Overexpression of cytosolic NADP-malic enzyme 1 from the common ice plant enhances water-deficit and high-light stress tolerance by modulating water-use efficiency and flavonoid biosynthesis

Innovative strategies are essential to enhance crop resilience against drought and heat stress intensified by climate change. Crassulacean acid metabolism (CAM) is a specialized photosynthetic pathway that improves water-use efficiency (WUE) by shifting CO₂ fixation to the nighttime. The common ice plant (Mesembryanthemum crystallinum) utilizes NADP-malic enzymes (ME) for malate decarboxylation during its facultative CAM transition. In this study, we characterized the cytosolic McNADP-ME1 enzyme, which is highly expressed in the ice plant under water-deficit stress. Transgenic Arabidopsis thaliana plants overexpressing McNADP-ME1 exhibited reduced stomatal density, size, and conductance, leading to enhanced instantaneous WUE. Although these modifications resulted in reduced biomass and seed yield under low-light conditions, the transgenic lines showed significantly improved survival and growth under both acute and chronic water-deficit stress. Additionally, McNADP-ME1 overexpression conferred improved vegetative growth under high-light stress conditions. Notably, McNADP-ME1 overexpression upregulated many genes within the flavonoid biosynthetic pathway, resulting in a marked increase in total flavonoid content. These flavonoids acted as effective antioxidants that facilitated the scavenging of reactive oxygen species (ROS), thereby reducing oxidative damage and malondialdehyde (MDA) levels under both water-deficit and high-light stress conditions. These results demonstrate that cytosolic McNADP-ME1 is a key enzyme for conferring water-deficit stress tolerance by integrating stomatal-mediated water conservation with flavonoid-driven ROS-scavenging mechanisms.