Structural and Functional Determinants of Conserved Lipid Interaction Domains of Inward Rectifying Kir6.2 Channels

Cukras, Catherine A; Jeliazkova, Iana; Nichols, Colin G.

doi:10.1085/jgp.20028562

Cited by 58 publications

(55 citation statements)

References 62 publications

(124 reference statements)

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“…single or multiple amino acid exchanges (Table 1). In Xenopus oocytes expressing Kir6.2ΔC26 W311A and Kir6.2ΔC26 D329A, no current could be detected in single-channel recordings from membrane patches using the inside-out configuration, in agreement with a previous report [23]. Replacing the lysine (K) with an alanine (A) at position 332 (K332A) resulted in a basal K + current at the same intensity as Kir6.2ΔC26 (Table 1).…”

Section: Resultssupporting

confidence: 90%

See 1 more Smart Citation

Single residue (K332A) substitution in Kir6.2 abolishes the stimulatory effect of long-chain acyl-CoA esters: indications for a long-chain acyl-CoA ester binding motif

Bränström

Leibiger

et al. 2007

Diabetologia

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Aims/hypothesis The pancreatic beta cell ATP-sensitive potassium (K ATP ) channel, composed of the pore-forming α subunit Kir6.2, a member of the inward rectifier K+ channel family, and the regulatory β subunit sulfonylurea receptor 1 (SUR1), a member of the ATP-binding cassette superfamily, couples the metabolic state of the cell to electrical activity. Several endogenous compounds are known to modulate K ATP channel activity, including ATP, ADP, phosphatidylinositol diphosphates and long-chain acyl coenzyme A (LC-CoA) esters. LC-CoA esters have been shown to interact with Kir6.2, but the mechanism and binding site(s) have yet to be identified. Materials and methods Using multiple sequence alignment of known acyl-CoA ester interacting proteins, we were able to identify four conserved amino acid residues that could potentially serve as an acyl-CoA ester-binding motif. The motif was also recognised in the C-terminal region of Kir6.2 (R311-332) but not in SUR1.Results Oocytes expressing Kir6.2ΔC26 K332A repeatedly generated K + currents in inside-out membrane patches that were sensitive to ATP, but were only weakly activated by 1 μmol/l palmitoyl-CoA ester. Compared with the control channel (Kir6.2ΔC26), Kir6.2ΔC26 K332A displayed unaltered ATP sensitivity but significantly decreased sensitivity to palmitoyl-CoA esters. Coexpression of Kir6.2ΔC26 K332A and SUR1 revealed slightly increased activation by palmitoyl-CoA ester but significantly decreased activation by the acyl-CoA esters compared with the wild-type K ATP channel and Kir6.2ΔC26+SUR1. Computational modelling, using the crystal structure of KirBac1.1, suggested that K332 is located on the intracellular domain of Kir6.2 and is accessible to intracellular modulators such as LC-CoA esters. Conclusions/interpretation These results verify that LCCoA esters interact at the pore-forming subunit Kir6.2, and on the basis of these data we propose an acyl-CoA ester binding motif located in the C-terminal region.

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Section: Resultssupporting

confidence: 90%

“…However, the proposed LCCoA ester binding motif, which we have now identified as 311-332, is not close to the residues that have been proposed for ATP and PIP2 interaction [12,17,24,29,30], except for 314 [31] and 308-311 [23]. All members of the Kir channel family tested so far are activated by PIPs, especially PIP2.…”

Section: Discussionmentioning

confidence: 59%

Single residue (K332A) substitution in Kir6.2 abolishes the stimulatory effect of long-chain acyl-CoA esters: indications for a long-chain acyl-CoA ester binding motif

Bränström

Leibiger

et al. 2007

Diabetologia

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“…Binding of PIP 2 to both K ir 1 and K ir 6 channels promotes channel activation by stabilizing the open state (32)(33)(34), and PIP 3 has been proposed to activate both epithelial sodium channels and TRPC6 channels by direct binding (35,36). Like PIP 2 -binding regions identified in other ion channels (37)(38)(39), the stretch of amino acids between residues 61-90 of CNGA2 contains multiple basic residues that may be important for the interaction with negatively charged phospholipids.…”

Section: Discussionmentioning

confidence: 99%

Interplay between PIP ₃ and calmodulin regulation of olfactory cyclic nucleotide-gated channels

