Integrated Transcriptomics and Proteomics to Reveal Regulation Mechanism and Evolution of SmWRKY61 on Tanshinone Biosynthesis in Salvia miltiorrhiza and Salvia castanea.

Yanbo Huang,Xiaodan Zhang,Zhechen Qi,Dongfeng Yang,Yue Chen,Jian Yang,Ying Cheng,Yukun Wei,Yanting Wang,Mans Dennis,Luqi Huang,Zixuan Wang,Zongsuo Liang,Zewei Du,Juan Guo

doi:10.3389/fpls.2021.820582

Abstract

Tanshinones found in Salvia species are the main active compounds for the treatment of cardiovascular and cerebrovascular diseases, but their contents are hugely different in different species. For example, tanshinone IIA content in Salvia castanea Diels f. tomentosa Stib. is about 49 times higher than that in Salvia miltiorrhiza Bunge. The molecular mechanism responsible for this phenomenon remains largely unknown. To address this, we performed comparative transcriptomic and proteomic analyses of S. miltiorrhiza and S. castanea. A total of 296 genes in S. castanea and 125 genes in S. miltiorrhiza were highly expressed at both the transcriptional and proteome levels, including hormone signal regulation, fungus response genes, transcription factors, and CYP450. Among these differentially expressed genes, the expression of SmWRKY61 was particularly high in S. castanea. Overexpression of SmWRKY61 in S. miltiorrhiza could significantly increase the content of tanshinone I and tanshinone IIA, which were 11.09 and 33.37 times of the control, respectively. Moreover, SmWRKY61 had a strong regulatory effect, elevating the expression levels of tanshinone pathway genes such as DXS2, CMK, HMGS2, 1, KSL1, KSL2, CYP76AH1, and CYP76AK3. For the WRKY family, 79 SmWRKYs were originally obtained and classified into three main groups. Collinearity analysis indicated a more specific extension of WRKY gene family in Salvia genus. In 55 Salvia species, only 37 species contained the WRKY61 sequence, and high SmWRKY61 expression in some Salvia L. species was often accompanied by high tanshinone accumulation. The above results suggest that SmWRKY61 is a highly effective regulator of tanshinone accumulation and may be a key factor resulting in high tanshinone accumulation in S. castanea.

Highlights

Salvia miltiorrhiza Bunge (S. miltiorrhiza), a perennial herb in Salvia L. (Lamiaceae: Nepetoideae: Mentheae: Salviinae) (Xu et al, 2016), has high medicinal and economic value (Ma et al, 2012)
The distribution pattern and accumulation of tanshinones in the roots of 37 Salvia species were analyzed by HPLC
We found that tanshinones were widely distributed in the roots of Salvia species from southwestern China such as S. przewalskii, S. trijuga, and S. castanea

Summary

Introduction

Salvia miltiorrhiza Bunge (S. miltiorrhiza), a perennial herb in Salvia L. (Lamiaceae: Nepetoideae: Mentheae: Salviinae) (Xu et al, 2016), has high medicinal and economic value (Ma et al, 2012). (Lamiaceae: Nepetoideae: Mentheae: Salviinae) (Xu et al, 2016), has high medicinal and economic value (Ma et al, 2012). It is known for its pharmacologically effective chemical components (Fang et al, 2018), primarily diterpene quinone compounds (e.g., tanshinone IIA, cryptotanshinone, dihydrotanshinone I, isocryptotanshinone, and przewaquinone) and polyphenolic compounds (e.g., rosmarinic acid, caffeic acid, and salvianolic acid) (Kai et al, 2011). MVA and MEP pathways result in the formation of intermediates, isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP) (Shi et al, 2016). The CPS and KSL may act as important key enzymes for the formation of various diterpenoids in S. miltiorrhiza, including tanshinones, gibberellins, and ent-13-epi-manoyl oxide (Gao et al, 2009). Many downstream steps of the tanshinone biosynthetic pathway may be catalyzed by cytochrome P450 (CYP450s), including CYP76AHs, CYP76AKs, and CYP71Ds (Guo et al, 2013; Zi et al, 2014; Guo et al, 2016; Ma et al, 2021)

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Results

Conclusion