Relative positioning of Kv11.1 (hERG) K+ channel cytoplasmic domain-located fluorescent tags toward the plasma membrane

Abstract Recent cryo-EM data have provided a view of the KCNH potassium channels molecular structures. However, some details about the cytoplasmic domains organization and specially their rearrangements associated to channel functionality are still lacking. Here we used the voltage-dependent dipicry...

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Autores principales:	Francisco Barros, Pedro Domínguez, Pilar de la Peña
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2018
Materias:	hERG Channel Proximal Domain HEKA Elektronic Long Depolarizing Hyperpolarization-activated Cyclic Nucleotide-gated (HCN) Medicine R Science Q
Acceso en línea:	https://doaj.org/article/dbdbb6a14e064facaba483b71d6a2752
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spelling	oai:doaj.org-article:dbdbb6a14e064facaba483b71d6a27522021-12-02T15:08:24ZRelative positioning of Kv11.1 (hERG) K+ channel cytoplasmic domain-located fluorescent tags toward the plasma membrane10.1038/s41598-018-33492-x2045-2322https://doaj.org/article/dbdbb6a14e064facaba483b71d6a27522018-10-01T00:00:00Zhttps://doi.org/10.1038/s41598-018-33492-xhttps://doaj.org/toc/2045-2322Abstract Recent cryo-EM data have provided a view of the KCNH potassium channels molecular structures. However, some details about the cytoplasmic domains organization and specially their rearrangements associated to channel functionality are still lacking. Here we used the voltage-dependent dipicrylamine (DPA)-induced quench of fluorescent proteins (FPS) linked to different positions at the cytoplasmic domains of KCNH2 (hERG) to gain some insights about the coarse structure of these channel parts. Fast voltage-clamp fluorometry with HEK293 cells expressing membrane-anchored FPs under conditions in which only the plasma membrane potential is modified, demonstrated DPA voltage-dependent translocation and subsequent FRET-triggered FP quenching. Our data demonstrate for the first time that the distance between an amino-terminal FP tag and the intracellular plasma membrane surface is shorter than that between the membrane and a C-terminally-located tag. The distances varied when the FPs were attached to other positions along the channel cytoplasmic domains. In some cases, we also detected slower fluorometric responses following the fast voltage-dependent dye translocation, indicating subsequent label movements orthogonal to the plasma membrane. This finding suggests the existence of additional conformational rearrangements in the hERG cytoplasmic domains, although their association with specific aspects of channel operation remains to be established.Francisco BarrosPedro DomínguezPilar de la PeñaNature PortfolioarticlehERG ChannelProximal DomainHEKA ElektronicLong DepolarizingHyperpolarization-activated Cyclic Nucleotide-gated (HCN)MedicineRScienceQENScientific Reports, Vol 8, Iss 1, Pp 1-14 (2018)
institution	DOAJ
collection	DOAJ
language	EN
topic	hERG Channel Proximal Domain HEKA Elektronic Long Depolarizing Hyperpolarization-activated Cyclic Nucleotide-gated (HCN) Medicine R Science Q
spellingShingle	hERG Channel Proximal Domain HEKA Elektronic Long Depolarizing Hyperpolarization-activated Cyclic Nucleotide-gated (HCN) Medicine R Science Q Francisco Barros Pedro Domínguez Pilar de la Peña Relative positioning of Kv11.1 (hERG) K+ channel cytoplasmic domain-located fluorescent tags toward the plasma membrane
description	Abstract Recent cryo-EM data have provided a view of the KCNH potassium channels molecular structures. However, some details about the cytoplasmic domains organization and specially their rearrangements associated to channel functionality are still lacking. Here we used the voltage-dependent dipicrylamine (DPA)-induced quench of fluorescent proteins (FPS) linked to different positions at the cytoplasmic domains of KCNH2 (hERG) to gain some insights about the coarse structure of these channel parts. Fast voltage-clamp fluorometry with HEK293 cells expressing membrane-anchored FPs under conditions in which only the plasma membrane potential is modified, demonstrated DPA voltage-dependent translocation and subsequent FRET-triggered FP quenching. Our data demonstrate for the first time that the distance between an amino-terminal FP tag and the intracellular plasma membrane surface is shorter than that between the membrane and a C-terminally-located tag. The distances varied when the FPs were attached to other positions along the channel cytoplasmic domains. In some cases, we also detected slower fluorometric responses following the fast voltage-dependent dye translocation, indicating subsequent label movements orthogonal to the plasma membrane. This finding suggests the existence of additional conformational rearrangements in the hERG cytoplasmic domains, although their association with specific aspects of channel operation remains to be established.
format	article
author	Francisco Barros Pedro Domínguez Pilar de la Peña
author_facet	Francisco Barros Pedro Domínguez Pilar de la Peña
author_sort	Francisco Barros
title	Relative positioning of Kv11.1 (hERG) K+ channel cytoplasmic domain-located fluorescent tags toward the plasma membrane
title_short	Relative positioning of Kv11.1 (hERG) K+ channel cytoplasmic domain-located fluorescent tags toward the plasma membrane
title_full	Relative positioning of Kv11.1 (hERG) K+ channel cytoplasmic domain-located fluorescent tags toward the plasma membrane
title_fullStr	Relative positioning of Kv11.1 (hERG) K+ channel cytoplasmic domain-located fluorescent tags toward the plasma membrane
title_full_unstemmed	Relative positioning of Kv11.1 (hERG) K+ channel cytoplasmic domain-located fluorescent tags toward the plasma membrane
title_sort	relative positioning of kv11.1 (herg) k+ channel cytoplasmic domain-located fluorescent tags toward the plasma membrane
publisher	Nature Portfolio
publishDate	2018
url	https://doaj.org/article/dbdbb6a14e064facaba483b71d6a2752
work_keys_str_mv	AT franciscobarros relativepositioningofkv111hergkchannelcytoplasmicdomainlocatedfluorescenttagstowardtheplasmamembrane AT pedrodominguez relativepositioningofkv111hergkchannelcytoplasmicdomainlocatedfluorescenttagstowardtheplasmamembrane AT pilardelapena relativepositioningofkv111hergkchannelcytoplasmicdomainlocatedfluorescenttagstowardtheplasmamembrane
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Relative positioning of Kv11.1 (hERG) K+ channel cytoplasmic domain-located fluorescent tags toward the plasma membrane

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