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JXB Advance Access published online on August 13, 2004
Journal of Experimental Botany, doi:10.1093/jxb/erh231
© 2004 by Oxford University Press
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Received January 8, 2004
Accepted June 18, 2004

RESEARCH PAPER

Effect of NO3- transport and reduction on intracellular pH: an in vivo NMR study in maize roots

Luca Espen 1, Fabio F. Nocito 1, Maurizio Cocucci 1*

1 Dipartimento di Produzione Vegetale, University of Milan, Via Celoria 2, I-20133 Milano, Italy

* To whom correspondence should be addressed. E-mail: maurizio.cocucci{at}unimi.it.


  Abstract

The effect of NO3- uptake on cellular pH was studied in maize roots by an in vivo 31P-NMR technique. In order to separate the effects on cytoplasmic pH due to NO3- uptake from those due to NO3- reduction, tungstate was used to inhibit nitrate reductase (NR). The results confirm that in maize roots tungstate inhibited NR activity. 15N-NMR in vivo experiments demonstrated the cessation of nitrogen flux from nitrate to organic compounds. Tungstate affected neither NO3- uptake nor the levels of the main phosphorylated compounds. Slight changes in cytoplasmic pH were observed during NO3- uptake and reduction (i.e. control). By contrast, in the presence of tungstate, a consistent decrease in cytoplasmic pH occurred. The vacuolar pH did not change in any of the conditions tested. These data show that NO3- uptake is an acidifying process and suggest a possible involvement of NO3- reduction in pH homeostasis. In the presence of NO3-, a transient depolarization of transmembrane electric potential difference (Em) was observed in all the conditions analysed. However, in tungstate-treated roots, a lesser depolarization accompanied by a greater ability to recover Em was found. This was related to a higher activity of the plasma membrane (PM) H+-ATPase. When NO3- was administered as potassium salt, its uptake increased and a greater depolarization of Em took place, whilst the changes in cytoplasmic pH were remarkably reduced, according to the central role played by K+ in the control of plasma membrane activities and cell pH homeostasis. A possible involvement of cytoplasmic pH in the control of PM H+-ATPase expression during nitrate exposure is suggested.

Keywords: Cytoplasmic pH; maize; nitrate reductase; nitrate uptake; NMR spectroscopy; tungstate.
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