Voltage- and Ca²⁺-inducible PLC activity for analyzing PI(4,5)P₂ sensitivity of ion channels in Xenopus oocytes

Kawai, Takafumi; Mizutani, Natsuki; Okamura, Yasushi

Biochimica et Biophysica Acta (BBA) - Biomembranes, 2024, 1867(1), 184396

Number of Access:411（2025-07-18 10:06 Counts）

Identifier to cite or link to this item: https://hdl.handle.net/11094/98373

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Title
Voltage- and Ca²⁺-inducible PLC activity for analyzing PI(4,5)P₂ sensitivity of ion channels in Xenopus oocytes
Creator
NRID
1000070614915 1000070614915
researchmap permalink
t_kawa t_kawa
Scopus Author ID
55465095900 55465095900
CreatorName
Kawai, Takafumi
ORCID
0009-0005-9575-9961 0009-0005-9575-9961
researchmap permalink
zdl134skhf zdl134skhf
CreatorName
Mizutani, Natsuki
NRID
1000080201987 1000080201987
researchmap permalink
yasushiokamura yasushiokamura
Scopus Author ID
55322083200 55322083200
ORCID
0000-0001-5386-7968 0000-0001-5386-7968
CreatorName
Okamura, Yasushi
Subject
PLCζ
Ion channel
Membrane potential
Calcium
Xenopus oocyte
Description
Kawai Takafumi, Mizutani Natsuki, Okamura Yasushi. Voltage- and Ca²⁺-inducible PLC activity for analyzing PI(4,5)P₂ sensitivity of ion channels in Xenopus oocytes. Biochimica et Biophysica Acta (BBA) - Biomembranes 1867, 184396 (2024); https://doi.org/10.1016/j.bbamem.2024.184396.
Abstract
Phosphatidylinositol 4,5-bisphosphate (PIP₂) is a key membrane lipid regulating various ion channel activities. Currently, several molecular tools are used to modulate PIP₂ levels, each of which has distinct advantages and drawbacks. In this study, we proposed a novel methodology using heterologous Xenopus oocytes to precisely manipulate PIP₂ levels using phospholipase C (PLC)-ζ, which hydrolyzes PIP₂. Xenopus oocytes injected with PLCζ exhibited notable hyperpolarization-induced Ca²⁺ influx driven by the increased driving force of Ca²⁺. High Ca²⁺ sensitivity of PLCζ facilitated hyperpolarization-induced PLC activity in Xenopus oocytes that was voltage- and Ca²⁺-dependent. This study demonstrated the regulatory capacity of PLCζ in modulating PIP₂-sensitive ion channels, such as the KCNQ2/3 and GIRK channels, in a voltage- and Ca²⁺-dependent manner. Moreover, activation pathway of PLCζ only requires a two-electrode voltage clamp setup, making it a convenient molecular tool to manipulate PIP₂ levels in combination with a voltage-sensing phosphatase (VSP). PLCζ has distinct characteristics and advantages compared to VSP: (1) Hyperpolarization, but not depolarization, reduced the PIP₂ levels, (2) PIP₂ levels were decreased without any increase in phosphatidylinositol 4-monophosphate (PIP) levels, and (3) PIP₂ levels were reduced by Ca²⁺ administration. Therefore, PLCζ effectively supports understanding how PIP₂ regulates ion channels, alongside VSP. Overall, this study highlights the unique characteristics of PLCζ and its distinct advantages in analyzing ion channel regulation by PIP₂ and the PLC pathway in Xenopus oocytes.
Publisher
Elsevier BV
Source Title
Biochimica et Biophysica Acta (BBA) - Biomembranes
Volume (Issue)
1867(1)
Page
184396
Date of Issued
2024-11-05
Language
English
Handle URL
https://hdl.handle.net/11094/98373
PISSN
00052736
Relation.isIdenticalTo
DOI (Publisher Version)
https://doi.org/10.1016/j.bbamem.2024.184396
Access Rights
open access
CopyRight
This article is licensed under a Creative Commons Attribution 4.0 International License.
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Version of Record
Category
学術雑誌論文 Journal Article