Identification of Abietane-Type Diterpenoids and Phenolic Acids Biosynthesis Genes in Salvia apiana Jepson Through Full-Length Transcriptomic and Metabolomic Profiling

Abstract

Salvia apiana (S. apiana) Jepson is a medicinal plant that is frequently used by the Chumash Indians in southern California as a diaphoretic, calmative, diuretic, or antimicrobial agent. Abietane-type diterpenoids (ATDs) and phenolic acids (PAs) are the main bioactive ingredients in S. apiana. However, few studies have looked into the biosynthesis of ATDs and PAs in S. apiana. In this study, using metabolic profiling focused on the ATDs and PAs in the roots and leaves of S. apiana, we found a distinctive metabolic feature with all-around accumulation of ATDs, but absence of salvianolic acid B. To identify the candidate genes involved in these biosynthesis pathways, full-length transcriptome was performed by PacBio single-molecule real-time (SMRT) sequencing. A total of 50 and 40 unigenes were predicted to be involved in ATDs and PAs biosynthesis, respectively. Further transcriptional profile using Illumina HiSeq sequencing showed that the transcriptional variations of these pathways were consistent with the accumulation patterns of corresponding metabolites. A plant kingdom-wide phylogenetic analysis of cytochromes (CYPs) identified two CYP76AK and two CYP76AH subfamily genes that might contribute for the specific ATDs biosynthesis in S. apiana. We also noticed that the clade VII laccase gene family was significantly expanded in Salvia miltiorrhiza compared with that of S. apiana, indicating their involvements in the formation of salvianolic acid B. In conclusion, our results will enable the further understanding of ATDs and PAs biosynthesis in S. apiana and Salvia genus.

Comparative transcriptomics of two Salvia subg. Perovskia species contribute towards molecular background of abietane-type diterpenoid biosynthesis

“…Additionally, it has been shown that the catalytic promiscuity of CYP450s creates a metabolic grid for tanshinone biosynthesis 52 , 63 . Nonetheless, despite numerous experimental strategies to elucidate the tanshinone biosynthesis pathway has been implemented to date, including: investigation of a spatial organization of the diterpenoid biosynthetic gene cluster 64 , comparative metabolomic, transcriptomics and proteomic analysis 33 , 40 , 44 , 65 , targeted mutagenesis approaches using RNA interference and CRISPR/Cas9 system 41 , 66 , 67 , hairy roots cultures and elicitor treatments 31 , 68 – 74 , or endophytic fungi infection 75 , several steps from the pathway remain unresolved. Particularly, the most intriguing transformation – the hypothetical conversion of 11,20-dihydroxyferruginol to miltirone, which requires loss of C20 (directly via demethylation or via decarboxylation of carnosic acid as an intermediate) and aromatization of the “B” ring, has not been determined yet.…”

“…Multiple pharmacological studies have proven that tanshinones possess various biological activities including antibacterial, antioxidant, anti-inflammatory and anti-tumor effects 34 – 36 . Due to the structural uniqueness of tanshinones, their biosynthesis has been studied intensively in the last two decades, in several species of Lamiaceae , incuding: S.miltiorrhiza 30 , 33 , 37 – 41 , Salvia rosmarinus 26 , 42 , Salvia pomifera 43 , Salvia fruticosa 26 or Salvia apiana 44 . Recently, a comprehensive, in-depth review of the tanshinone biosynthesis has been published 45 .…”

Comparative Transcriptomic Analysis of two Asian Medicinal and Aromatic Species of Salvia subg. Perovskia – a Contribution Towards Molecular Background of Abietane-type Diterpenoid Biosynthesis

“…comparative metabolomic, transcriptomics and proteomic analysis [33,40,44,66], targeted mutagenesis approaches using RNA interference and CRISPR/Cas9 system [41,67,68], hairy roots cultures and elicitor treatments [31,[69][70][71][72][73][74][75], or endophytic fungi infection [76], several steps from the pathway remain unresolved. Particularly, the most intriguing transformation -the hypothetical conversion of 11,20-dihydroxyferruginol to miltirone, which requires loss of C20 (directly via demethylation or via decarboxylation of carnosic acid as an intermediate) and aromatization of the "B" ring, has not been determined yet.…”

“…Multiple pharmacological studies have proven that tanshinones possess various biological activities including antibacterial, antioxidant, anti-in ammatory and anti-tumor effects [34][35][36]. Due to the structural uniqueness of tanshinones, their biosynthesis has been studied intensively in the last two decades, in several species of Lamiaceae, incuding: S.miltiorrhiza [30,33,[37][38][39][40][41], Salvia rosmarinus [26,42], Salvia pomifera [43], Salvia fruticosa [26] or Salvia apiana [44]. Recently, a comprehensive, in-depth review of the tanshinone biosynthesis has been published [45].…”

Integration of high-throughput omics technologies in medicinal plant research: The new era of natural drug discovery

“…In commercial Danshen ( Salvia ) decoctions, phenolic acids, especially salvianolic acid, are the major marker component used for quality assessment according to the official Chinese Pharmacopoeia. A proposed biosynthetic pathway of phenolic acids in Salvia apiana has been schemed based on full-length transcriptomic and metabolomic profiling ( Shi et al., 2021 ; Hu et al., 2022 ). In addition, functional genes and transcription factors as regulators of phenolic acids biosynthesis in Salvia miltiorrhiza have been reported ( Yu et al., 2018 ; Deng et al., 2020 ; Zhou et al., 2021c ).…”