Pore collapse underlies irreversible inactivation of TRPM2 cation channel currents

Balázs Tóth, L. Csanády

Research output: Contribution to journalArticle

31 Citations (Scopus)

Abstract

The Ca 2+-permeable cation channel transient receptor potential melastatin 2 (TRPM2) plays a key role in pathogen-evoked phagocyte activation, postischemic neuronal apoptosis, and glucoseevoked insulin secretion, by linking these cellular responses to oxidative stress. TRPM2 channels are coactivated by binding of intracellular ADP ribose and Ca 2+ to distinct cytosolically accessible sites on the channels. These ligands likely regulate the activation gate, conserved in the voltage-gated cation channel superfamily, that comprises a helix bundle formed by the intracellular ends of transmembrane helix six of each subunit. For several K + and TRPM family channels, activation gate opening requires the presence of phosphatidylinositol- bisphosphate (PIP 2) in the inner membrane leaflet. Most TRPM family channels inactivate upon prolonged stimulation in inside-out patches; this "rundown" is due to PIP 2depletion. TRPM2 currents also run down within minutes, but the molecular mechanism of this process is unknown. Here we report that high-affinity PIP 2 binding regulates Ca 2+ sensitivity of TRPM2 activation. Nevertheless, TRPM2 inactivation is not due to PIP 2 depletion; rather, it is state dependent, sensitive to permeating ions, and can be completely prevented by mutations in the extracellular selectivity filter. Introduction of two negative charges plus a single-residue insertion, to mimic the filter sequence of TRPM5, results in TRPM2 channels that maintain unabated maximal activity for over 1 h, and display altered permeation properties but intact ADP ribose/Ca 2+-dependent gating. Thus, upon prolonged stimulation, the TRPM2 selectivity filter undergoes a conformational change reminiscent of that accompanying C-type inactivation of voltage-gated K + channels. The noninactivating TRPM2 variant will be invaluable for gating studies.

Original languageEnglish
Pages (from-to)13440-13445
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume109
Issue number33
DOIs
Publication statusPublished - Aug 14 2012

Fingerprint

Adenosine Diphosphate Ribose
Cations
Transient Receptor Potential Channels
Voltage-Gated Potassium Channels
Phagocytes
Phosphatidylinositols
Oxidative Stress
Ions
Insulin
Apoptosis
Ligands
Mutation
Membranes

Keywords

  • C-type inactivation
  • Cation permeation
  • Electrostatic effect
  • Gating kinetics
  • Pi(4,5)P

ASJC Scopus subject areas

  • General

Cite this

Pore collapse underlies irreversible inactivation of TRPM2 cation channel currents. / Tóth, Balázs; Csanády, L.

In: Proceedings of the National Academy of Sciences of the United States of America, Vol. 109, No. 33, 14.08.2012, p. 13440-13445.

Research output: Contribution to journalArticle

@article{a7c758760e7149c39f9210d74c3951a6,
title = "Pore collapse underlies irreversible inactivation of TRPM2 cation channel currents",
abstract = "The Ca 2+-permeable cation channel transient receptor potential melastatin 2 (TRPM2) plays a key role in pathogen-evoked phagocyte activation, postischemic neuronal apoptosis, and glucoseevoked insulin secretion, by linking these cellular responses to oxidative stress. TRPM2 channels are coactivated by binding of intracellular ADP ribose and Ca 2+ to distinct cytosolically accessible sites on the channels. These ligands likely regulate the activation gate, conserved in the voltage-gated cation channel superfamily, that comprises a helix bundle formed by the intracellular ends of transmembrane helix six of each subunit. For several K + and TRPM family channels, activation gate opening requires the presence of phosphatidylinositol- bisphosphate (PIP 2) in the inner membrane leaflet. Most TRPM family channels inactivate upon prolonged stimulation in inside-out patches; this {"}rundown{"} is due to PIP 2depletion. TRPM2 currents also run down within minutes, but the molecular mechanism of this process is unknown. Here we report that high-affinity PIP 2 binding regulates Ca 2+ sensitivity of TRPM2 activation. Nevertheless, TRPM2 inactivation is not due to PIP 2 depletion; rather, it is state dependent, sensitive to permeating ions, and can be completely prevented by mutations in the extracellular selectivity filter. Introduction of two negative charges plus a single-residue insertion, to mimic the filter sequence of TRPM5, results in TRPM2 channels that maintain unabated maximal activity for over 1 h, and display altered permeation properties but intact ADP ribose/Ca 2+-dependent gating. Thus, upon prolonged stimulation, the TRPM2 selectivity filter undergoes a conformational change reminiscent of that accompanying C-type inactivation of voltage-gated K + channels. The noninactivating TRPM2 variant will be invaluable for gating studies.",
keywords = "C-type inactivation, Cation permeation, Electrostatic effect, Gating kinetics, Pi(4,5)P",
author = "Bal{\'a}zs T{\'o}th and L. Csan{\'a}dy",
year = "2012",
month = "8",
day = "14",
doi = "10.1073/pnas.1204702109",
language = "English",
volume = "109",
pages = "13440--13445",
journal = "Proceedings of the National Academy of Sciences of the United States of America",
issn = "0027-8424",
number = "33",

}

TY - JOUR

T1 - Pore collapse underlies irreversible inactivation of TRPM2 cation channel currents

AU - Tóth, Balázs

AU - Csanády, L.

