H2O2 Participates in the Induction and Formation of Potato Tubers by Activating Tuberization-Related Signal Transduction Pathways

Reactive oxygen species (ROS), especially hydrogen peroxide, H2O2, act as signaling molecules to widely mediate growth, development, and stress response of plants. In the present study, internal ROS accumulation, effects of exogenous H2O2 treatment, the expression of the key tuberization-related genes, and the effect of knockout of Solanum tuberosum self-pruning 6A (StSP6A) on H2O2-induced tuber formation were investigated to elucidate whether and how H2O2 is involved in induction and formation of potato tubers using two diploid landraces, Solanum phureja and S. ajanhuiri. The results showed that there was a significant accumulation of ROS (including H2O2, superoxide anion, O2&minus:, and total ROS) during tuber induction and formation in stolons/tubers, especially in the hook-like subapical part of stolons prior to tuberization, as detected by staining observation and quantitative measurement. Furthermore, exogenous H2O2 treatment significantly enhanced percentage of tuber formation. By contrast, addition of either the ROS inhibitor diphenyleneiodonium chloride (DPI) or H2O2 scavenger catalase (CAT) resulted in a decline of tuber formation. In addition, expression analysis of nine key tuberization-related genes demonstrated that the H2O2-induced tuberization could be associated with H2O2-controlled regulation of these tuberization- and signaling-pathway-related genes, especially StSP6A, which was dramatically up-regulated during the early stage of tuber induction and H2O2 treatment. When StSP6A was knocked out by CRISPR-Cas9-mediated genome editing, the tuberization frequency of StSP6A null-mutants became significantly lower at various H2O2 concentration treatments. These findings indicate that H2O2 accumulation in stolons might play an important role by acting as a signaling molecule to initiate tuber induction, H2O2-induced tuber formation is triggered by regulating the tuberization-related gene expression and activating signal transduction pathways, and StSP6A is a pivotal player in H2O2-induced tuber formation in potato.