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Structural and functional properties of the rat P2X4 purinoreceptor extracellular vestibule during gating

P2X receptors are ATP-gated cation channels consisting of three subunits that are mutually intertwined and form an upper, central, and extracellular vestibule with three lateral portals and the channel pore. Here we used cysteine and alanine scanning mutagenesis of the rat P2X4R receptor V47–V61 and...

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Autores principales: Rokic, Milos B., Stojilkovic, Stanko S., Zemkova, Hana
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2014
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3905210/
https://www.ncbi.nlm.nih.gov/pubmed/24523669
http://dx.doi.org/10.3389/fncel.2014.00003
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author Rokic, Milos B.
Stojilkovic, Stanko S.
Zemkova, Hana
author_facet Rokic, Milos B.
Stojilkovic, Stanko S.
Zemkova, Hana
author_sort Rokic, Milos B.
collection PubMed
description P2X receptors are ATP-gated cation channels consisting of three subunits that are mutually intertwined and form an upper, central, and extracellular vestibule with three lateral portals and the channel pore. Here we used cysteine and alanine scanning mutagenesis of the rat P2X4R receptor V47–V61 and K326–N338 sequences to study structural and functional properties of extracellular vestibule during gating. Cysteine mutants were used to test the accessibility of these residue side chains to cadmium during closed-open-desensitized transitions, whereas alanine mutants served as controls. This study revealed the accessibility of residues E51, T57, S59, V61, K326, and M336 to cadmium in channels undergoing a transition from a closed-to-open state and the accessibility of residues V47, G53, D331, I332, I333, T335, I337, and N338 in channels undergoing a transition from an open-to-desensitized state; residues E56 and K329 were accessible during both transitions. The effect of cadmium on channel gating was stimulatory in all reactive V47–V61 mutants and inhibitory in the majority of reactive K326–N338 mutants. The rat P2X4 receptor homology model suggests that residues affected by cadmium in the closed-to-open transition were located within the lumen of the extracellular vestibule and toward the central vestibule; however, the residues affected by cadmium in the open-to-desensitized state were located at the bottom of the vestibule near the pore. Analysis of the model assumed that there is ion access to extracellular and central vestibules through lateral ports when the channel is closed, with residues above the first transmembrane domain being predominantly responsible for ion uptake. Upon receptor activation, there is passage of ions toward the residues located on the upper region of the second transmembrane domain, followed by permeation through the gate region.
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spelling pubmed-39052102014-02-12 Structural and functional properties of the rat P2X4 purinoreceptor extracellular vestibule during gating Rokic, Milos B. Stojilkovic, Stanko S. Zemkova, Hana Front Cell Neurosci Neuroscience P2X receptors are ATP-gated cation channels consisting of three subunits that are mutually intertwined and form an upper, central, and extracellular vestibule with three lateral portals and the channel pore. Here we used cysteine and alanine scanning mutagenesis of the rat P2X4R receptor V47–V61 and K326–N338 sequences to study structural and functional properties of extracellular vestibule during gating. Cysteine mutants were used to test the accessibility of these residue side chains to cadmium during closed-open-desensitized transitions, whereas alanine mutants served as controls. This study revealed the accessibility of residues E51, T57, S59, V61, K326, and M336 to cadmium in channels undergoing a transition from a closed-to-open state and the accessibility of residues V47, G53, D331, I332, I333, T335, I337, and N338 in channels undergoing a transition from an open-to-desensitized state; residues E56 and K329 were accessible during both transitions. The effect of cadmium on channel gating was stimulatory in all reactive V47–V61 mutants and inhibitory in the majority of reactive K326–N338 mutants. The rat P2X4 receptor homology model suggests that residues affected by cadmium in the closed-to-open transition were located within the lumen of the extracellular vestibule and toward the central vestibule; however, the residues affected by cadmium in the open-to-desensitized state were located at the bottom of the vestibule near the pore. Analysis of the model assumed that there is ion access to extracellular and central vestibules through lateral ports when the channel is closed, with residues above the first transmembrane domain being predominantly responsible for ion uptake. Upon receptor activation, there is passage of ions toward the residues located on the upper region of the second transmembrane domain, followed by permeation through the gate region. Frontiers Media S.A. 2014-01-29 /pmc/articles/PMC3905210/ /pubmed/24523669 http://dx.doi.org/10.3389/fncel.2014.00003 Text en Copyright © 2014 Rokic, Stojilkovic and Zemkova. http://creativecommons.org/licenses/by/3.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
spellingShingle Neuroscience
Rokic, Milos B.
Stojilkovic, Stanko S.
Zemkova, Hana
Structural and functional properties of the rat P2X4 purinoreceptor extracellular vestibule during gating
title Structural and functional properties of the rat P2X4 purinoreceptor extracellular vestibule during gating
title_full Structural and functional properties of the rat P2X4 purinoreceptor extracellular vestibule during gating
title_fullStr Structural and functional properties of the rat P2X4 purinoreceptor extracellular vestibule during gating
title_full_unstemmed Structural and functional properties of the rat P2X4 purinoreceptor extracellular vestibule during gating
title_short Structural and functional properties of the rat P2X4 purinoreceptor extracellular vestibule during gating
title_sort structural and functional properties of the rat p2x4 purinoreceptor extracellular vestibule during gating
topic Neuroscience
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3905210/
https://www.ncbi.nlm.nih.gov/pubmed/24523669
http://dx.doi.org/10.3389/fncel.2014.00003
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