Transcriptome profiling shows gene regulation patterns in a flavonoid pathway in response to exogenous phenylalanine in Boesenbergia rotunda cell culture

Abstract

BackgroundPanduratin A extracted from Boesenbergia rotunda is a flavonoid reported to possess a range of medicinal indications which include anti-dengue, anti-HIV, anti-cancer, antioxidant and anti-inflammatory properties. Boesenbergia rotunda is a plant from the Zingiberaceae family commonly used as a food ingredient and traditional medicine in Southeast Asia and China. Reports on the health benefits of secondary metabolites extracted from Boesenbergia rotunda over the last few years has resulted in rising demands for panduratin A. However large scale extraction has been hindered by the naturally low abundance of the compound and limited knowledge of its biosynthetic pathway.ResultsTranscriptome sequencing and digital gene expression (DGE) analysis of native and phenylalanine treated Boesenbergia rotunda cell suspension cultures were carried out to elucidate the key genes differentially expressed in the panduratin A biosynthetic pathway. Based on experiments that show increase in panduratin A production after 14 days post treatment with exogenous phenylalanine, an aromatic amino acid derived from the shikimic acid pathway, total RNA of untreated and 14 days post-phenylalanine treated cell suspension cultures were extracted and sequenced using next generation sequencing technology employing an Illumina-Solexa platform. The transcriptome data generated 101, 043 unigenes with 50, 932 (50.41%) successfully annotated in the public protein databases; including 49.93% (50, 447) in the non-redundant (NR) database, 34.63% (34, 989) in Swiss-Prot, 24,07% (24, 316) in Kyoto Encyclopedia of Genes and Genomes (KEGG) and 16.26% (16, 426) in Clusters of Orthologous Groups (COG). Through DGE analysis, we found that 14, 644 unigenes were up-regulated and 14, 379 unigenes down-regulated in response to exogenous phenylalanine treatment. In the phenylpropanoid pathway leading to the proposed panduratin A production, 2 up-regulated phenylalanine ammonia-lyase (PAL), 3 up-regulated 4-coumaroyl:coenzyme A ligase (4CL) and 1 up-regulated chalcone synthase (CHS) were found.ConclusionsThis is the first report of Boesenbergia rotunda de novo transcriptome data that could serve as a reference for gene or enzyme functional studies in the Zingiberaceae family. Although enzymes that are directly involved in the panduratin A biosynthetic pathway were not completely elucidated, the data provides an overall picture of gene regulation patterns leading to panduratin A production.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2164-15-984) contains supplementary material, which is available to authorized users.

Analyses of Boesenbergia Rotunda Cell Suspension Cultures With Overexpressed Chalcone Synthase (Chs) Manifesting Changes on the Expression Level of Flavonoid-Related Genes

“…3). Compared with previous B. rotunda transcriptome databases (Md-Mustafa et al, 2014), the average length and the N50 of unigenes in this study were much longer with nearly half of total unigenes were more than 1000bp in size. Therefore, all the data generated in this study indicated a relatively good overall integrity of the unigenes.…”

Expression profiles of flavonoid-related gene, 4 coumarate: coenzyme A ligase, and optimization of culturing conditions for the selected flavonoid production in Boesenbergia rotunda

“…For instance, the expression levels of 4CL gene in T10 cultures were up-regulated by twofold in comparison to C10 cultures, whereas T14 cultures were threefold higher than C14 cultures. In line with our previous study (Tan et al 2012;Md-Mustafa et al 2014), these findings indicated the promotive effect of phenylalanine in enhancing the selected flavonoid production.…”

“…Hence, in this study, we analyzed the 4CL gene expression level after adding a precursor, phenylalanine, into B. rotunda cell which grown in full strength MS basal medium containing 1 mg/l NAA, 1 mg/l BA, and 3 % sucrose. Since our previous study showed that the amount of flavonoids produced was highest after 14 days of phenylalanine treatment before the cell growth decreased (Tan 2005;Tan et al 2012;Md-Mustafa et al 2014), the analysis of 4CL gene expression in this study was performed at 10 and 14 days of phenylalanine treatment.…”

Transcriptome profiling shows gene regulation patterns in ginsenoside pathway in response to methyl jasmonate in Panax Quinquefolium adventitious root

“…For medicinal plants, RNA-seq has been used to identify genes that are directly or indirectly involved in the biosynthetic pathways of target bioactive compounds22. Transcriptome analysis from only one plant organ or tissue does not provide a full transcript catalogue, even though it can serve numerous specific genetic and breeding objectives.…”

“…Primary metabolism is essential for plant growth, plant development and plant reproduction. In cell suspension cultures, primary metabolism is essential for plant cells to propagate in liquid media22.…”