Drought and other abiotic stresses negatively affect plant growth and development and thus reduce productivity. The plant-specific NAM/ATAF1/2/CUC2 (NAC) transcription factors have important roles in abiotic stress-responsive signaling. Here, we show that Arabidopsis thaliana NAC016 is involved in drought stress responses; nac016 mutants have high drought tolerance, and NAC016-overexpressing (NAC016-OX) plants have low drought tolerance. Using genome-wide gene expression microarray analysis and MEME motif searches, we identified the NAC016-specific binding motif (NAC16BM), GATTGGAT[AT]CA, in the promoters of genes downregulated in nac016-1 mutants. The NAC16BM sequence does not contain the core NAC binding motif CACG (or its reverse complement CGTG). NAC016 directly binds to the NAC16BM in the promoter of ABSCISIC ACID-RESPONSIVE ELEMENT BINDING PROTEIN1 (AREB1), which encodes a central transcription factor in the stress-responsive abscisic acid signaling pathway and represses AREB1 transcription. We found that knockout mutants of the NAC016 target gene NAC-LIKE, ACTIVATED BY AP3/PI (NAP) also exhibited strong drought tolerance; moreover, NAP binds to the AREB1 promoter and suppresses AREB1 transcription. Taking these results together, we propose that a trifurcate feed-forward pathway involving NAC016, NAP, and AREB1 functions in the drought stress response, in addition to affecting leaf senescence in Arabidopsis.

Purpose: Gamma-rays and carbon ions are frequently used for mutation breeding in diverse plant species, whereas proton ions have been rarely used for this purpose. This study assessed the potential of proton ions for plant mutation breeding. Materials and methods: We compared the effects of radiation on creeping bentgrass seeds with γ-rays, proton ions, and carbon ions on seed germination, plant growth parameters, and DNA fragmentation. Results and conclusions: The lethal dose 50 (LD₅₀) doses based on seed germinability were 115.9 Gy (γ-rays), 225.1 Gy (proton ions), and 57.7 Gy (carbon ions). Threshold doses for survival were 150 Gy (γ-rays), 150 Gy (proton ions), and 25 Gy (carbon ions). Suppression of plant growth was displayed at 100 Gy (γ-rays), 25 Gy (proton ions), and 25 Gy (carbon ions). Similar patterns of decreasing head DNA percentage were observed for γ-rays and proton ions. Carbon ions induced the lowest frequency of DNA fragmentation. The biological effects of the ionizing radiation types on creeping bentgrass are summarizable as follows: germination, carbon ions (C) > γ-rays (G) > proton ions (P); survival, C > P = G; growth, C ≥ P > G; DNA fragmentation, G ≥ P > C. These results indicate that proton ions are useful as a physical mutagen in plant mutation breeding.