PY - 2012/8/14

Y1 - 2012/8/14

N2 - The Ca 2+-permeable cation channel transient receptor potential melastatin 2 (TRPM2) plays a key role in pathogen-evoked phagocyte activation, postischemic neuronal apoptosis, and glucoseevoked insulin secretion, by linking these cellular responses to oxidative stress. TRPM2 channels are coactivated by binding of intracellular ADP ribose and Ca 2+ to distinct cytosolically accessible sites on the channels. These ligands likely regulate the activation gate, conserved in the voltage-gated cation channel superfamily, that comprises a helix bundle formed by the intracellular ends of transmembrane helix six of each subunit. For several K + and TRPM family channels, activation gate opening requires the presence of phosphatidylinositol- bisphosphate (PIP 2) in the inner membrane leaflet. Most TRPM family channels inactivate upon prolonged stimulation in inside-out patches; this "rundown" is due to PIP 2depletion. TRPM2 currents also run down within minutes, but the molecular mechanism of this process is unknown. Here we report that high-affinity PIP 2 binding regulates Ca 2+ sensitivity of TRPM2 activation. Nevertheless, TRPM2 inactivation is not due to PIP 2 depletion; rather, it is state dependent, sensitive to permeating ions, and can be completely prevented by mutations in the extracellular selectivity filter. Introduction of two negative charges plus a single-residue insertion, to mimic the filter sequence of TRPM5, results in TRPM2 channels that maintain unabated maximal activity for over 1 h, and display altered permeation properties but intact ADP ribose/Ca 2+-dependent gating. Thus, upon prolonged stimulation, the TRPM2 selectivity filter undergoes a conformational change reminiscent of that accompanying C-type inactivation of voltage-gated K + channels. The noninactivating TRPM2 variant will be invaluable for gating studies.

AB - The Ca 2+-permeable cation channel transient receptor potential melastatin 2 (TRPM2) plays a key role in pathogen-evoked phagocyte activation, postischemic neuronal apoptosis, and glucoseevoked insulin secretion, by linking these cellular responses to oxidative stress. TRPM2 channels are coactivated by binding of intracellular ADP ribose and Ca 2+ to distinct cytosolically accessible sites on the channels. These ligands likely regulate the activation gate, conserved in the voltage-gated cation channel superfamily, that comprises a helix bundle formed by the intracellular ends of transmembrane helix six of each subunit. For several K + and TRPM family channels, activation gate opening requires the presence of phosphatidylinositol- bisphosphate (PIP 2) in the inner membrane leaflet. Most TRPM family channels inactivate upon prolonged stimulation in inside-out patches; this "rundown" is due to PIP 2depletion. TRPM2 currents also run down within minutes, but the molecular mechanism of this process is unknown. Here we report that high-affinity PIP 2 binding regulates Ca 2+ sensitivity of TRPM2 activation. Nevertheless, TRPM2 inactivation is not due to PIP 2 depletion; rather, it is state dependent, sensitive to permeating ions, and can be completely prevented by mutations in the extracellular selectivity filter. Introduction of two negative charges plus a single-residue insertion, to mimic the filter sequence of TRPM5, results in TRPM2 channels that maintain unabated maximal activity for over 1 h, and display altered permeation properties but intact ADP ribose/Ca 2+-dependent gating. Thus, upon prolonged stimulation, the TRPM2 selectivity filter undergoes a conformational change reminiscent of that accompanying C-type inactivation of voltage-gated K + channels. The noninactivating TRPM2 variant will be invaluable for gating studies.

KW - C-type inactivation

KW - Cation permeation

KW - Electrostatic effect

KW - Gating kinetics

KW - Pi(4,5)P

UR - http://www.scopus.com/inward/record.url?scp=84865190336&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84865190336&partnerID=8YFLogxK

U2 - 10.1073/pnas.1204702109

DO - 10.1073/pnas.1204702109

M3 - Article

VL - 109

SP - 13440

EP - 13445

JO - Proceedings of the National Academy of Sciences of the United States of America

JF - Proceedings of the National Academy of Sciences of the United States of America

SN - 0027-8424

IS - 33

ER -