The superoxide-mediated ascorbate-glutathione cycle modulates the transition of growth to reproduction in Ulva prolifera

Summary The major reactive oxygen species superoxide (O 2 − ) plays important roles in plant growth, development, and stress responses. However, the molecular mechanism underlying O 2 − mediation of reproduction in Ulva prolifera is unclear. Here, we used inhibitors and promoters alongside phenotypes, histochemical staining, transcriptomic analysis, complementation tests, and related physiology parameter assays to determine that O 2 − was the main ROS that regulated reproduction cell formation by modulating the ascorbate-glutathione cycle. We found that O 2 − was the main ROS involved in regulating reproduction and was regulated by Ca 2+ levels. Analysis of the effects of different sources of O 2 − showed that chloroplast O 2 − mainly affected early reproductive progress and mitochondrial O 2 − impacted the entire reproduction process by regulating energy metabolism. Moreover, ascorbic acid (AsA) transmitted oxidative signals through the AsA-GSH cycle, thus interacting with O 2 − . However, an imbalance between AsA and ROS significantly impacted the oxidative signal transduction required for reproduction. Additionally, transcriptomic analysis revealed that the expression of genes related to O 2 − production and signal transmission, such as DHAR, were significantly downregulated by DPI and EGTA treatment, while the upregulated expression of GR, GSS, and GST could increase GSH content, resulting in a high GSH/GSSG ratio, which weakened oxidative signaling and disrupted reproduction. Taken together, our findings demonstrated that Ca 2+ level might regulate O 2 − production, and through DHAR affect the redox state change of the AsA-GSH cycle involved in signal transmission, which participates in the ROS signal transmit pathway and affects reproduction. These results provide new viewpoint about the reproduction and breeding of U. prolifera , then provide a scientific basis for land-based cultivation.