Stratification of apple seeds in the context of ROS metabolism.

Apple (Malus domestica Borkh.) seeds exhibit deep embryonic dormancy. Uniform germination of isolated apple embryos is observed after 40-day-long cold stratification of the seeds. Stratification treatment modifies the level of reactive oxygen species (ROS), which are regarded as key regulators of seed dormancy. In this study, axes of embryos isolated from seeds stratified for 7, 14, 21, and 40 days differing in dormancy depth were used. After one week of stratification, the increased polyamine oxidase activity enables ROS generation, which is followed by an upregulation of the NADPH oxidase gene expression. Catalase activity increased after 14 days of stratification, suggesting the requirement to maintain ROS concentrations at an optimal level already in the early phase of dormancy removal. When cold stratification was prolonged, accompanied by a significant increase in ROS level, ROS scavenging by catalase was supported by elevated phenolic compounds content. Then, peroxidase activity was also the highest. As ROS-induced phenylalanine (Phe) oxidation leads to the formation of meta-tyrosine (m-Tyr) - a potentially toxic component, the levels of these amino acids were examined. The fluctuation in m-Tyr content indicates the existence of mechanisms in the tissue for the disposal of this compound. Finally, its presence may be mitigated by an increase in Phe levels. Maintaining oxidised RNA at elevated levels from the 14th day of stratification may be crucial for seed dormancy removal, ensuring translation regulation as metabolism resumes. We concluded that dormancy removal of apple seeds by stratification requires a time-dependent sequence of biochemical events reflecting ROS metabolism alterations.