Relationship between H₂O₂ accumulation and NO synthesis during osmotic stress: promoted somatic embryogenesis of Fraxinus mandshurica

Abstract

Osmotic stress promotes somatic embryogenesis of Fraxinus mandshurica, which leads to accumulation of reactive oxygen species (ROS). The single pieces of cotyledons of F. mandshurica were used as explants to induce somatic embryogenesis in osmotic-stress medium. Furthermore, the hydrogen peroxide (H₂O₂) content of explanted cells was varied by adding exogenous H₂O₂ or catalase solution to assess the effects of the exogenous H₂O₂ on somatic embryogenesis, intracellular H₂O₂ accumulation, and the relationship between signaling mediated by ROS or reactive nitrogen species. The results revealed that exogenous H₂O₂ (100‒300 μmol L^–1) increased the number of somatic embryos. On 60th day of exogenous H₂O₂ (200 μmol L^–1) treatment, the number of somatic embryos of explants treated, which was 136.54%, was higher than the control. Moreover, exogenous H₂O₂ (100 μmol L^–1) significantly increased the intracellular H₂O₂ content and enhanced the activities of superoxidase dismutase and peroxidase. Finally, exogenous H₂O₂ (100 μmol L^–1) activated the intracellular non-enzymatic pathway for nitric oxide (NO) synthesis. The somatic embryogenesis in broadleaf trees increases with the change of endogenic ROS content, and depends on the upregulation of antioxidant enzymes. Both H₂O₂ and NO, as signaling molecules, were found to be involved in the process of somatic embryogenesis in broadleaf trees. In the process of exogenous H₂O₂ promoting somatic embryogenesis, NO synthesis depended on non-enzymatic reactions. These results provide a scientific basis for resolving the mechanism by which ROS levels are regulated during somatic embryogenesis of broadleaf trees and establish a reasonable and efficient technology system for regulating somatic embryogenesis of trees.

Keywords

Fraxinus mandshurica / Somatic embryogenesis / Hydrogen peroxide / Nitric oxide / Osmotic stress

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