Transcriptome and metabolite profiling to identify genes associated with rhizome lignification and the function of ZoCSE in ginger (Zingiber officinale).

Ginger (Zingiber officinale Roscoe) is an important spice crop in China, and fresh ginger rhizomes are consumed as vegetable in Sichuan and Chongqing. However, tissue lignification accelerates with rhizome maturation, resulting in the loss of edible quality. To understand the molecular mechanisms of texture modification during rhizome development, we investigated lignin accumulation patterns and identified the key genes associated with lignin biosynthesis using gas chromatography-mass spectrometry (GC-MS), liquid chromatography-tandem mass spectrometry (LC-MS/MS) and RNA-sequencing (RNA-Seq). Results showed that the contents of total lignin and its precursors exhibited notable declines with tissue maturation. However, the lignin composition was remarkably modified and syringyl lignin was deposited in mature rhizomes, leading to ginger lignification. Transcriptome analysis displayed 32 lignin biosynthetic genes were dramatically downregulated with rhizome development, including caffeoylshikimate esterase (CSE ), 4-coumarate-CoA ligase , laccase , cinnamoyl-CoA reductase , cinnamyl-alcohol dehydrogenase , peroxidase and caffeic acid 3-O-methyltransferase , indicating that lignin reduction might be attributed to deficiency in intermediates or the downregulation of key biosynthetic enzymes. Furthermore, overexpressing ZoCSE in Nicotiana benthamiana L. enhanced the total lignin content, suggesting its fundamental role in lignin biosynthesis. RNA-Seq also identified candidate lignin production regulators, including hormone-related genes and NAC/MYB transcription factors (ZoNAC1 , ZoNAC4 , ZoMYB14 and ZoMYB17 ). This result provides a molecular basis for lignin accumulation in ginger.

A R2R3-MYB transcriptional activator LmMYB15 regulates chlorogenic acid biosynthesis and phenylpropanoid metabolism in Lonicera macranthoides.

Lonicera macranthoides Hand-Mazz is an important medicinal plant widely distributed in southern China that has long been used in Chinese traditional medicines. Chlorogenic acid (CGA, 3-caffeoylquinic acid) is the major biologically active ingredient in L. macranthoides. Although key CGA biosynthetic genes have been well documented, their transcriptional regulation remains largely unknown. In this study, we observed that a R2R3 MYB transcription factor LmMYB15 showed a significant correlation with CGA content, indicating its potential role in CGA biosynthesis. A yeast two-hybrid assay suggested that LmMYB15 functions as a transcriptional activator. Overexpression of LmMYB15 in tobacco led to increased accumulation of CGA compared to those in wild-type leaves. To elucidate its functional mechanism, genome-wide DAP-seq was employed and identified the conserved binding motifs of LmMYB15, that is [(C/T) (C/T) (C/T) ACCTA(C/A) (C/T) (A/T)], as well as its direct downstream target genes, including 4CL, MYB3, MYB4, KNAT6/7, IAA26, and ETR2. Subsequently, yeast one-hybrid and dual-luciferase reporter assays verified that LmMYB15 could bind and activate the promoters of 4CL, MYB3 and MYB4, thereby facilitating CGA biosynthesis and phenylpropanoid metabolism. Our findings provide a new track for breeding strategies aiming to enhance CGA content in L. macranthoides that can significantly contribute to better mechanical properties.