Energetics and Location of Phosphoinositide Binding in Human Kir2.1 Channels

D’Avanzo, Nazzareno; Lee, Sun-Joo; Cheng, Wayland W.L.; Nichols, Colin G.

doi:10.1074/jbc.m113.452540

Cited by 35 publications

(46 citation statements)

References 44 publications

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“…54,55 Relative to other Kir channels, Kir 7.1 has a weak affinity for PIP 2 which results in a more pronounced inhibition by PIP 2 hydrolysis compared to other Kir channels. 19,56 We observed an OXT-mediated decrease in the Kir7.1 channel current by 15.5 pA/pF or 68±3% in HEK-OXTR cells (Fig.…”

Section: Discussionmentioning

confidence: 99%

Oxytocin (OXT)-stimulated inhibition of Kir7.1 activity is through PIP 2 -dependent Ca 2+ response of the oxytocin receptor in the retinal pigment epithelium in vitro

York

Halbach

Chiu

et al. 2017

Cellular Signalling

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Oxytocin (OXT) is a neuropeptide that activates the oxytocin receptor (OXTR), a rhodopsin family G-protein coupled receptor. Our localization of OXTR to the retinal pigment epithelium (RPE), in close proximity to OXT in the adjacent photoreceptor neurons, leads us to propose that OXT plays an important role in RPE-retinal communication. An increase of RPE [Ca2+]i in response to OXT stimulation implies that the RPE may utilize oxytocinergic signaling as a mechanism by which it accomplishes some of its many roles. In this study, we used an established human RPE cell line, a HEK293 heterologous OXTR expression system, and pharmacological inhibitors of Ca2+ signaling to demonstrate that OXTR utilizes capacitative Ca2+ entry (CCE) mechanisms to sustain an increase in cytoplasmic Ca2+. These findings demonstrate how multiple functional outcomes of OXT-OXTR signaling could be integrated via a single pathway. In addition, the activated OXTR was able to inhibit the Kir7.1 channel, an important mediator of sub retinal waste transport and K+ homeostasis.

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Section: Discussionmentioning

confidence: 99%

Oxytocin (OXT)-stimulated inhibition of Kir7.1 activity is through PIP 2 -dependent Ca 2+ response of the oxytocin receptor in the retinal pigment epithelium in vitro

York

Halbach

Chiu

et al. 2017

Cellular Signalling

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“…Instead, each PIP ligand can bind with in the same overall location with varying energies, but in non-identical conformations (D'Avanzo et al, 2013). Biochemical assays suggests that the key interactions which govern binding of each particular PIP are different.…”

Section: Structural Determinants Of Pip Sensitivity and Selectivitymentioning

confidence: 99%

Phosphoinositide regulation of inward rectifier potassium (Kir) channels

2014

Self Cite

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Inward rectifier potassium (Kir) channels are integral membrane proteins charged with a key role in establishing the resting membrane potential of excitable cells through selective control of the permeation of K+ ions across cell membranes. In conjunction with secondary anionic phospholipids, members of this family are directly regulated by phosphoinositides (PIPs) in the absence of other proteins or downstream signaling pathways. Different Kir isoforms display distinct specificities for the activating PIPs but all eukaryotic Kir channels are activated by PI(4,5)P2. On the other hand, the bacterial KirBac1.1 channel is inhibited by PIPs. Recent crystal structures of eukaryotic Kir channels in apo and lipid bound forms reveal one specific binding site per subunit, formed at the interface of N- and C-terminal domains, just beyond the transmembrane segments and clearly involving some of the key residues previously identified as controlling PI(4,5)P2 sensitivity. Computational, biochemical, and biophysical approaches have attempted to address the energetic determinants of PIP binding and selectivity among Kir channel isoforms, as well as the conformational changes that trigger channel gating. Here we review our current understanding of the molecular determinants of PIP regulation of Kir channel activity, including in context with other lipid modulators, and provide further discussion on the key questions that remain to be answered.

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“…On the other hand, crystal structures of Kir2.2 and Kir3.2 channels bound to PIP2 show only one specific site per subunit [4,16]. This result was further elucidated with a PIP strip assay [17], which suggests R218 as one most essential residue for PIP2 binding. However, these later approaches including X-ray crystallography used proteins solubilized by detergents.…”

Section: Interaction Site For Pip2mentioning

confidence: 99%

Novel Methodology to Detect Physical Interaction of Inositol Phospholipid with Potassium Ion Channel Using Luminescence Resonance Energy Transfer

Chin

Cheng

2019

Preprint

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Membrane proteins like ion channels are located within a lipid environment. While the bulk lipids provide a supporting architecture for proteins, some lipids also play important signaling roles in modulating protein activity. This modulation can occur through direct interaction with proteins without involving second messenger pathways. Phosphoinositides are one such lipids, which have been implicated in interactions with a diverse range of proteins and such interactions mediate a variety of cellular functions. Towards understanding how these lipids play their roles in regulating protein functions It is necessary to detect interactions between proteins and lipids and to identify binding sites on proteins. However, detection of protein-lipid interactions has been difficult especially in their native environment because only a fraction of lipids are bound to proteins while the bulk is freely diffusing. The purpose of this study is to develop an experimental method capable of identifying protein-lipid interactions in membrane environments. The strategy involves luminescence energy transfer to locally illuminate lipids around proteins thereby suppressing the large background of light from unbound lipid molecules. The approach is applied to an inward rectifier potassium ion channel for its interactions with phosphoinositide lipids. Experiments show that it can successfully detect phosphoinositides bound to proteins, and when combined with mutagenic approaches, can further identify binding site. The approach thus provides a valuable methodology for probing protein-lipid interactions in a native-like lipid environment.

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Energetics and Location of Phosphoinositide Binding in Human Kir2.1 Channels

Cited by 35 publications

References 44 publications

Oxytocin (OXT)-stimulated inhibition of Kir7.1 activity is through PIP 2 -dependent Ca 2+ response of the oxytocin receptor in the retinal pigment epithelium in vitro

Oxytocin (OXT)-stimulated inhibition of Kir7.1 activity is through PIP 2 -dependent Ca 2+ response of the oxytocin receptor in the retinal pigment epithelium in vitro

Phosphoinositide regulation of inward rectifier potassium (Kir) channels

Novel Methodology to Detect Physical Interaction of Inositol Phospholipid with Potassium Ion Channel Using Luminescence Resonance Energy Transfer

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