Reactive Nitrogen and Oxygen Species in Defense/Stress Responses Activated by Chitosan in Sycamore Cultured Cells

Chitosan (CHT) is a natural, non-toxic and inexpensive compound obtained by partial alkaline deacetylation of chitin, the main component of the exoskeleton of crustaceans and other arthropods. Although the exact mode of action of CHT is still unknown, in agriculture it has been shown to be a versatile compound that controls numerous pre and postharvest diseases on various horticultural commodities1. In sycamore (Acer pseudoplatanus L.) cultured cells, a material well characterized both biochemically and physiologically, CHT rapidly induces a set of defense/stress responses: accumulation of dead cells and of cells with fragmented DNA, production of H2O2 and nitric oxide (NO), accumulation of regulative 14-3-3 proteins in the cytosol and of HSP70 molecular chaperone Binding Protein (BiP) in the endoplasmic reticulum, release of cytochrome c from the mitochondrion2. These findings open the possibility to use sycamore cultured cells to investigate the signalling pathways leading to the CHT-induced responses. In this work we investigated the possible signalling role of reactive nitrogen and oxygen species in the CHT-induced responses by means of inhibitors of production and/or scavengers. In particular, sycamore cultured cells have been challenged with CHT in the presence or in the absence of Tiron (4,5-dihydroxy-1,3-benzene disulfonic acid), a specific scavenger of superoxide anion (O2.-), DPI (diphenilene iodonium), a specific inhibitor of the NADPH oxidase (O2.- synthase) of the plasma membrane, and cPTIO (2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide), a specific NO scavenger. The results show that both reactive nitrogen and oxygen species are involved in the responses activated by CHT in sycamore cultured cells.