Characterization and Function of the 1-Deoxy-D-xylose-5-Phosphate Synthase (DXS) Gene Related to Terpenoid Synthesis in Pinus massoniana

Abstract

In the methyl-D-erythritol-4-phosphate (MEP) pathway, 1-deoxy-D-xylose-5-phosphate synthase (DXS) is considered the key enzyme for the biosynthesis of terpenoids. In this study, <i>PmDXS</i> (MK970590) was isolated from <i>Pinus massoniana</i>. Bioinformatics analysis revealed homology of MK970590 with DXS proteins from other species. Relative expression analysis suggested that <i>PmDXS</i> expression was higher in roots than in other plant parts, and the treatment of <i>P. massoniana</i> seedlings with mechanical injury via 15% polyethylene glycol 6000, 10 mM H₂O₂, 50 μM ethephon (ETH), 10 mM methyl jasmonate (MeJA), and 1 mM salicylic acid (SA) resulted in an increased expression of <i>PmDXS</i>. pET28a-<i>PmDXS</i> was expressed in <i>Escherichia coli</i> TransB (DE3) cells, and stress analysis showed that the recombinant protein was involved in resistance to NaCl and drought stresses. The subcellular localization of <i>PmDXS</i> was in the chloroplast. We also cloned a full-length 1024 bp <i>PmDXS</i> promoter. <i>GUS</i> expression was observed in <i>Nicotiana benthamiana</i> roots, stems, and leaves. <i>PmDXS</i> overexpression significantly increased carotenoid, chlorophyll a, and chlorophyll b contents and DXS enzyme activity, suggesting that <i>DXS</i> is important in isoprenoid biosynthesis. This study provides a theoretical basis for molecular breeding for terpene synthesis regulation and resistance.