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Real-time monitoring of subcellular H(2)O(2) distribution in Chlamydomonas reinhardtii

Hydrogen peroxide (H(2)O(2)) is recognized as an important signaling molecule in plants. We sought to establish a genetically encoded, fluorescent H(2)O(2) sensor that allows H(2)O(2) monitoring in all major subcompartments of a Chlamydomonas cell. To this end, we used the Chlamydomonas Modular Clon...

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Detalles Bibliográficos
Autores principales: Niemeyer, Justus, Scheuring, David, Oestreicher, Julian, Morgan, Bruce, Schroda, Michael
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Oxford University Press 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8462822/
https://www.ncbi.nlm.nih.gov/pubmed/34196712
http://dx.doi.org/10.1093/plcell/koab176
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author Niemeyer, Justus
Scheuring, David
Oestreicher, Julian
Morgan, Bruce
Schroda, Michael
author_facet Niemeyer, Justus
Scheuring, David
Oestreicher, Julian
Morgan, Bruce
Schroda, Michael
author_sort Niemeyer, Justus
collection PubMed
description Hydrogen peroxide (H(2)O(2)) is recognized as an important signaling molecule in plants. We sought to establish a genetically encoded, fluorescent H(2)O(2) sensor that allows H(2)O(2) monitoring in all major subcompartments of a Chlamydomonas cell. To this end, we used the Chlamydomonas Modular Cloning toolbox to target the hypersensitive H(2)O(2) sensor reduction–oxidation sensitive green fluorescent protein2-Tsa2ΔC(R) to the cytosol, nucleus, mitochondrial matrix, chloroplast stroma, thylakoid lumen, and endoplasmic reticulum (ER). The sensor was functional in all compartments, except for the ER where it was fully oxidized. Employing our novel sensors, we show that H(2)O(2) produced by photosynthetic linear electron transport (PET) in the stroma leaks into the cytosol but only reaches other subcellular compartments if produced under nonphysiological conditions. Furthermore, in heat-stressed cells, we show that cytosolic H(2)O(2) levels closely mirror temperature up- and downshifts and are independent from PET. Heat stress led to similar up- and downshifts of H(2)O(2) levels in the nucleus and, more mildly, in mitochondria but not in the chloroplast. Our results thus suggest the establishment of steep intracellular H(2)O(2) gradients under normal physiological conditions with limited diffusion into other compartments. We anticipate that these sensors will greatly facilitate future investigations of H(2)O(2) biology in plant cells.
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spelling pubmed-84628222021-09-27 Real-time monitoring of subcellular H(2)O(2) distribution in Chlamydomonas reinhardtii Niemeyer, Justus Scheuring, David Oestreicher, Julian Morgan, Bruce Schroda, Michael Plant Cell Breakthrough Report Hydrogen peroxide (H(2)O(2)) is recognized as an important signaling molecule in plants. We sought to establish a genetically encoded, fluorescent H(2)O(2) sensor that allows H(2)O(2) monitoring in all major subcompartments of a Chlamydomonas cell. To this end, we used the Chlamydomonas Modular Cloning toolbox to target the hypersensitive H(2)O(2) sensor reduction–oxidation sensitive green fluorescent protein2-Tsa2ΔC(R) to the cytosol, nucleus, mitochondrial matrix, chloroplast stroma, thylakoid lumen, and endoplasmic reticulum (ER). The sensor was functional in all compartments, except for the ER where it was fully oxidized. Employing our novel sensors, we show that H(2)O(2) produced by photosynthetic linear electron transport (PET) in the stroma leaks into the cytosol but only reaches other subcellular compartments if produced under nonphysiological conditions. Furthermore, in heat-stressed cells, we show that cytosolic H(2)O(2) levels closely mirror temperature up- and downshifts and are independent from PET. Heat stress led to similar up- and downshifts of H(2)O(2) levels in the nucleus and, more mildly, in mitochondria but not in the chloroplast. Our results thus suggest the establishment of steep intracellular H(2)O(2) gradients under normal physiological conditions with limited diffusion into other compartments. We anticipate that these sensors will greatly facilitate future investigations of H(2)O(2) biology in plant cells. Oxford University Press 2021-07-01 /pmc/articles/PMC8462822/ /pubmed/34196712 http://dx.doi.org/10.1093/plcell/koab176 Text en © The Author(s) 2021. Published by Oxford University Press on behalf of American Society of Plant Biologists. https://creativecommons.org/licenses/by/4.0/This is an Open Access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.
spellingShingle Breakthrough Report
Niemeyer, Justus
Scheuring, David
Oestreicher, Julian
Morgan, Bruce
Schroda, Michael
Real-time monitoring of subcellular H(2)O(2) distribution in Chlamydomonas reinhardtii
title Real-time monitoring of subcellular H(2)O(2) distribution in Chlamydomonas reinhardtii
title_full Real-time monitoring of subcellular H(2)O(2) distribution in Chlamydomonas reinhardtii
title_fullStr Real-time monitoring of subcellular H(2)O(2) distribution in Chlamydomonas reinhardtii
title_full_unstemmed Real-time monitoring of subcellular H(2)O(2) distribution in Chlamydomonas reinhardtii
title_short Real-time monitoring of subcellular H(2)O(2) distribution in Chlamydomonas reinhardtii
title_sort real-time monitoring of subcellular h(2)o(2) distribution in chlamydomonas reinhardtii
topic Breakthrough Report
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8462822/
https://www.ncbi.nlm.nih.gov/pubmed/34196712
http://dx.doi.org/10.1093/plcell/koab176
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