The effect of endogenous hydrogen peroxide induced by cold treatment in the improvement of tissue regeneration efficiency

We propose that oxidative stress resulting from an imbalance between generation and scavenging hydrogen peroxide contributes to tissue regeneration efficiency during somatic embryogenesis of hexaploid winter wheat ( Triticum aestivum cv. Kamila) and organogenesis of faba bean ( Vicia faba ssp. minor cv. Nadwislanski). Endogenous hydrogen peroxide content and antioxidant capacity of cells were determined in initial explants and callus cultures derived from these explants. Regeneration-competent explants (immature embryos) contained more endogenous H 2 O 2 than explants initiated from regeneration-recalcitrant tissue (mature wheat embryos and faba bean epicotyls). Higher H 2 O 2 levels were observed despite the higher activity of antioxidative enzymes (superoxide dismutase and catalase) and the induction of their gene expression. Calli originating from immature embryos retained the capacity of the initial explants: high H 2 O 2 production was observed during the whole culture period. Low temperature treatment (4°C) was found to be an effective factor, which improved both regeneration ability and H 2 O 2 production. Exogenous application to the medium of H 2 O 2 and catalase blocker (3-aminotriazole), but not FeEDTA and superoxide dismutase blocker (diethyldithiocarbamate), also resulted in the enhancement of regeneration efficiency. These results clearly indicate that plant regeneration is specifically regulated by endogenous H 2 O 2 and by factors, which improve its accumulation. Moreover, a study of the activity of various SOD isoforms suggests that not only the absolute concentration of H 2 O 2 , but also its localisation might be responsible for controlling regeneration processes.