Transcriptome Characterization of Gnetum parvifolium Reveals Candidate Genes Involved in Important Secondary Metabolic Pathways of Flavonoids and Stilbenoids

Abstract

Gnetum is a small, unique group of Gnetophyta with a controversial phylogenetic position. Gnetum parvifolium is an important Chinese traditional medicinal plant, which is rich in bioactive compounds such as flavonoids and stilbenoids. These compounds provide significant medicinal effects, mostly as antioxidant, anticancer, and antibacterial agents. However, the mechanisms involved in the biosynthesis and regulation of these compounds in G. parvifolium are still unknown. In this study, we found that flavonoids and stilbene compounds accumulated at different levels in various tissues of G. parvifolium. We further obtained and analyzed massive sequence information from pooled samples of G. parvifolium by transcriptome sequencing, which generated 94,816 unigenes with an average length of 724 bp. Functional annotation of all these unigenes revealed that many of them were associated with several important secondary metabolism pathways including flavonoids and stilbenoids. In particular, several candidate unigenes (PAL-, C4H-, 4CL-, and STS-like genes) involved in stilbenoids biosynthesis were highly expressed in leaves and mature fruits. Furthermore, high temperature and UV-C strongly induced the expression of these genes and enhanced stilbene production (i.e., resveratrol and piceatannol) in leaves of young seedlings. Our present transcriptomic and biochemical data on secondary metabolites in G. parvifolium should encourage further investigation on evolution, ecology, functional genomics, and breeding of this plant with strong pharmaceutical potential.

Phytochemical and comparative transcriptome analyses reveal different regulatory mechanisms in the terpenoid biosynthesis pathways between Matricaria recutita L. and Chamaemelum nobile L.

“…Transcriptomic analyses can be used to identify functional elements in the genome, metabolic pathways, and differentially expressed genes in model and non-model organisms [17][18][19][20]. The transcriptomes of many important medicinal plants and their individual tissues such as leaves, roots, and stems have been reported [21][22][23][24][25]. Genomic resources and transcriptome sequences for German chamomile and Roman chamomile are very limited at present, and the complex regulatory mechanisms that control carbon flux through the terpenoid biosynthetic pathway and their cooperation in the biosynthesis of volatile terpenoids remain unknown.…”

Regulation of stilbene biosynthesis in plants

“…Using LC-QTOF-MS-based metabolite profiling, Degu et al (2016) demonstrated that high light treatment (2500 lmol m -2 s -1 ) applied to detached Shiraz grape berries led to a transient increase in stilbene levels at pre-veraison (piceid) and veraison (piceid and viniferin) stages, while high temperature (40°C) specifically induced viniferin content at all sampling times only at the pre-veraison stage. Recent results obtained by Deng et al (2016) showed that high temperature (40°C) induced expression of the PAL-, C4H-, 4CL-, and STS-like genes and increased piceatannol levels in the leaves of Gnetum parvifolium, but did not affect resveratrol levels. However, the following analysis revealed that the high temperature conditions inhibited the accumulation of both resveratrol and piceatannol in G. parvifolium stems and roots, while expression of the stilbene biosynthesis genes was increased with some variability (Deng et al 2017).…”

“…Multiple studies show that UV-C light exposure induced stilbene biosynthesis in different plant species (e.g., grapevine, pine, peanut, and gnetum), and the induction was accompanied by a transcriptional activation, protein accumulation, and enzyme activation of STS and some other stilbene biosynthesis enzymes (Wang et al 2010Tang et al 2010;Suzuki et al 2015;Deng et al 2016Deng et al , 2017Kiselev et al 2017). For grapevine berries, it has been shown that the induction of resveratrol biosynthesis after UV-C irradiation occurred primarily in the form of t-resveratrol (not piceids) and was dependent on the berry developmental stage .…”

Identification of the candidate genes related to sanshool biosynthesis in Zanthoxylum bungeanum by comparative RNA sequencing analysis

“…2a, a total of 76 744 (46.98%) unigenes were successfully annotated in at least one database, 9074 (5.55%) were annotated in all databases, and 86 596 unigenes (53.02%) were not annotated in the public databases chosen in this study. The low rate of annotation may be attributed to the limited genomic information available for Z. bungeanum or because most of unannotated unigenes were too short to generate a blast hit and the same results also appeared in the transcriptomes of Carya illinoinensis 32 , Pseudostellaria heterophylla 12 , and G. parvifolium 15 which were constructed of several libraries and generated many unigenes. Additionally, species distribution analysis based on blast hits against the non-redundant database showed that Citrus sinensis and Citrus clementine were the top-hit species, which are in the same evolutionary unit as Rutaceae with Z. bungeanum 33 , and accounted for 64.83% (34 631) of the identified unigenes.…”

“…To screen and identify the pathways and genes related to the biosynthesis of secondary metabolites, RNA-seq based on high-throughput sequencing technology is a cost-effective and powerful tool to obtain many transcripts from different tissues or the same tissue from different developmental stages, which can be used for differentially expressed gene (DEG) analysis combined with Gene Ontology (GO) classification and Kyoto Encyclopedia of genes and Genomes (KEGG) pathway functional enrichment analysis to identify key genes [12][13][14] . For example, pathways and genes involved in the production of secondary metabolites, such as flavonoid and stil-benoid biosynthesis in Gnetum parvifolium 15 , terpenoid and thenylpropanoid biosynthesis in Ananas comosus 16 and caffeine and theanine biosynthesis in Camellia sinensis 17 have been verified by comparative transcriptome sequencing.…”