Dynamics and localization of H₂O₂ production in elicited plant cells

• Published in: Protoplasma 2007-03, Vol.230 (1-2), p.89-97
• Main Authors: Bóka, K, Orbán, N, Kristóf, Z
• Format: Article
• Language: English
• Subjects: Acids; Cell Survival - drug effects; Cells, Cultured; Cyclopentanes - pharmacology; Electron energy loss spectroscopy; Elicitor; hydrogen peroxide; Hydrogen Peroxide - metabolism; Microscopy, Electron; Oxylipins; Rubia - drug effects; Rubia - metabolism; Rubia - ultrastructure; Rubia tinctorum; Salicylic Acid - pharmacology; Spectrofluorometry; spectroscopy; Spectroscopy, Electron Energy-Loss
• ISSN: 0033-183X; 1615-6102
• DOI: 10.1007/s00709-006-0225-8

H₂O₂ produced in plant cells plays a dual role. In addition to its antimicrobial effect, it also acts as a secondary messenger initiating and modulating responses of plants exposed to unfavorable external signals. A suspension culture of Rubia tinctorum cells challenged with elicitors was used as a model system to investigate H₂O₂ formation. Cellular H₂O₂ was measured by a modified titanium(IV) method, while that in the medium was detected with scopoletin fluorescence. Localization of H₂O₂ production at the ultrastructural level was carried out by the CeCl₃ reaction. A fungal elicitor induced H₂O₂ production with transient maxima, the first of which appeared 4 min after treatment. Three subsequent maxima appeared in the cells up to 48 h after treatment. Exposure of cells to exogenous jasmonic acid and salicylic acid also changed the H₂O₂ concentration maxima over 48 h; however, their timing was slightly shifted. Fungal-elicitor, jasmonic acid, and salicylic acid treatments had different effects on the H₂O₂ concentration in the medium. Ultrastructural investigations revealed that electron-dense precipitates were present at the plasmalemma and in some nearby vesicular cytoplasmic structures 30 min after treatment. Later samples showed cytochemical-precipitate accumulation in the cell walls. These deposits appeared to be local and independent of the direction of the external signal. We could not detect the presence of H₂O₂ in peroxisomes, mitochondria, plastids, or the central vacuolar space. Electron energy loss spectroscopy investigations distinguished between the cerium-containing precipitates and other electrondense particles, thereby proving that H₂O₂ generation occurs locally.