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Plant and Cell Physiology Advance Access published online on June 4, 2005
Plant and Cell Physiology, doi:10.1093/pcp/pci142
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Plant and Cell Physiology 2005 © The Japanese Society of Plant Physiologists (JSPP); all rights reserved.
Received March 17, 2005
Accepted May 26, 2005

Regular Paper

Modulation of Reactive Oxygen Species Production During Osmotic Stress in Arabidopsis thaliana Cultured Cells: Involvement of the Plasma Membrane Ca2+-ATPase and H+-ATPase

Nicoletta Beffagna 1*, Barbara Buffoli 1, and Chiara Busi 1

1 Istituto di Biofisica del CNR - Sezione di Milano, Dipartimento di Biologia, Università degli Studi di Milano, Via Celoria 26 - 20133 Milano, Italy

* To whom correspondence should be addressed.
Nicoletta Beffagna, E-mail: Nicoletta.Beffagna{at}unimi.it


  Abstract

In Arabidopsis thaliana cells hypoosmotic treatment initially stimulates Ca2+ influx and inhibits its efflux and, concurrently, promotes a large H2O2 accumulation in the external medium, representative of reactive oxygen species (ROS) production. After the first 10-15 min, Ca2+ influx rate is however lowered and a large rise in Ca2+ efflux, concomitant with a rapid decline in H2O2 level, takes place. The drop of H2O2 peak, as well as the efflux of Ca2+, are prevented by treatment with sub-µM concentrations of eosin yellow (EY), selectively inhibiting the Ca2+-ATPase of plasma membrane (PM). Comparable changes of Ca2+ fluxes are induced by hyperosmotic treatment too. However, in this case, H2O2 level does not rise, but declines below control levels when Ca2+ efflux is activated. Also K+ and H+ net fluxes across the PM and cytoplasmic pH (pHcyt) are very differently influenced by the two opposite stresses: strongly decreased by hypoosmotic stress and increased under hyperosmotic treatment. The H2O2 accumulation kinetics, followed as a function of the pHcyt changes imposed by modulation of the PM H+-ATPase activity or weak acid treatment, show a close correlation between pHcyt and H2O2 formed, the larger amount being produced for changes towards acidic pH values. Overall, these results confirm a relevant role for the PM Ca2+-ATPase in switching off the signal triggering ROS production, and propose a role for the PM H+-ATPase in modulating the development of the oxidative wave through the pHcyt changes following the changes of its activity induced by stress conditions.

Keywords: Arabidopsis thaliana cells; Ca2+ fluxes; cytoplasmic pH; osmotic stress; plasma membrane ATPases; reactive oxygen species.
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