NUCLEAR TRANSPORT FACTOR 2‐LIKE improves drought tolerance by modulating leaf water loss in alfalfa ( Medicago sativa L.)

Drought is a major environmental factor that limits the production of alfalfa (Medicago sativa). In the present study, M. sativa NUCLEAR TRANSPORT FACTOR 2-LIKE (MsNTF2L) was identified as a nucleus-, cytoplasm-, and plasma membrane-localized protein. Its transcriptional expression was highly induced by ABA and drought stress. Overexpression of MsNTF2L in Arabidopsis resulted in hypersensitivity to ABA during both the seed germination and seedling growth stages. However, transgenic Arabidopsis plants exhibited enhanced tolerance to drought stress by reducing the levels of reactive oxygen species (ROS) and increasing the expression of stress/ABA-inducible genes. Consistently, analysis of MsNTF2L overexpression (OE) and RNA interference (RNAi) alfalfa plants revealed that MsNTF2L confers drought tolerance through promoting ROS scavenging, a decrease in stomatal density, ABA-induced stomatal closure, and epicuticular wax crystal accumulation. MsNTF2L highly affected epicuticular wax deposition, as a large group of wax biosynthesis and transport genes were influenced in the alfalfa OE and RNAi lines. Furthermore, transcript profiling of drought-treated alfalfa WT, OE, and RNAi plants showed a differential drought response for genes related to stress/ABA signaling, antioxidant defense, and photosynthesis. Taken together, these results reveal that MsNTF2L confers drought tolerance in alfalfa via modulation of leaf water loss (by regulating both stomata and wax deposition), antioxidant defense, and photosynthesis.