Identification of potential genes involved in triterpenoid saponins biosynthesis in Gleditsia sinensis by transcriptome and metabolome analyses

Yusuke Kuwahara,Mami Yamazaki,Noriaki Kawano,Nobuo Kawahara,Sayaka Shinpo,Michimi Nakamura,Daisuke Nakajima,Hideki Hirakawa,Hideyuki Suzuki,Kazuki Saito

doi:10.1007/s11418-018-1270-2

Abstract

Gleditsia sinensis is widely used as a medicinal plant in Asia, especially in China. Triterpenes, alkaloids, and sterols were isolated from Gleditsia species. Among them, triterpenoid saponins are very important metabolites owing to their various pharmacological activities. However, the triterpenoid saponin biosynthesis pathway has not been well characterized. In the present study, we performed de novo transcriptome assembly for 14.3 Gbps of clean reads sequenced from nine tissues of G. sinensis. The results showed that 81,511 unique transcripts (unitranscripts) (47,855 unigenes) were constructed, of which 31,717 unigenes were annotated with Gene Ontology and EC numbers by Blast2GO against the NCBI-nr protein database. We also analyzed the metabolite contents in the same nine tissues by LS–MS/MS, and saponins including gleditsioside I were found in fruit at higher levels. Many of the genes with tissue-specific expression in fruit are involved in the flavonoid biosynthesis pathway, and many of those have UDP-glucosyltransferase (UGT) activity. We constructed a saponin biosynthesis pathway and identified two key enzyme families in the triterpenoid saponin biosynthesis pathway, cytochrome P450 and UDP-glucosyltransferase, that are encoded by 37 unigenes and 77 unigenes, respectively. CYP72A, CYP716A, and CYP88D, which are known as key enzymes for saponin biosynthesis, were also identified among the P450s. Our results provide insight into the secondary metabolite biosynthesis and serve as important resources for future research and cultivation of G. sinensis.

Highlights

Gleditsia is a genus from the legume family (Fabaceae) and grown worldwide, especially in Asia and America
A total of 14.3 Gbp reads from the nine tissues of G. sinensis were checked for quality and applied to the transcriptome analysis
The top 20 GO terms at level 3 based on the number of assigned unigenes for biological process (BP), molecular function (MF), and cellular component (CC) were investigated using Blast2GO

Summary

Introduction

Gleditsia is a genus from the legume family (Fabaceae) and grown worldwide, especially in Asia and America. Many species in the Gleditsia genus have been identified, including G. sinensis, G. japonica, and G. triacanthos, and these and other Gleditsia species are used as traditional diuretics and expectorants [1]. Recent phytochemical studies have revealed the presence of triterpenoid saponins, alkaloids, sterols, and flavonoids in these species. The extracts of these compounds exhibit several important biological activities, including antitumor, anti-inflammatory, and anti-HIV activities [2,3,4]. Over 30 compounds of triterpenoid saponins were identified in the tissues of G. sinensis [1], but the biosynthesis of these physiologically active substances is not yet fully understood.

Methods

Results

Conclusion