Ectopic expression of Dryopteris fragrans DfMTPSL6, a directly target gene of DfWRKY16/45, enhanced drought tolerance in tobacco plants

Abstract

Terpenoid synthesis in seed plants is primarily catalyzed by Typical Terpene Synthase (TPS) enzymes. However, terrestrial non-seed plants also possess microbial terpene synthase-like (MTPSL) enzymes for terpene synthesis. A previous study has demonstrated the presence of both TPSs and MTPSLs in Dryopteris fragrans (L.) Schott. Specifically, DfMTPSL6 has been identified as the enzyme responsible for catalyzing the conversion of farnesyl diphosphate (FPP) to nerolidol. Nerolidol has several functions, including insect, disease and chilling resistance, although its biological role in eukaryotes remains to be confirmed. Transcription factors regulate the terpenoid biosynthesis by binding to gene promoters. However, the regulation of terpene metabolism by transcription factors, including MTPSLs, has not been investigated in ferns.

This study analyzed the conservation of DfMTPSL6, a nerolidol synthase, expressed in tobacco plants to enhance drought tolerance. Promoter analysis revealed specific expression in glandular hairs, with the active site responsive to MeJA and PEG treatments and containing a W-box, a binding site for WRKY transcription factors. 48 DfWRKY transcription factors were identified, and their expression patterns under MeJA and PEG treatments were analyzed. Yeast one-hybrid assays identified DfWRKY16 and DfWRKY45 as potential regulators of DfMTPSL6. Subcellular localization and transcriptional activation analysis confirmed that DfWRKY16 and DfWRKY45 are transcriptional activator and promotion of DfMTPSL6 expression.