Genome-Wide Identification and Genetic Characterization of Eucommia ulmoides NAC Family Genes and Functional Analysis of EuNAC9 in Relieving Mn2+ Stress

NAC (NAM/ATAF1,2/CUC2) is one of the most important plant-specific transcription factors, playing a significant role in plant growth, morphogenesis, and stress resilience. In this investigation, 77 members of the NAC gene family were identified from the Eucommia ulmoides database, and these E. ulmoides NAC families were classified into 18 subfamilies based on their protein sequences. Currently, the architectural design, conserved domains, and cis-acting elements of EuNAC genes, as well as physicochemical properties of EuNAC proteins, have also been analyzed. In addition, the expression patterns of selected EuNAC genes under five heavy metal (Cu2+, Cd2+, Pb2+, Mn2+, and As5+) imposed stresses were investigated. The results showed that the expression of EuNAC9 experienced a significant induction to upregulate under these five heavy metal stresses, suggesting that EuNAC9 may mediate the response to these five heavy metal stresses. In addition, the function of EuNAC9 was investigated in both transgenic yeasts and tobaccos. The transgenic yeasts exhibited enhanced resistance to Mn2+ stress by increasing the expression of ScSMF1 and ScSOD2, whereas the transgenic tobacco grew better under Mn2+ stress and showed intense antioxidant enzyme activity to alleviate the accumulation of H2O2 and O2-. Moreover, subcellular localization indicated EuNAC9 was a nuclear localization protein, and the transcriptional activity domain of EuNAC9 resided at the C-terminal. This study will provide a theoretical framework for analyzing the molecular regulatory mechanisms of plant responses to heavy metals mediated by EuNAC9 genes and elucidating the biological roles of E. ulmoides NACs in responding to heavy metal stress.