Localization of the ATP/Phosphatidylinositol 4,5 Diphosphate-binding Site to a 39-Amino Acid Region of the Carboxyl Terminus of the ATP-regulated K+ Channel Kir1.1

Dong, Ke; Tang, Lieqi; MacGregor, Gordon G.; Hebert, Steven C.

doi:10.1074/jbc.m208679200

Cited by 28 publications

(24 citation statements)

References 29 publications

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“…PIP 2 is a well-known modulator of channel function 41,42 and has been shown to depend upon positively charged amino acids in cytosolic domains. [43][44][45] However, we (unpublished observations) and Donald Hilgemann (personal communication) have found no effect of PIP 2 on hIK1 channel function. Similarly, we found no effect of PI(3)P on hIK1 function in inside-out patches (unpublished observations).…”

Section: Discussionmentioning

confidence: 57%

An NH2-Terminal Multi-Basic RKR Motif Is Required for the ATP-Dependent Regulation of hIK1

et al. 2007

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We previously demonstrated that the ATP/PKA-dependent activation of the human intermediate conductance, Ca 2+ -activated K + channel, hIK1, is dependent upon a C-terminal motif. The NH 2 -terminus of hIK1 contains a multi-basic 13 RRRKR 17 motif, known to be important in the trafficking and function of ion channels. While individual mutations within this domain have no effect on channel function, the triple mutation ( 15 RKR 17 /AAA), as well as additional double mutations, result in a near complete loss of functional channels, as assessed by whole-cell patch-clamp. However, cell-surface immunoprecipitation studies confirmed expression of these mutated channels at the plasma membrane. To elucidate the functional consequences of the 15 RKR 17 /AAA mutation we performed inside-out patch clamp recordings where we observed no difference in Ca 2+ affinity between the wild-type and mutated channels. However, in contrast to wild-type hIK1, channels expressing the 15 RKR 17 /AAA mutation exhibited rundown, which could not be reversed by the addition of ATP. Wild-type hIK1 channel activity was reduced by alkaline phosphatase both in the presence and absence of ATP, indicative of a phosphorylation event, whereas the 15 RKR 17 /AAA mutation eliminated this effect of alkaline phosphatase. Further, single channel analysis demonstrated that the 15 RKR 17 /AAA mutation resulted in a four-fold lower channel open probability (P o ), in the presence of saturating Ca 2+ and ATP, compared to wild-type hIK1. In conclusion, these results represent the first demonstration for a role of the NH 2 -terminus in the second messenger-dependent regulation of hIK1 and, in combination with our previous findings, suggest that this regulation is dependent upon a close NH 2 /C-terminal association.

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Section: Discussionmentioning

confidence: 57%

An NH2-Terminal Multi-Basic RKR Motif Is Required for the ATP-Dependent Regulation of hIK1

et al. 2007

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“…2A, this region of the channel contains several well-conserved, positively charged arginines and lysines. In this regard, then, the putative PIP 2 binding site in ENaC is similar to that proposed for Kir and TRP channels, which all have an amino acid track containing several well-conserved, positively charged residues that potentially facilitate electrostatic interactions with the polar head groups of the phosphatidylinositides in the inner leaflet of the plasma membrane (17,58,66,73,97). Mutation of this domain in ␤-ENaC reduces ENaC activity without affecting surface expression (39).…”

Section: Pip 2 Directly Interacts With Enacmentioning

confidence: 64%

“…Point mutation of the conserved positively charged residues in this region also disrupts regulation of the channel by PI3-K. This PIP 3 binding site in ENaC then is similar to the phosphatidylinositide binding site in other channels in the respect that they all contain several positively charged conserved residues (17,58,66,73,97). Moreover, the location of this PIP 3 binding site in ENaC just distal to the second transmembrane domain is ideally suited to modulate open probability.…”

Section: Pip 3 Binding Sites In Enacmentioning

confidence: 92%

“…This mechanism for ion channel modulation is recognized to be physiologically important, for its disruption, in some instances, leads to disease (e.g., Bartter's and Andersen's syndromes) (16,43,56,65). Diverse types of ion channels are directly modulated by phosphatidylinositides (3,17,42,58,74,84,96,97). Phosphatidylinositides, such as phosphatidylinositol 4,5-bisphosphate (PIP 2 ) and phosphatidylinositol 3,4,5-triphosphate (PIP 3 ), directly interact with these channels to modulate gating (58,93,97).…”

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confidence: 99%

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Regulation of the epithelial Na⁺ channel (ENaC) by phosphatidylinositides