Brady

Rich

Martens

et al. 2006

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

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Phosphatidylinositol-3,4,5-trisphosphate (PIP3) has been proposed to modulate the odorant sensitivity of olfactory sensory neurons by inhibiting activation of cyclic nucleotide-gated (CNG) channels in the cilia. When applied to the intracellular face of excised patches, PIP3 has been shown to inhibit activation of heteromeric olfactory CNG channels, composed of CNGA2, CNGA4, and CNGB1b subunits, and homomeric CNGA2 channels. In contrast, we discovered that channels formed by CNGA3 subunits from cone photoreceptors were unaffected by PIP3. Using chimeric channels and a deletion mutant, we determined that residues 61-90 within the N terminus of CNGA2 are necessary for PIP3 regulation, and a biochemical ''pulldown'' assay suggests that PIP3 directly binds this region. The N terminus of CNGA2 contains a previously identified calcium-calmodulin (Ca 2؉ ͞CaM)-binding domain (residues 68 -81) that mediates Ca 2؉ ͞CaM inhibition of homomeric CNGA2 channels but is functionally silent in heteromeric channels. We discovered, however, that this region is required for PIP3 regulation of both homomeric and heteromeric channels. Furthermore, PIP3 occluded the action of Ca 2؉ ͞CaM on both homomeric and heteromeric channels, in part by blocking Ca 2؉ ͞CaM binding. Our results establish the importance of the CNGA2 N terminus for PIP3 inhibition of olfactory CNG channels and suggest that PIP3 inhibits channel activation by disrupting an autoexcitatory interaction between the N and C termini of adjacent subunits. By dramatically suppressing channel currents, PIP3 may generate a shift in odorant sensitivity that does not require prior channel activity.lipid signaling ͉ olfaction ͉ phosphatidylinositide ͉ sensory adaptation O dorant binding to specialized receptors in the cilia of olfactory sensory neurons triggers an increase in intracellular cAMP (1-4), which directly opens cyclic nucleotide-gated (CNG) channels (5). Calcium influx through CNG channels activates an atypical chloride current (6-8), leading to depolarization of the cell membrane. The elevated calcium also causes rapid adaptation to odorants by triggering a calcium-calmodulin (Ca 2ϩ ͞CaM)-dependent decrease in the sensitivity of CNG channels to cAMP (9). Recent evidence suggests that phosphatidylinositol-3,4,5-trisphosphate (PIP 3 ) also decreases the sensitivity of olfactory CNG channels and reduces the response of olfactory sensory neurons to complex odors, but the mechanism has yet to be elucidated (10, 11).Ca 2ϩ ͞CaM inhibits homomeric CNGA2 channel activation by binding to a Baa-like motif in the N terminus (12-14), thereby disrupting an autostimulatory interaction with the C terminus of an adjacent subunit (15-17). Deletion of the Ca 2ϩ ͞CaM-binding domain (amino acids 68-81) in CNGA2 produces channels that are resistant to inhibition by Ca 2ϩ ͞CaM and exhibit dramatically reduced sensitivity to cyclic nucleotides due to the loss of the autostimulatory interaction. Native olfactory CNG channels are tetrameric assemblies of three different pore-forming subunits, C...

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“…Recently, long chain acyl-CoA esters were found to modulate ATP inhibition of the K ATP channels by the same mechanisms as PIP 2 (14), suggesting these lipid metabolites shared a common site of interaction on the channel. Indeed, it has been proposed that there is a novel lipid-interacting motif for inward rectifier K ϩ channels termed KIRLI domain (45). The core of this domain is composed of residues 170 -320, and the structure is composed of antiparallel ␤ strands and an ␣ helix, which closely resembles that of the pleckstrin homology (PH) domain, a known PIP 2 binding motif.…”

Section: Discussionmentioning

confidence: 99%

Molecular Determinants of Cardiac KATP Channel Activation by Epoxyeicosatrienoic Acids

