Expression and promoter analysis of MEP pathway enzyme-encoding genes in Pinus massoniana Lamb

Abstract

The methylerythritol phosphate (MEP) pathway provides the universal basic blocks for the biosynthesis of terpenoids and plays a critical role in the growth and development of higher plants. Pinus massoniana is the most valuable oleoresin producer tree with an extensive terrestrial range. It has the potential to produce more oleoresin with commercial value, while being resistant to pine wood nematode (PWN) disease. For this study, eleven MEP pathway associated enzyme-encoding genes and ten promoters were isolated from P. massoniana. Three PmDXS and two PmHDR existed as multi-copy genes, whereas the other six genes existed as single copies. All eleven of these MEP enzymes exhibited chloroplast localization with transient expression. Most of the MEP genes showed higher expression in the needles, while PmDXS2, PmDXS3, and PmHDR1 had high expression in the roots. The expressions of a few MEP genes could be induced under exogenous elicitor conditions. The functional complementation in a dxs-mutant Escherichia coli strain showed the DXS enzymatic activities of the three PmDXSs. High throughput TAIL PCR was employed to obtain the upstream sequences of the genes encoding for enzymes in the MEP pathway, whereby abundant light responsive cis-elements and transcription factor (TF) binding sites were identified within the ten promoters. This study provides a theoretical basis for research on the functionality and transcriptional regulation of MEP enzymes, as well as a potential strategy for high-resin generation and improved genetic resistance in P. massoniana.

Keywords

• Pinus massoniana
• MEP pathway
• Expression
• Subcellular localization
• Promoter