Integrated Metabolomic and Transcriptomic Analysis of triterpenoid Accumulation in the Roots of Codonopsis pilosula. var. modesta (Nannf.) L.T.Shen at Different Altitudes

Abstract Background: Codonopsis Radix is a benefit Traditional Chinese Medicine and triterpenoid are the major bioactive constituents of Codonopsis Radix. Codonopsis pilosula. var. modesta(Nannf.) L.T. Shen (CPM)is a precious variety and the legal source of Codonopsis Radix, and it roots are known as Wen Dangshen, which is distribute in high mountains area and the altitudes are ranging from 1300 to 4300 m. Environment plays an important role in the synthesis and metabolism of active ingredients in medicinal plants, but there is no report elaborate the effect of altitude on terpenoid metabolites accumulation in CPM. In this study, we integrated metabolomic and transcriptomic to explain the effects of altitude on terpenoid biosynthetic pathways and secondary metabolite accumulation in CPM, fresh root samples from CPM grown at low altitude (1480 m) and high altitude (2300m) at the same harvest stage were selected for analysis. Results: Untargeted metabolic results were shown that there were significant differences in the total secondary metabolites between high altitude and low altitude CPM group. Based on our laboratory previously established UPLC-Q-TOF-MS method, 10 triterpenoids in the above two altitude CPM fresh roots were quantitatively analyzed. According to their chemical structure and isomerism, they can be divided into 6 categories including Gansuidine-type tetracyclic triterpenes (codopitirol A), Cycloaneurane tetracyclic triterpenes (24-methylenecycloartanol), Xylorane-type pentacyclic triterpenes (kokoonol, friedel-1-en-3-one, friedelin), Dandelion pentacyclic triterpenoids (codopimodol A, taraxerol), Oleanocarpine pentacyclic triterpenoids (bryonolol, glut 5-en-3β-ol), Ursulane-type pentacyclic triterpenes (α-amyrin), our result showed that there are significant differences in the content and the types of terpenoids between different altitude CPM and the content were higher in the high altitude samples ofCPM. The results of transcriptome study showed that CPM could significantly up-regulate the gene expression levels of seven key enzymes in the biosynthetic pathway of triterpenoid precursor substances. These enzymes include isoprenyl diphosphate isomerase (IDI), geraniol pyrophosphate synthase (GPPS), farnesyl pyrophosphate synthase (FPPS), farnesyl pyroacyltransferase (SS), squalene synthase (SE), beta-amyrin synthase (β-AS) and lupiol synthase (LS). The quantitative polymerase chain reaction was used to further verify the accuracy of the transcriptome data. Conclusions: This study shows that there were significant differences between high altitude and low altitude groups of CPM, which was manifested in the following aspects including overall secondary metabolites, content and types of triterpenoids, and gene expression levels. The CPM at high altitude were more likely to accumulate triterpenes than those at low altitude, which was related to the up-regulation of the gene expression levels of seven key enzymes in the triterpenoid precursor biosynthetic pathway, thereby increasing the accumulation of triterpenoids. These results expand our understanding of how altitude affects plant metabolite biosynthesis.