Computational Identification and Systematic Classification of Novel Cytochrome P450 Genes in Salvia miltiorrhiza.

Bin Wu,Mukesh Jain,David R Nelson,Chang Liu,Haimei Chen,Kai Wu

doi:10.1371/journal.pone.0115149

Abstract

Salvia miltiorrhiza is one of the most economically important medicinal plants. Cytochrome P450 (CYP450) genes have been implicated in the biosynthesis of its active components. However, only a dozen full-length CYP450 genes have been described, and there is no systematic classification of CYP450 genes in S. miltiorrhiza. We obtained 77,549 unigenes from three tissue types of S. miltiorrhiza using RNA-Seq technology. Combining our data with previously identified CYP450 sequences and scanning with the CYP450 model from Pfam resulted in the identification of 116 full-length and 135 partial-length CYP450 genes. The 116 genes were classified into 9 clans and 38 families using standard criteria. The RNA-Seq results showed that 35 CYP450 genes were co-expressed with CYP76AH1, a marker gene for tanshinone biosynthesis, using r≥0.9 as a cutoff. The expression profiles for 16 of 19 randomly selected CYP450 obtained from RNA-Seq were validated by qRT-PCR. Comparing against the KEGG database, 10 CYP450 genes were found to be associated with diterpenoid biosynthesis. Considering all the evidence, 3 CYP450 genes were identified to be potentially involved in terpenoid biosynthesis. Moreover, we found that 15 CYP450 genes were possibly regulated by antisense transcripts (r≥0.9 or r≤–0.9). Lastly, a web resource (SMCYP450, http://www.herbalgenomics.org/samicyp450) was set up, which allows users to browse, search, retrieve and compare CYP450 genes and can serve as a centralized resource.

Highlights

Salvia miltiorrhiza Bunge (Chinese name: Danshen) is a perennial plant belonging to the Lamiaceae family
Despite several studies focusing on the identification and characterization of Cytochrome P450 (CYP450) genes from S. miltiorrhiza, information relating to these genes is scattered in the literature, and many of their sequences remain proprietary and not accessible to the general public
A lack of a systematic nomenclature, as well as a centralized resource for CYP450 genes from S. miltiorrhiza, has become a bottleneck in research efforts attempting to elucidate the biosynthetic pathways for active compounds in S. miltiorrhiza

Summary

Introduction

Salvia miltiorrhiza Bunge (Chinese name: Danshen) is a perennial plant belonging to the Lamiaceae family. The root of S. miltiorrhiza has been used as a traditional Chinese medicine in the treatment of dysmenorrhea, amenorrhea, cardiovascular diseases, blood circulation disturbances, inflammation, and angina pectoris [1]. The active components of S. miltiorrhiza consist of water-soluble and lipid-soluble compounds. The lipid-soluble tanshinones, such as tanshinone I, tanshinone IIA, tanshinone IIB, and cryptotanshinone, are abietane-type norditerpenoid naphthoquinones, which are considered as the main lipophilic bioactive components of S. miltiorrhiza [2]. The annual sales of medicinal products containing active components of S. miltiorrhiza exceed US$ 1 billion. Due to the medical and economic importance of tanshinones, increasing the production of these compounds through marker-assisted breeding and metabolic engineering has become an active area of research. The identification of all genes involved in the biosynthesis of tanshinones and the elucidation of the corresponding pathways are critical for achieving these goals

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Conclusion