Skip to main content

Thank you for visiting You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

Reactive oxygen species produced by NADPH oxidase regulate plant cell growth


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.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.


All prices are NET prices.

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.


  1. 1

    Cramer, G. R. & Jones, R. L. Osmotic stress and abscisic acid reduce cytosolic calcium activities in roots of Arabidopsis thaliana. Plant Cell Environ. 19, 1291–1298 (1996)

    CAS  Article  Google Scholar 

  2. 2

    Wymer, C. L., Bibikova, T. N. & Gilroy, S. Cytoplasmic free calcium distributions during the development of root hairs of Arabidopsis thaliana. Plant J. 12, 427–439 (1997)

    CAS  Article  Google Scholar 

  3. 3

    Schiefelbein, J. W. & Somerville, C. Genetic control of root hair development in Arabidopsis thaliana. Plant Cell 2, 235–243 (1990)

    CAS  Article  Google Scholar 

  4. 4

    Carroll, A. D. et al. Ca2+, annexins, and GTP modulate exocytosis from maize root cap protoplasts. Plant Cell 10, 1267–1276 (1998)

    CAS  PubMed  PubMed Central  Google Scholar 

  5. 5

    Demidchik, V. et al. Arabidopsis thaliana root nonselective cation channels mediate calcium uptake and are involved in growth. Plant J. 32, 799–808 (2002)

    CAS  Article  Google Scholar 

  6. 6

    Kiegle, E., Gilliham, M., Haseloff, J. & Tester, M. Hyperpolarisation-activated calcium channels found only in cells from the elongation zone of Arabidopsis thaliana roots. Plant J. 21, 225–229 (2000)

    CAS  Article  Google Scholar 

  7. 7

    Véry, A.-A. & Davies, J. M. Hyperpolarization-activated calcium channels at the tip of Arabidopsis root hairs. Proc. Natl Acad. Sci. USA 97, 9801–9806 (2000)

    ADS  Article  Google Scholar 

  8. 8

    Miedema, H., Bothwell, J. H. F., Brownlee, C. & Davies, J. M. Calcium uptake by plant cells—channels and pumps acting in concert. Trends Plant Sci. 11, 514–519 (2001)

    Article  Google Scholar 

  9. 9

    Dolan, L. et al. Clonal relationships and cell patterning in the root epidermis of Arabidopsis. Development 120, 2465–2474 (1994)

    CAS  Google Scholar 

  10. 10

    Keller, T. et al. A plant homolog of the neutrophil NADPH oxidase gp91phox subunit gene encodes a plasma membrane protein with Ca2+ binding motifs. Plant Cell 10, 255–266 (1998)

    CAS  PubMed  PubMed Central  Google Scholar 

  11. 11

    Torres, M. A. et al. Six Arabidopsis thaliana homologues of the human respiratory burst oxidase (gp91phox). Plant J. 14, 365–370 (1998)

    CAS  Article  Google Scholar 

  12. 12

    Segal, A. W. & Abo, A. The biochemical basis of the NADPH oxidase of phagocytes. Trends Biochem. Sci. 18, 43–47 (1993)

    CAS  Article  Google Scholar 

  13. 13

    Torres, M. A., Dangl, J. L. & Jones, J. D. Arabidopsis gp91phox homologues AtrbohD and AtrbohF are required for accumulation of reactive oxygen intermediates in the plant defence response. Proc. Natl Acad. Sci. USA 99, 517–522 (2002)

    ADS  CAS  Article  Google Scholar 

  14. 14

    Williams, A. J. & Cole, P. J. Investigation of alveolar macrophage function using lucigenin-dependent chemiluminescence. Thorax 36, 866–869 (1981)

    CAS  Article  Google Scholar 

  15. 15

    Bolwell, G. P. & Wojtaszek, P. Mechanisms for the generation of reactive oxygen species in plant defence—a broad perspective. Physiol. Mol. Plant Pathol. 51, 347–366 (1997)

    CAS  Article  Google Scholar 

  16. 16

    Pei, Z. M. et al. Calcium channels activated by hydrogen peroxide mediate abscisic acid signalling in guard cells. Nature 406, 731–734 (2000)

    ADS  CAS  Article  Google Scholar 

  17. 17

    Fry, S. C. Oxidative scission of plant cell wall polysaccharides by ascorbate-induced hydroxyl radicals. Biochem. J. 332, 507–515 (1998)

    CAS  Article  Google Scholar 

  18. 18

    Halliwell, B. & Gutteridge, J. M. C. Free Radicals in Biology and Medicine (Oxford Univ. Press, Oxford, 1999)

    Google Scholar 

  19. 19

    Hirsch, R. E., Lewis, B. D., Spalding, E. P. & Sussman, M. R. A role for the AKT1 potassium channel in plant nutrition. Science 280, 918–921 (1998)

    ADS  CAS  Article  Google Scholar 

  20. 20

    Kiss, T. & Osipenko, O. N. Toxic effects of heavy metals on ionic channels. Pharmacol. Rev. 46, 245–267 (1994)

    CAS  PubMed  Google Scholar 

  21. 21

    Felle, H. & Hepler, P. K. The cytosolic Ca2+ concentration gradient of Sinapsis alba root hairs as revealed by Ca2+-selective microelectrode tests and fura-dextran ratio imaging. Plant Physiol. 114, 39–45 (1997)

    CAS  Article  Google Scholar 

  22. 22

    Molendijk, A. J. et al. Arabidopsis thaliana Rop GTPases are localized to tips of root hairs and control polar growth. EMBO J. 20, 2779–2788 (2001)

    CAS  Article  Google Scholar 

  23. 23

    Jones, M. A. et al. The Arabidopsis Rop2 GTPase is a positive regulator of both root hair initiation and tip growth. Plant Cell 14, 763–776 (2002)

    CAS  Article  Google Scholar 

  24. 24

    Tissier, A. F. et al. Multiple independent defective Suppressor-mutator transposon insertions in Arabidopsis: A tool for functional genomics. Plant Cell 11, 1841–1852 (1999)

    CAS  Article  Google Scholar 

  25. 25

    Coen, E. S. et al. floricaula: a homeotic gene required for flower development in Antirrhinum majus. Cell 63, 1311–1322 (1990)

    CAS  Article  Google Scholar 

  26. 26

    Altschul, S. F. et al. Gapped BLAST and PSI-BLAST: A new generation of protein database search programs. Nucleic Acids Res. 25, 3389–3402 (1997)

    CAS  Article  Google Scholar 

  27. 27

    Thompson, J. D., Higgins, D. G. & Gibson, T. J. CLUSTAL W: Improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Res. 22, 4673–4680 (1994)

    CAS  Article  Google Scholar 

  28. 28

    Page, R. D. M. TREEVIEW: An application to display phylogenetic trees on personal computers. Computer Applic. Biosci. 12, 357–358 (1996)

    CAS  Google Scholar 

  29. 29

    Zhang, W. H., Rengel, Z. & Kuo, J. Determination of intracellular Ca2+ in cells of intact wheat roots: loading of acetoxymethyl ester of Fluo-3 under low temperature. Plant J. 15, 147–151 (1998)

    Article  Google Scholar 

Download references


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.

Author information



Corresponding author

Correspondence to Liam Dolan.

Ethics declarations

Competing interests

The authors declare that they have no competing financial interests.

Supplementary information

Rights and permissions

Reprints and Permissions

About this article

Cite this article

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

Download citation

Further reading


By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.


Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing