Functional Identification and Regulatory Active Site Screening of the DfDXS Gene of Dryopteris fragrans

<i>Dryopteris fragrans</i> (L.) Schott has anti-inflammatory and antioxidant properties, and terpenoids are important components of its active constituents. The methyl-D-erythritol 4-phosphate (MEP) pathway is one of the major pathways for the synthesis of terpene precursors in plants, and 1-deoxy-D-xylulose-5-phosphate synthase (DXS) is the first rate-limiting enzyme in this pathway. <i>DXS</i> has been shown to be associated with increased stress tolerance in plants. In this experiment, two <i>DXS</i> genes were extracted from the <i>D. fragrans</i> transcriptome and named <i>DfDXS1</i> and <i>DfDXS2</i>. Based on phylogenetic tree and conserved motif analyses, DXS was shown to be highly conserved evolutionarily and its localization to chloroplasts was determined by subcellular localization. Prokaryotic expression results showed that the number and growth status of recombinant colonies were better than the control under 400 mM NaCl salt stress and 800 mM mannitol-simulated drought stress. In addition, the <i>DfDXS1</i> and <i>DfDXS2</i> transgenic tobacco plants showed improved resistance to drought and salt stress. <i>DfDXS1</i> and <i>DfDXS2</i> responded strongly to methyl jasmonate (MeJA) and PEG-mimicked drought stress following exogenous hormone and abiotic stress treatments of <i>D. fragrans</i>. The transcriptional active sites were investigated by dual luciferase and GUS staining assays, and the results showed that the STRE element (AGGGG), the ABRE element (ACGTGGC), and the MYC element (CATTTG) were the important transcriptional active sites in the promoters of the two <i>DXS</i> genes, which were closely associated with hormone response and abiotic stress. These results suggest that the <i>DfDXS</i> gene of <i>D. fragrans</i> plays an important role in hormone signaling and response to stress. This study provides a reference for analyzing the molecular mechanisms of stress tolerance in <i>D. fragrans</i>.