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Abstract
As one type of the most important alkaloids in the world, terpenoid indole alkaloids (TIAs) show a wide range of pharmaceutical activities that are beneficial for clinical treatments. Catharanthus roseus produces approximately 130 identified TIAs and is considered to be a model plant to study TIA biosynthesis. In order to increase the production of high medical value metabolites whose yields are extremely low in C. roseus, genetic engineering combined with transcriptional regulation has been applied in recent years. By using bioinformatics which is based on RNA sequencing (RNA-seq) data from methyl jasmonate (MeJA)-treated C. roseus as well as phylogenetic analysis, the present work aims to screen candidate genes that may be involved in the regulation of TIA biosynthesis, resulting in a novel AP2/ERF transcription factor, CR1 (Catharanthus roseus 1). Subsequently, virus-induced gene silencing (VIGS) of CR1 was carried out to identify the involvement of CR1 in the accumulations of several TIAs and quantitative real-time PCR (qRT-PCR) was then applied to detect the expression levels of 7 genes in the related biosynthetic pathway in silenced plants. The results show that all the 7 genes were upregulated in CR1-silenced plants. Furthermore, metabolite analyses indicate that silencing CR1 could increase the accumulations of vindoline and serpentine in C. roseus. These results suggest a novel negative regulator which may be involved in the TIAs biosynthetic pathway.
Highlights
Plants produce a vast number of secondary metabolites, when they adapt themselves to variable ecological environment during long-term evolution
In order to excavate candidate transcription factors that may be involved in the regulation of terpenoid indole alkaloids (TIAs) biosynthesis, we collected the leaves of 1-month-old C. roseus with methyl jasmonate (MeJA) treatment and extracted their RNA to generate the transcriptome sequencing data using Illumina platform
A phylogenetic tree was constructed based on the protein sequences of the three candidate transcription factors, together with the previously reported AP2/ERF transcription factors which are potentially involved in secondary metabolite biosynthesis, to analyze their evolutionary relationships (Figure 3)
Summary
Plants produce a vast number of secondary metabolites, when they adapt themselves to variable ecological environment during long-term evolution. Secondary metabolites are usually derived from primary metabolites and have their own unique metabolic pathways and different properties. They are generally classified into three major groups: phenols, terpenoids and alkaloids. One from Apocynaceae, Catharanthus roseus, produces approximately 130 identified TIAs and is considered to be a model plant to study TIA biosynthesis. Most of the TIAs produced by C. roseus, such as ajmaline, serpentine and catharanthine, show significant clinical medical values, Regulation of TIAs Biosynthesis in Catharanthus roseus vinblastine and vincristine, which are recognized to be the most valuable antineoplastic agents, have been widely used in the world. The yields of these high medical value metabolites are extremely low (Guéritte and Fahy, 2005)
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