Disruption of putative anion channel gene AtCLC-a in Arabidopsis suggests a role in the regulation of nitrate content

Authors

  • Danny Geelen,

    1. Institut des Sciences Végétales, Centre National de la Recherche Scientifique (CNRS), Bât 22, avenue de la Terrasse, F-91198 Gif-sur-Yvette Cedex, France, and
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    • Present address: Laboratorium voor Genetica, Departement Planten Genetica, Vlaams Interuniversitai, Instituut voor Biotechnologie (VIB), Universiteit Gent, K.L. Ledeganckstraat 35, B-9000 Gent, Belgium.

  • Claire Lurin,

    1. Institut des Sciences Végétales, Centre National de la Recherche Scientifique (CNRS), Bât 22, avenue de la Terrasse, F-91198 Gif-sur-Yvette Cedex, France, and
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    • Present address: Departement of Biology, University of Pennsylvania, 415 South University Avenue, Philadelphia, PA 19104-6018, USA.

  • David Bouchez,

    1. Laboratoire de Biologie Cellulaire, Institut National de la Recherche Agronomique, Route de Saint-Cyr, F-78026 Versailles Cedex, France
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  • Jean-Marie Frachisse,

    1. Institut des Sciences Végétales, Centre National de la Recherche Scientifique (CNRS), Bât 22, avenue de la Terrasse, F-91198 Gif-sur-Yvette Cedex, France, and
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  • Françoise Lelièvre,

    1. Institut des Sciences Végétales, Centre National de la Recherche Scientifique (CNRS), Bât 22, avenue de la Terrasse, F-91198 Gif-sur-Yvette Cedex, France, and
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  • Béatrice Courtial,

    1. Laboratoire de Biologie Cellulaire, Institut National de la Recherche Agronomique, Route de Saint-Cyr, F-78026 Versailles Cedex, France
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  • Hélène Barbier-Brygoo,

    1. Institut des Sciences Végétales, Centre National de la Recherche Scientifique (CNRS), Bât 22, avenue de la Terrasse, F-91198 Gif-sur-Yvette Cedex, France, and
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  • Christophe Maurel

    Corresponding author
    1. Institut des Sciences Végétales, Centre National de la Recherche Scientifique (CNRS), Bât 22, avenue de la Terrasse, F-91198 Gif-sur-Yvette Cedex, France, and
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    • §

      Present address: Biochimie et Physiologie Moléculaire des Plantes, ENSA-M/INRA/CNRS UMR 5004, 2 Place Viala, 34060 Montpellier Cedex 1, France.


*For correspondence (fax +33 4 6752 5737; e-mail maurel@ensam.inra.fr).

Summary

In animals and yeast, voltage-dependent chloride channels of the CLC family play a role in basic cellular functions such as epithelial transport, plasma membrane excitability, and control of pH and membrane potential in intracellular compartments. To assess the function of CLCs in plants, we searched for CLC insertion mutants in a library of Arabidopsis lines transformed by Agrobacterium tumefaciens transferred DNA (T-DNA). Using a polymerase chain reaction-based screening procedure, an Arabidopsis line that carries a T-DNA insertion within the C-terminus of the AtCLC-a coding sequence was identified. Progeny from this plant line, clca-1, showed dramatically altered transcription of the AtCLC-a gene. Plants homozygous for the clca-1 mutation exhibited normal development and a morphology indistinguishable from the wild-type. However, their capacity to accumulate nitrate under conditions of nitrate excess was reduced in roots and shoots, by approximately 50%, while chloride, sulphate and phosphate levels were similar to the wild-type. In addition, the herbicide chlorate, an analogue of nitrate, induced a faster and more pronounced chlorosis in mutant plants. Hypersensitivity to chlorate as well as decreased nitrate levels co-segregated with the T-DNA insertion. They were found at various time points of the clca-1 life cycle, supporting the idea that AtCLC-a has a general role in the control of the nitrate status in Arabidopsis. Concordant with such a function, AtCLC-a mRNA was found in roots and shoots, and its levels rapidly increased in both tissues upon addition of nitrate but not ammonium to the culture medium. The specificity of AtCLC-a function with respect to nitrate is further supported by a similar free amino acid content in wild-type and clca-1 plants. Although the cellular localization of AtCLC-a remains unclear, our results suggest that AtCLC-a plays a role in controlling the intracellular nitrate status.

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