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Reactive oxygen species produced by NADPH oxidase regulate plant cell growth

Abstract

Cell expansion is a central process in plant morphogenesis, and the elongation of roots and root hairs is essential for uptake of minerals and water from the soil. Ca2+ influx from the extracellular store is required for (and sets the rates of) cell elongation in roots1. Arabidopsis thaliana rhd2 mutants are defective in Ca2+ uptake and consequently cell expansion is compromised—rhd2 mutants have short root hairs2,3 and stunted roots. To determine the regulation of Ca2+ acquisition in growing root cells we show here that RHD2 is an NADPH oxidase, a protein that transfers electrons from NADPH to an electron acceptor leading to the formation of reactive oxygen species (ROS). We show that ROS accumulate in growing wild-type (WT) root hairs but their levels are markedly decreased in rhd2 mutants. Blocking the activity of the NADPH oxidase with diphenylene iodonium (DPI) inhibits ROS formation and phenocopies Rhd2-. Treatment of rhd2 roots with ROS partly suppresses the mutant phenotype and stimulates the activity of plasma membrane hyperpolarization-activated Ca2+ channels, the predominant root Ca2+ acquisition system. This indicates that NADPH oxidases control development by making ROS that regulate plant cell expansion through the activation of Ca2+ channels.

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Figure 1: Characterization of RHD2/AtrbohC gene.
Figure 2: ROS accumulation (CM-H2DCF imaging) during root-hair elongation: transmission (top) and pseudocolour fluorescent images (bottom) are displayed for WT (a), rhd2 (b) and WT (c) after treatment with DPI.
Figure 3: ROS elevates [Ca2+]c in an rhd2 root-hair bulge.
Figure 4: Activation of hyperpolarization-activated Ca2+ channels by ROS.

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Acknowledgements

We thank D. Graham and A. Dark for help with screening and cultivation, respectively; E. Ryan, P. Shaw and K. Roberts for comments on the manuscript; and P. Doerner for support. This work was funded by the BBSRC, the Gatsby Foundation, the Leverhulme Trust and the European Union.

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Correspondence to Liam Dolan.

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Foreman, J., Demidchik, V., Bothwell, J. et al. Reactive oxygen species produced by NADPH oxidase regulate plant cell growth. Nature 422, 442–446 (2003). https://doi.org/10.1038/nature01485

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