Pochynyuk

Tong

Staruschenko

et al. 2006

American Journal of Physiology-Renal Physiology

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The epithelial Na ϩ channel (ENaC) is an endeffector of diverse cellular signaling cascades, including those with phosphatidylinositide second messengers. Recent evidence also suggests that in some instances, phospatidylinositides can directly interact with ENaC to increase channel activity by increasing channel open probability and/or membrane localization. We review here findings relevant to regulation of ENaC by phosphatidylinositol 4,5-bisphosphate (PIP 2) and phosphatidylinositol 3,4,5-triphosphate (PIP3). Similar to its actions on other ion channels, PIP2 is permissive for ENaC openings having a direct effect on gating. The PIP2 binding site in ENaC involved in this regulation is most likely localized to the NH 2 terminus of ␤-ENaC. PIP3 also affects ENaC gating but, rather than being permissive, augments open probability. The PIP3 binding site in ENaC involved in this regulation is localized to the proximal region of the COOH terminus of ␥-ENaC just following the second transmembrane domain. In complementary pathways, PIP 3 also impacts ENaC membrane levels through both direct actions on the channel and via a signaling cascade involving phosphoinositide 3-OH kinase (PI3-K) and the aldosterone-induced gene product serum and glucocorticoid-inducible kinase. The putative PIP 3 binding site in ENaC involved in direct regulation of channel membrane levels has not yet been identified. phosphatidylinositol 4,5-bisphosphate; phosphatidylinositol 3,4,5-triphosphate; receptor tyrosine kinase; insulin G protein-coupled receptor ION CHANNELS PLAY CRITICAL roles in every aspect of physiology. Moreover, ion channels have long been recognized to be important end-effectors of diverse cellular signaling cascades, including those having phosphatidylinositide second messengers. Direct regulation of ion channel activity by phosphatidylinositide-signaling molecules, in addition, is now becoming widely appreciated (reviewed in Ref. 33). This mechanism for ion channel modulation is recognized to be physiologically important, for its disruption, in some instances, leads to disease (e.g., Bartter's and Andersen's syndromes) (16,43,56,65). Diverse types of ion channels are directly modulated by phosphatidylinositides (3,17,42,58,74,84,96,97). Phosphatidylinositides, such as phosphatidylinositol 4,5-bisphosphate (PIP 2 ) and phosphatidylinositol 3,4,5-triphosphate (PIP 3 ), directly interact with these channels to modulate gating (58,93,97). Several recent studies identify the epithelial Na ϩ channel (ENaC) as being a channel sensitive to direct phosphatidylinositide regulation.ENaC is a heteromeric channel composed of three distinct but similar subunits: ␣, ␤, and ␥ (8, 9). All ENaC subunits have NH 2 -and COOH-terminal cytosolic domains separated by two transmembrane domains and a large extracellular region. ENaC serves an essential physiological function, for its activity is limiting for Na ϩ absorption across many epithelia, including that in the distal renal nephron (reviewed in Refs. 27, 37, and 69). Thus ENaC is well positio...

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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. Similar to PIP 2 inhibition of TRPV1 channels (40), PIP 3 inhibited activation of olfactory CNG channels.…”

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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Localization of the ATP/Phosphatidylinositol 4,5 Diphosphate-binding Site to a 39-Amino Acid Region of the Carboxyl Terminus of the ATP-regulated K+ Channel Kir1.1

Cited by 28 publications

References 29 publications

An NH2-Terminal Multi-Basic RKR Motif Is Required for the ATP-Dependent Regulation of hIK1

An NH2-Terminal Multi-Basic RKR Motif Is Required for the ATP-Dependent Regulation of hIK1

Regulation of the epithelial Na⁺ channel (ENaC) by phosphatidylinositides

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

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Localization of the ATP/Phosphatidylinositol 4,5 Diphosphate-binding Site to a 39-Amino Acid Region of the Carboxyl Terminus of the ATP-regulated K+ Channel Kir1.1

Cited by 28 publications

References 29 publications

An NH2-Terminal Multi-Basic RKR Motif Is Required for the ATP-Dependent Regulation of hIK1

An NH2-Terminal Multi-Basic RKR Motif Is Required for the ATP-Dependent Regulation of hIK1

Regulation of the epithelial Na+ channel (ENaC) by phosphatidylinositides

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

Contact Info

Product

Resources

About

Regulation of the epithelial Na⁺ channel (ENaC) by phosphatidylinositides

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