, and placenta (2, 3) where it plays a crucial role in a variety of physiological functions. Indeed hIK1 plays a seminal role in modulating Ca 2ϩ -dependent transepithelial ion transport (4, 5), has recently been confirmed to be the Gardos channel of red blood cells (6), and has been proposed to play a role in both vascular remodeling (7) and in modulating vascular tone (8). Because of these critical physiological roles, the ...

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“…Also, openers have been shown to alter vascular tone and hence may modulate peripheral blood pressure (16,17). A great deal of research has focused on both the biophysical and pharmacological properties of hIK1 (18 -20) and more recently on the assembly and second messenger activation (21)(22)(23)(24). In this study, we have begun to investigate the role that the S4-S5 linker region plays in the function and trafficking of IK and SK channels.…”

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confidence: 99%

“…Electrophysiology-All electrophysiological recordings were carried out as described previously (21,22,35). During whole-cell patch-clamp experiments, the bath contained 140 mM NaCl, 4 mM KCl, 2 mM CaCl 2 , 1 mM MgCl 2 , and 10 mM HEPES (pH adjusted to 7.4 with NaOH).…”

mentioning

confidence: 99%

Role of an S4-S5 Linker Lysine in the Trafficking of the Ca2+-activated K+ Channels IK1 and SK3

Jones

Hamilton

Devor

2005

Journal of Biological Chemistry

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We have investigated the role of the S4-S5 linker in the trafficking of the intermediate (human (h) IK1) and small (rat SK3) conductance K ؉ channels using a combination of patch-clamp, protein biochemical, and immunofluorescence-based techniques. We demonstrate that a lysine residue (Lys 197 ) located on the intracellular loop between the S4 and S5 domains is necessary for the correct trafficking of hIK1 to the plasma membrane. Mutation of this residue to either alanine or methionine precluded trafficking of the channel to the membrane, whereas the charge-conserving arginine mutation had no effect on channel localization or function. Immunofluorescence localization demonstrated that the K197A mutation resulted in a channel that was primarily retained in the endoplasmic reticulum, and this could not be rescued by incubation at 27°C. Furthermore, immunoblot analysis revealed that the K197A mutation was overexpressed compared with wild-type hIK1 and that this was due to a greatly diminished rate of channel degradation. Co-immunoprecipitation studies demonstrated that the K197A mutation did not preclude multimer formation. Indeed, the K197A mutation dramatically suppressed expression of wild-type hIK1 at the cell surface. Finally, mutation of this conserved lysine in rat SK3 similarly resulted in a channel that failed to correctly traffic to the plasma membrane. These results are the first to demonstrate a critical role for the S4-S5 linker in the trafficking and/or function of IK and SK channels.The human (h) 3 intermediate conductance Ca 2ϩ -activated K ϩ channel IK1 is a member of the KCNN gene family, which also includes SK1-3, the small conductance Ca 2ϩ -activated K ϩ channels. Within this channel family, there is ϳ40% amino acid sequence homology, with the greatest level of identity occurring in the pore and proximal C terminus, a region known to constitutively bind calmodulin (1, 2), thereby conferring Ca 2ϩ sensitivity to these channels. The hIK1 channel is now known to be the Gardos channel involved in red blood cell volume regulation (3). hIK1 is also known to be involved in the Ca 2ϩ -dependent regulation of Cl Ϫ secretion across intestinal and airway epithelia (4 -8). More recent work has demonstrated that the progression of breast cancer cells through the cell cycle is dependent on membrane hyperpolarization resulting from the activation of hIK1 channels (9). hIK1 has also been shown to play a role in both macrophage activation and T-lymphocyte proliferation (10 -12). The critical role that pharmacological modulation of hIK1 may play has been revealed by the demonstration that blockers of hIK1 reduce the autoimmune response to experimental encephalomyelitis in mice (13), reduce brain edema following traumatic brain injury (14), and prevent restenosis following balloon angioplasty (15). Also, openers have been shown to alter vascular tone and hence may modulate peripheral blood pressure (16,17). A great deal of research has focused on both the biophysical and pharmacological properties of hIK1 (18 -20)...

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Molecular Localization of the Inhibitory Arachidonic Acid Binding Site to the Pore of hIK1

Cited by 54 publications

References 50 publications

HCN Pacemaker Channel Activation Is Controlled by Acidic Lipids Downstream of Diacylglycerol Kinase and Phospholipase A2

HCN Pacemaker Channel Activation Is Controlled by Acidic Lipids Downstream of Diacylglycerol Kinase and Phospholipase A2

Role of the NH2 Terminus in the Assembly and Trafficking of the Intermediate Conductance Ca2+-activated K+ Channel hIK1

Role of an S4-S5 Linker Lysine in the Trafficking of the Ca2+-activated K+ Channels IK1 and SK3

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