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Carbenoxolone inhibits Pannexin1 channels through interactions in the first extracellular loop

Pannexin1 (Panx1) is an ATP release channel important for controlling immune responses and synaptic strength. Various stimuli including C-terminal cleavage, a high concentration of extracellular potassium, and voltage have been demonstrated to activate Panx1. However, it remains unclear how Panx1 se...

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Autores principales: Michalski, Kevin, Kawate, Toshimitsu
Formato: Online Artículo Texto
Lenguaje:English
Publicado: The Rockefeller University Press 2016
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4727946/
https://www.ncbi.nlm.nih.gov/pubmed/26755773
http://dx.doi.org/10.1085/jgp.201511505
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author Michalski, Kevin
Kawate, Toshimitsu
author_facet Michalski, Kevin
Kawate, Toshimitsu
author_sort Michalski, Kevin
collection PubMed
description Pannexin1 (Panx1) is an ATP release channel important for controlling immune responses and synaptic strength. Various stimuli including C-terminal cleavage, a high concentration of extracellular potassium, and voltage have been demonstrated to activate Panx1. However, it remains unclear how Panx1 senses and integrates such diverse stimuli to form an open channel. To provide a clue on the mechanism underlying Panx1 channel gating, we investigated the action mechanism of carbenoxolone (CBX), the most commonly used small molecule for attenuating Panx1 function triggered by a wide range of stimuli. Using a chimeric approach, we discovered that CBX reverses its action polarity and potentiates the voltage-gated channel activity of Panx1 when W74 in the first extracellular loop is mutated to a nonaromatic residue. A systematic mutagenesis study revealed that conserved residues in this loop also play important roles in CBX function, potentially by mediating CBX binding. We extended our experiments to other Panx1 inhibitors such as probenecid and ATP, which also potentiate the voltage-gated channel activity of a Panx1 mutant at position 74. Notably, probenecid alone can activate this mutant at a resting membrane potential. These data suggest that CBX and other inhibitors, including probenecid, attenuate Panx1 channel activity through modulation of the first extracellular loop. Our experiments are the first step toward identifying a previously unknown mode of CBX action, which provide insight into the role of the first extracellular loop in Panx1 channel gating.
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spelling pubmed-47279462016-08-01 Carbenoxolone inhibits Pannexin1 channels through interactions in the first extracellular loop Michalski, Kevin Kawate, Toshimitsu J Gen Physiol Research Articles Pannexin1 (Panx1) is an ATP release channel important for controlling immune responses and synaptic strength. Various stimuli including C-terminal cleavage, a high concentration of extracellular potassium, and voltage have been demonstrated to activate Panx1. However, it remains unclear how Panx1 senses and integrates such diverse stimuli to form an open channel. To provide a clue on the mechanism underlying Panx1 channel gating, we investigated the action mechanism of carbenoxolone (CBX), the most commonly used small molecule for attenuating Panx1 function triggered by a wide range of stimuli. Using a chimeric approach, we discovered that CBX reverses its action polarity and potentiates the voltage-gated channel activity of Panx1 when W74 in the first extracellular loop is mutated to a nonaromatic residue. A systematic mutagenesis study revealed that conserved residues in this loop also play important roles in CBX function, potentially by mediating CBX binding. We extended our experiments to other Panx1 inhibitors such as probenecid and ATP, which also potentiate the voltage-gated channel activity of a Panx1 mutant at position 74. Notably, probenecid alone can activate this mutant at a resting membrane potential. These data suggest that CBX and other inhibitors, including probenecid, attenuate Panx1 channel activity through modulation of the first extracellular loop. Our experiments are the first step toward identifying a previously unknown mode of CBX action, which provide insight into the role of the first extracellular loop in Panx1 channel gating. The Rockefeller University Press 2016-02 /pmc/articles/PMC4727946/ /pubmed/26755773 http://dx.doi.org/10.1085/jgp.201511505 Text en © 2016 Michalski and Kawate This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 3.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/3.0/).
spellingShingle Research Articles
Michalski, Kevin
Kawate, Toshimitsu
Carbenoxolone inhibits Pannexin1 channels through interactions in the first extracellular loop
title Carbenoxolone inhibits Pannexin1 channels through interactions in the first extracellular loop
title_full Carbenoxolone inhibits Pannexin1 channels through interactions in the first extracellular loop
title_fullStr Carbenoxolone inhibits Pannexin1 channels through interactions in the first extracellular loop
title_full_unstemmed Carbenoxolone inhibits Pannexin1 channels through interactions in the first extracellular loop
title_short Carbenoxolone inhibits Pannexin1 channels through interactions in the first extracellular loop
title_sort carbenoxolone inhibits pannexin1 channels through interactions in the first extracellular loop
topic Research Articles
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4727946/
https://www.ncbi.nlm.nih.gov/pubmed/26755773
http://dx.doi.org/10.1085/jgp.201511505
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