Transcriptome Analysis of Salicylic Acid Treatment in Rehmannia glutinosa Hairy Roots Using RNA-seq Technique for Identification of Genes Involved in Acteoside Biosynthesis

Abstract

Rehmannia glutinosa is a common bulk medicinal material that has been widely used in China due to its active ingredients. Acteoside, one of the ingredients, has antioxidant, antinephritic, anti-inflammatory, hepatoprotective, immunomodulatory, and neuroprotective effects, is usually selected as a quality-control component for R. glutinosa herb in the Chinese Pharmacopeia. The acteoside biosynthesis pathway in R. glutinosa has not yet been clearly established. Herein, we describe the establishment of a genetic transformation system for R. glutinosa mediated by Agrobacterium rhizogenes. We screened the optimal elicitors that markedly increased acteoside accumulation in R. glutinosa hairy roots. We found that acteoside accumulation dramatically increased with the addition of salicylic acid (SA); the optimal SA dose was 25 μmol/L for hairy roots. RNA-seq was applied to analyze the transcriptomic changes in hairy roots treated with SA for 24 h in comparison with an untreated control. A total of 3,716, 4,018, and 2,715 differentially expressed transcripts (DETs) were identified in 0 h-vs.-12 h, 0 h-vs.-24 h, and 12 h-vs.-24 h libraries, respectively. KEGG pathway-based analysis revealed that 127 DETs were enriched in “phenylpropanoid biosynthesis.” Of 219 putative unigenes involved in acteoside biosynthesis, 54 were found to be up-regulated at at least one of the time points after SA treatment. Selected candidate genes were analyzed by quantitative real-time PCR (qRT-PCR) in hairy roots with SA, methyl jasmonate (MeJA), AgNO3 (Ag+), and putrescine (Put) treatment. All genes investigated were up-regulated by SA treatment, and most candidate genes were weakly increased by MeJA to some degree. Furthermore, transcription abundance of eight candidate genes in tuberous roots of the high-acteoside-content (HA) cultivar QH were higher than those of the low-acteoside-content (LA) cultivar Wen 85-5. These results will pave the way for understanding the molecular basis of acteoside biosynthesis in R. glutinosa, and can serve as a basis for future validation studies.

Molecular Regulation of Catalpol and Acteoside Accumulation in Radial Striation and non-Radial Striation of Rehmannia glutinosa Tuberous Root

“…The biosynthetic pathway of acteoside in Olea europaea has been characterized [22]. In R. glutinosa , two biosynthetic pathways are involved in the biosynthesis of the processor of phenylethanoid glycosides: the phenylalanine-derived pathway and the tyrosine-derived pathway [18] (Figure 7A). In the phenylalanine-derived pathway, phenylalanine is metabolized to caffeoyl-CoA through phenylalanine ammonia-lyase (PAL), cinnamate-4-hydroxylase (C4H), coumarate-3-hydroxylase (C3H), and 4-coumarate-CoA ligase (4CL).…”

“…Catalpol and acteoside are usually selected as the two quality-control components in the R. glutinosa herb and have been included in the Chinese Pharmacopoeia since the 2015 edition. Considering limited genomic information coupled with genome complexity (polyploidy), several reports about the catalpol and acteoside biosynthesis pathway have been seen in some studies [16,17,18], but the molecular regulation mechanism needs to be deeply studied.…”

Comparative Transcriptome Analysis of Pine Trees Treated with Resistance-Inducing Substances against the Nematode Bursaphelenchus xylophilus

“…Given that studies focussing solely on isolated contributary factors often fail to yield meaningful insights into the important elements of biological processes, we designed the present study with a view towards characterising an integrated environment comprising pine hosts, RISs, and PWNs. Simple treatment using RISs is limited to the detection of general genes (termed pathogenesis-related genes) under SAR conditions [ 67 , 68 ], whereas PWN infection in pine trees predominantly reflects the pathogenic mechanisms within dying cells [ 69 ]. Consequently, the integrated design of the present study enabled us to assess the sequential stages of RIS treatment and infection, as well as the activation of related genes under conditions of SAR induction, thereby facilitating subsequent analyses of the essential mechanisms underlying the protection against and inhibition of PWN.…”

Phospholipid Signaling Is a Component of the Salicylic Acid Response in Plant Cell Suspension Cultures

“…Much effort has been spent to understand the cascades of reactions that result in the formation and accumulation of secondary metabolites in plant cells in response to SA. In this sense, it has been shown that signaling molecules lead to genetic expression and biochemical changes in a specific metabolic pathway [93,94]. Additionally, a whole-genome approaches such as metabolic and transcriptomic profiling in the roots of Arabidopsis [95], in the seeds of Triticum estivum L. [96], in Rehmannia glutinosa hairy roots [94] and in Scutellaria baicalensis [97], have been used to determine the genes specifically regulated in response to SA treatment.…”