Lü

Hong

Lee

2005

Journal of Biological Chemistry

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We have previously reported that epoxyeicosatrienoic acids (EETs), the cytochrome P450 epoxygenase metabolites of arachidonic acid, are potent stereospecific activators of the cardiac K ATP channel. The epoxide group in EET is critical for reducing channel sensitivity to ATP, thereby activating the channel. This study is to identify the molecular sites on the K ATP channels for EET-mediated activation. We investigated the effects of EETs on Kir6.2⌬C26 with or without the coexpression of SUR2A and on Kir6. The cardiac ATP-sensitive K ϩ (K ATP ) 1 channels are biological sensors that respond to intracellular metabolic changes and play important roles in regulating cardiac functions (1). Although the role of the cardiac K ATP channels under normal physiological conditions remains unclear, it is crucial in ischemic preconditioning, and activation of K ATP channels occurs during cardiac ischemia and hypoxia, leading to reduced Ca 2ϩ influx and intracellular Ca 2ϩ overload (2). Some important insights were provided by studies using the Kir6.2 knock-out mice, which have compromised ability in modulating cardiac electrophysiological properties, contractility (3, 4), and in handling cardiac ischemia and stress (3,5,6). Recently, a form of human dilated cardiomyopathy was found to be associated with mutations of the cardiac K ATP channels (7).The cardiac K ATP channel is a heterooctamer containing four inward rectifier K ϩ channel (Kir6.2) and four sulfonylurea receptor (SUR2A) subunits. SUR contains two cytoplasmic nucleotide binding folds that were initially thought to be important for channel regulation by ATP (8). However, a truncation mutant of the Kir6.2 involving deletion of the C-terminal 26 residues (Kir6.2⌬C26) gives rise to channels that retain much of the ATP sensitivity in the absence of SUR (9). Also, mutations in the SUR produced only small effects on ATP inhibition of Kir6.2/SUR currents (10). Previous studies have demonstrated that both the N and C termini of Kir6.2 contribute to the site(s) that regulates ATP sensitivity, and they include Arg-50, Cys-166, Ile-167, Thr-171, Arg-176, Arg-177, Glu-179, Ile-182, Lys-185, Arg-192, Arg-201, and Gly-344 residues. Of these, Arg-50, Lys-185, and Arg-201 residues are particularly crucial for ATP sensitivity and are implicated for interaction with the ␣, ␤, and ␥ phosphates of ATP (11-13). However, the mechanism of Kir6.2 channel inhibition by ATP remains to be elucidated.Lipid metabolites are important modulators of the K ATP channels, and these include the long chain acyl-CoA esters (14), phosphatidylinositol 4,5-bisphosphate (PIP 2 ), phosphoinositides (15), L-palmitoylcarnitine (16), and epoxyeicosatrienoic acids (EETs) (17, 18). Arachidonic acid is converted by the cytochrome P450 epoxygenase into 4 EET regioisomers, 5.6-, 8,9-, 11,12-, and 14,15-EET (Fig. 1) (19). EETs are abundant endogenous constituents of the human and rat hearts, measured at 70 ng of total EETs per g of rat heart (20). 8,9-, 11,12-, and 14,15-EET contribute 39, 28, and 33%, respectively, ...

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Structural and Functional Determinants of Conserved Lipid Interaction Domains of Inward Rectifying Kir6.2 Channels

Cited by 58 publications

References 62 publications

Single residue (K332A) substitution in Kir6.2 abolishes the stimulatory effect of long-chain acyl-CoA esters: indications for a long-chain acyl-CoA ester binding motif

Single residue (K332A) substitution in Kir6.2 abolishes the stimulatory effect of long-chain acyl-CoA esters: indications for a long-chain acyl-CoA ester binding motif

Interplay between PIP ₃ and calmodulin regulation of olfactory cyclic nucleotide-gated channels

Molecular Determinants of Cardiac KATP Channel Activation by Epoxyeicosatrienoic Acids

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Structural and Functional Determinants of Conserved Lipid Interaction Domains of Inward Rectifying Kir6.2 Channels

Cited by 58 publications

References 62 publications

Single residue (K332A) substitution in Kir6.2 abolishes the stimulatory effect of long-chain acyl-CoA esters: indications for a long-chain acyl-CoA ester binding motif

Single residue (K332A) substitution in Kir6.2 abolishes the stimulatory effect of long-chain acyl-CoA esters: indications for a long-chain acyl-CoA ester binding motif

Interplay between PIP 3 and calmodulin regulation of olfactory cyclic nucleotide-gated channels

Molecular Determinants of Cardiac KATP Channel Activation by Epoxyeicosatrienoic Acids

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Interplay between PIP ₃ and calmodulin regulation of olfactory cyclic nucleotide-gated channels