Cargando…

Conformational decoupling in acid-sensing ion channels uncovers mechanism and stoichiometry of PcTx1-mediated inhibition

Acid-sensing ion channels (ASICs) are trimeric proton-gated cation channels involved in fast synaptic transmission. Pharmacological inhibition of ASIC1a reduces neurotoxicity and stroke infarct volumes, with the cysteine knot toxin psalmotoxin-1 (PcTx1) being one of the most potent and selective inh...

Descripción completa

Detalles Bibliográficos
Autores principales: Heusser, Stephanie A, Borg, Christian B, Colding, Janne M, Pless, Stephan A
Formato: Online Artículo Texto
Lenguaje:English
Publicado: eLife Sciences Publications, Ltd 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8871370/
https://www.ncbi.nlm.nih.gov/pubmed/35156612
http://dx.doi.org/10.7554/eLife.73384
_version_ 1784656980610121728
author Heusser, Stephanie A
Borg, Christian B
Colding, Janne M
Pless, Stephan A
author_facet Heusser, Stephanie A
Borg, Christian B
Colding, Janne M
Pless, Stephan A
author_sort Heusser, Stephanie A
collection PubMed
description Acid-sensing ion channels (ASICs) are trimeric proton-gated cation channels involved in fast synaptic transmission. Pharmacological inhibition of ASIC1a reduces neurotoxicity and stroke infarct volumes, with the cysteine knot toxin psalmotoxin-1 (PcTx1) being one of the most potent and selective inhibitors. PcTx1 binds at the subunit interface in the extracellular domain (ECD), but the mechanism and conformational consequences of the interaction, as well as the number of toxin molecules required for inhibition, remain unknown. Here, we use voltage-clamp fluorometry and subunit concatenation to decipher the mechanism and stoichiometry of PcTx1 inhibition of ASIC1a. Besides the known inhibitory binding mode, we propose PcTx1 to have at least two additional binding modes that are decoupled from the pore. One of these modes induces a long-lived ECD conformation that reduces the activity of an endogenous neuropeptide. This long-lived conformational state is proton-dependent and can be destabilized by a mutation that decreases PcTx1 sensitivity. Lastly, the use of concatemeric channel constructs reveals that disruption of a single PcTx1 binding site is sufficient to destabilize the toxin-induced conformation, while functional inhibition is not impaired until two or more binding sites are mutated. Together, our work provides insight into the mechanism of PcTx1 inhibition of ASICs and uncovers a prolonged conformational change with possible pharmacological implications.
format Online
Article
Text
id pubmed-8871370
institution National Center for Biotechnology Information
language English
publishDate 2022
publisher eLife Sciences Publications, Ltd
record_format MEDLINE/PubMed
spelling pubmed-88713702022-02-25 Conformational decoupling in acid-sensing ion channels uncovers mechanism and stoichiometry of PcTx1-mediated inhibition Heusser, Stephanie A Borg, Christian B Colding, Janne M Pless, Stephan A eLife Structural Biology and Molecular Biophysics Acid-sensing ion channels (ASICs) are trimeric proton-gated cation channels involved in fast synaptic transmission. Pharmacological inhibition of ASIC1a reduces neurotoxicity and stroke infarct volumes, with the cysteine knot toxin psalmotoxin-1 (PcTx1) being one of the most potent and selective inhibitors. PcTx1 binds at the subunit interface in the extracellular domain (ECD), but the mechanism and conformational consequences of the interaction, as well as the number of toxin molecules required for inhibition, remain unknown. Here, we use voltage-clamp fluorometry and subunit concatenation to decipher the mechanism and stoichiometry of PcTx1 inhibition of ASIC1a. Besides the known inhibitory binding mode, we propose PcTx1 to have at least two additional binding modes that are decoupled from the pore. One of these modes induces a long-lived ECD conformation that reduces the activity of an endogenous neuropeptide. This long-lived conformational state is proton-dependent and can be destabilized by a mutation that decreases PcTx1 sensitivity. Lastly, the use of concatemeric channel constructs reveals that disruption of a single PcTx1 binding site is sufficient to destabilize the toxin-induced conformation, while functional inhibition is not impaired until two or more binding sites are mutated. Together, our work provides insight into the mechanism of PcTx1 inhibition of ASICs and uncovers a prolonged conformational change with possible pharmacological implications. eLife Sciences Publications, Ltd 2022-02-14 /pmc/articles/PMC8871370/ /pubmed/35156612 http://dx.doi.org/10.7554/eLife.73384 Text en © 2022, Heusser et al https://creativecommons.org/licenses/by/4.0/This article is distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use and redistribution provided that the original author and source are credited.
spellingShingle Structural Biology and Molecular Biophysics
Heusser, Stephanie A
Borg, Christian B
Colding, Janne M
Pless, Stephan A
Conformational decoupling in acid-sensing ion channels uncovers mechanism and stoichiometry of PcTx1-mediated inhibition
title Conformational decoupling in acid-sensing ion channels uncovers mechanism and stoichiometry of PcTx1-mediated inhibition
title_full Conformational decoupling in acid-sensing ion channels uncovers mechanism and stoichiometry of PcTx1-mediated inhibition
title_fullStr Conformational decoupling in acid-sensing ion channels uncovers mechanism and stoichiometry of PcTx1-mediated inhibition
title_full_unstemmed Conformational decoupling in acid-sensing ion channels uncovers mechanism and stoichiometry of PcTx1-mediated inhibition
title_short Conformational decoupling in acid-sensing ion channels uncovers mechanism and stoichiometry of PcTx1-mediated inhibition
title_sort conformational decoupling in acid-sensing ion channels uncovers mechanism and stoichiometry of pctx1-mediated inhibition
topic Structural Biology and Molecular Biophysics
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8871370/
https://www.ncbi.nlm.nih.gov/pubmed/35156612
http://dx.doi.org/10.7554/eLife.73384
work_keys_str_mv AT heusserstephaniea conformationaldecouplinginacidsensingionchannelsuncoversmechanismandstoichiometryofpctx1mediatedinhibition
AT borgchristianb conformationaldecouplinginacidsensingionchannelsuncoversmechanismandstoichiometryofpctx1mediatedinhibition
AT coldingjannem conformationaldecouplinginacidsensingionchannelsuncoversmechanismandstoichiometryofpctx1mediatedinhibition
AT plessstephana conformationaldecouplinginacidsensingionchannelsuncoversmechanismandstoichiometryofpctx1mediatedinhibition