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

Abstract

Background: Panduratin 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. Results: Transcriptome 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. Conclusions: This 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.