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https://hdl.handle.net/2440/139886
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Type: | Journal article |
Title: | A new method to measure aquaporin-facilitated membrane diffusion of hydrogen peroxide and cations in plant suspension cells |
Author: | Ahmed, J. Ismail, A. Ding, L. Yool, A.J. Chaumont, F. |
Citation: | Plant, Cell and Environment, 2024; 47(2):527-539 |
Publisher: | Wiley |
Issue Date: | 2024 |
ISSN: | 0140-7791 1365-3040 |
Statement of Responsibility: | Jahed Ahmed, Ahmed Ismail, Lei Ding, Andrea J. Yool, François Chaumont |
Abstract: | Plant aquaporins (AQPs) facilitate the membrane diffusion of water and small solutes, including hydrogen peroxide (H2 O2 ) and, possibly, cations, essential signalling molecules in many physiological processes. While the determination of the channel activity generally depends on heterologous expression of AQPs in Xenopus oocytes or yeast cells, we established a genetic tool to determine whether they facilitate the diffusion of H2 O2 through the plasma membrane in living plant cells. We designed genetic constructs to co-express the fluorescent H2 O2 sensor HyPer and AQPs, with expression controlled by a heat shock-inducible promoter in Nicotiana tabacum BY-2 suspension cells. After induction of ZmPIP2;5 AQP expression, a HyPer signal was recorded when the cells were incubated with H2 O2 , suggesting that ZmPIP2;5 facilitates H2 O2 transmembrane diffusion; in contrast, the ZmPIP2;5W85A mutated protein was inactive as a water or H2 O2 channel. ZmPIP2;1, ZmPIP2;4 and AtPIP2;1 also facilitated H2 O2 diffusion. Incubation with abscisic acid and the elicitor flg22 peptide induced the intracellular H2 O2 accumulation in BY-2 cells expressing ZmPIP2;5. We also monitored cation channel activity of ZmPIP2;5 using a novel fluorescent photo-switchable Li+ sensor in BY-2 cells. BY-2 suspension cells engineered for inducible expression of AQPs as well as HyPer expression and the use of Li+ sensors constitute a powerful toolkit for evaluating the transport activity and the molecular determinants of PIPs in living plant cells. |
Keywords: | HyPer biosensor Li+ sensor live cell imaging plasma membrane intrinsic protein (PIP) tobacco BY-2 suspension cells |
Description: | First published: 09 November 2023 |
Rights: | © 2023 John Wiley & Sons Ltd. |
DOI: | 10.1111/pce.14763 |
Grant ID: | http://purl.org/au-research/grants/arc/DP190101745 |
Published version: | http://dx.doi.org/10.1111/pce.14763 |
Appears in Collections: | Physiology publications |
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