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Permeant calcium ion feed-through regulation of single inositol 1,4,5-trisphosphate receptor channel gating
The ubiquitous inositol 1,4,5-trisphosphate (InsP(3)) receptor (InsP(3)R) Ca(2+) release channel plays a central role in the generation and modulation of intracellular Ca(2+) signals, and is intricately regulated by multiple mechanisms including cytoplasmic ligand (InsP(3), free Ca(2+), free ATP(4−)...
Autores principales: | , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
The Rockefeller University Press
2012
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3514735/ https://www.ncbi.nlm.nih.gov/pubmed/23148262 http://dx.doi.org/10.1085/jgp.201210804 |
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author | Vais, Horia Foskett, J. Kevin Ullah, Ghanim Pearson, John E. Daniel Mak, Don-On |
author_facet | Vais, Horia Foskett, J. Kevin Ullah, Ghanim Pearson, John E. Daniel Mak, Don-On |
author_sort | Vais, Horia |
collection | PubMed |
description | The ubiquitous inositol 1,4,5-trisphosphate (InsP(3)) receptor (InsP(3)R) Ca(2+) release channel plays a central role in the generation and modulation of intracellular Ca(2+) signals, and is intricately regulated by multiple mechanisms including cytoplasmic ligand (InsP(3), free Ca(2+), free ATP(4−)) binding, posttranslational modifications, and interactions with cytoplasmic and endoplasmic reticulum (ER) luminal proteins. However, regulation of InsP(3)R channel activity by free Ca(2+) in the ER lumen ([Ca(2+)](ER)) remains poorly understood because of limitations of Ca(2+) flux measurements and imaging techniques. Here, we used nuclear patch-clamp experiments in excised luminal-side-out configuration with perfusion solution exchange to study the effects of [Ca(2+)](ER) on homotetrameric rat type 3 InsP(3)R channel activity. In optimal [Ca(2+)](i) and subsaturating [InsP(3)], jumps of [Ca(2+)](ER) from 70 nM to 300 µM reduced channel activity significantly. This inhibition was abrogated by saturating InsP(3) but restored when [Ca(2+)](ER) was raised to 1.1 mM. In suboptimal [Ca(2+)](i), jumps of [Ca(2+)](ER) (70 nM to 300 µM) enhanced channel activity. Thus, [Ca(2+)](ER) effects on channel activity exhibited a biphasic dependence on [Ca(2+)](i). In addition, the effect of high [Ca(2+)](ER) was attenuated when a voltage was applied to oppose Ca(2+) flux through the channel. These observations can be accounted for by Ca(2+) flux driven through the open InsP(3)R channel by [Ca(2+)](ER), raising local [Ca(2+)](i) around the channel to regulate its activity through its cytoplasmic regulatory Ca(2+)-binding sites. Importantly, [Ca(2+)](ER) regulation of InsP(3)R channel activity depended on cytoplasmic Ca(2+)-buffering conditions: it was more pronounced when [Ca(2+)](i) was weakly buffered but completely abolished in strong Ca(2+)-buffering conditions. With strong cytoplasmic buffering and Ca(2+) flux sufficiently reduced by applied voltage, both activation and inhibition of InsP(3)R channel gating by physiological levels of [Ca(2+)](ER) were completely abolished. Collectively, these results rule out Ca(2+) regulation of channel activity by direct binding to the luminal aspect of the channel. |
format | Online Article Text |
id | pubmed-3514735 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2012 |
publisher | The Rockefeller University Press |
record_format | MEDLINE/PubMed |
spelling | pubmed-35147352013-06-01 Permeant calcium ion feed-through regulation of single inositol 1,4,5-trisphosphate receptor channel gating Vais, Horia Foskett, J. Kevin Ullah, Ghanim Pearson, John E. Daniel Mak, Don-On J Gen Physiol Article The ubiquitous inositol 1,4,5-trisphosphate (InsP(3)) receptor (InsP(3)R) Ca(2+) release channel plays a central role in the generation and modulation of intracellular Ca(2+) signals, and is intricately regulated by multiple mechanisms including cytoplasmic ligand (InsP(3), free Ca(2+), free ATP(4−)) binding, posttranslational modifications, and interactions with cytoplasmic and endoplasmic reticulum (ER) luminal proteins. However, regulation of InsP(3)R channel activity by free Ca(2+) in the ER lumen ([Ca(2+)](ER)) remains poorly understood because of limitations of Ca(2+) flux measurements and imaging techniques. Here, we used nuclear patch-clamp experiments in excised luminal-side-out configuration with perfusion solution exchange to study the effects of [Ca(2+)](ER) on homotetrameric rat type 3 InsP(3)R channel activity. In optimal [Ca(2+)](i) and subsaturating [InsP(3)], jumps of [Ca(2+)](ER) from 70 nM to 300 µM reduced channel activity significantly. This inhibition was abrogated by saturating InsP(3) but restored when [Ca(2+)](ER) was raised to 1.1 mM. In suboptimal [Ca(2+)](i), jumps of [Ca(2+)](ER) (70 nM to 300 µM) enhanced channel activity. Thus, [Ca(2+)](ER) effects on channel activity exhibited a biphasic dependence on [Ca(2+)](i). In addition, the effect of high [Ca(2+)](ER) was attenuated when a voltage was applied to oppose Ca(2+) flux through the channel. These observations can be accounted for by Ca(2+) flux driven through the open InsP(3)R channel by [Ca(2+)](ER), raising local [Ca(2+)](i) around the channel to regulate its activity through its cytoplasmic regulatory Ca(2+)-binding sites. Importantly, [Ca(2+)](ER) regulation of InsP(3)R channel activity depended on cytoplasmic Ca(2+)-buffering conditions: it was more pronounced when [Ca(2+)](i) was weakly buffered but completely abolished in strong Ca(2+)-buffering conditions. With strong cytoplasmic buffering and Ca(2+) flux sufficiently reduced by applied voltage, both activation and inhibition of InsP(3)R channel gating by physiological levels of [Ca(2+)](ER) were completely abolished. Collectively, these results rule out Ca(2+) regulation of channel activity by direct binding to the luminal aspect of the channel. The Rockefeller University Press 2012-12 /pmc/articles/PMC3514735/ /pubmed/23148262 http://dx.doi.org/10.1085/jgp.201210804 Text en © 2012 Vais et al. 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 | Article Vais, Horia Foskett, J. Kevin Ullah, Ghanim Pearson, John E. Daniel Mak, Don-On Permeant calcium ion feed-through regulation of single inositol 1,4,5-trisphosphate receptor channel gating |
title | Permeant calcium ion feed-through regulation of single inositol
1,4,5-trisphosphate receptor channel gating |
title_full | Permeant calcium ion feed-through regulation of single inositol
1,4,5-trisphosphate receptor channel gating |
title_fullStr | Permeant calcium ion feed-through regulation of single inositol
1,4,5-trisphosphate receptor channel gating |
title_full_unstemmed | Permeant calcium ion feed-through regulation of single inositol
1,4,5-trisphosphate receptor channel gating |
title_short | Permeant calcium ion feed-through regulation of single inositol
1,4,5-trisphosphate receptor channel gating |
title_sort | permeant calcium ion feed-through regulation of single inositol
1,4,5-trisphosphate receptor channel gating |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3514735/ https://www.ncbi.nlm.nih.gov/pubmed/23148262 http://dx.doi.org/10.1085/jgp.201210804 |
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