In Vivo Intracellular pH Measurements in Tobacco and Arabidopsis Reveal an Unexpected pH Gradient in the Endomembrane System |
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| Authors: | Alexandre Martinière Elias Bassil Elodie Jublanc Carine Alcon Maria Reguera Hervé Sentenac Eduardo Blumwald Nadine Paris |
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| Affiliation: | aBiochemistry and Plant Molecular Biology Lab, Unité Mixte de Recherche 5004, 34060 Montpellier, France;bDepartment of Plant Sciences, University of California, Davis, California 95616;cInstitut National de la Recherche Agronomique, Unité Mixte de Recherche 866, Dynamique Musculaire et Métabolisme, 34060 Montpellier, France |
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| Abstract: | The pH homeostasis of endomembranes is essential for cellular functions. In order to provide direct pH measurements in the endomembrane system lumen, we targeted genetically encoded ratiometric pH sensors to the cytosol, the endoplasmic reticulum, and the trans-Golgi, or the compartments labeled by the vacuolar sorting receptor (VSR), which includes the trans-Golgi network and prevacuoles. Using noninvasive live-cell imaging to measure pH, we show that a gradual acidification from the endoplasmic reticulum to the lytic vacuole exists, in both tobacco (Nicotiana tabacum) epidermal (ΔpH −1.5) and Arabidopsis thaliana root cells (ΔpH −2.1). The average pH in VSR compartments was intermediate between that of the trans-Golgi and the vacuole. Combining pH measurements with in vivo colocalization experiments, we found that the trans-Golgi network had an acidic pH of 6.1, while the prevacuole and late prevacuole were both more alkaline, with pH of 6.6 and 7.1, respectively. We also showed that endosomal pH, and subsequently vacuolar trafficking of soluble proteins, requires both vacuolar-type H+ ATPase–dependent acidification as well as proton efflux mediated at least by the activity of endosomal sodium/proton NHX-type antiporters. |
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