Terpenoid Biosynthesis In Plants Research Articles

Contemporary understanding of transcription factor regulation of terpenoid biosynthesis in plants.

This review summarized how TFs function independently or in response to environmental factors to regulate terpenoid biosynthesis via fine-tuning the expression of rate-limiting enzymes. Terpenoids are derived from various species and sources. They are essential for interacting with the environment and defense mechanisms, such as antimicrobial, antifungal, antiviral, and antiparasitic properties. Almost all terpenoids have high medicinal value and economic performance. Recently, the control of enzyme genes on terpenoid biosynthesis has received a great deal of attention, but transcriptional factors regulatory network on terpenoid biosynthesis and accumulation has yet to get a thorough review. Transcription factors function as activators or suppressors independently or in response to environmental stimuli, fine-tuning terpenoid accumulation through regulating rate-limiting enzyme expression. This study investigates the advancements in transcription factors related to terpenoid biosynthesis and systematically summarizes previous works on the specific mechanisms of transcription factors that regulate terpenoid biosynthesis via hormone signal-transcription regulatory networks in plants. This will help us to better comprehend the regulatory network of terpenoid biosynthesis and build the groundwork for terpenoid development and effective utilization.

Effect of interfering TOR signaling pathway on the biosynthesis of terpenoids in Salvia miltiorrhiza Bge

ABSTRACT The TOR (Target of Rapamycin) signaling pathway, which takes TOR kinase as the core, regulates the absorption, distribution, and recycling of nutrients by integrating metabolic network and other signaling pathways, thus participating in the plant growth-defense trade-off. While terpenoids play an important role in plant growth, development, stress response, and signal transduction. The effect of the TOR signaling pathway on terpenoid biosynthesis in plants has yet to be studied in detail. In this study, the tissue culture seedlings of Salvia miltiorrhiza were treated with the TOR inhibitor AZD8055. The results show that the roots of the control group had begun to grow on the 8th day, while the seedlings treated with AZD8055 had no rooting signs. Combined with the expression changes of genes related to the TOR signaling pathway in the first 8 days, samples on the 3rd, 6th, and 8th days were selected for RNA-Seq analysis. Through RNA-Seq analysis, a total of 50,689 unigenes were obtained from the samples of these three periods, of which 4088 unigenes showed differential expression. The function enrichment and time-series analysis of differentially expressed genes (DEGs) showed that the main influence of the TOR signal pathway on plant growth-related processes was gradually transmitted with treatment time after TOR was inhibited. Pathway enrichment analysis of DEGs showed that the genes in the biosynthesis of terpenoids, such as diterpenoid and carotenoid biosynthetic pathways, could be regulated. Compared with other stages, DEGs related to terpenoid biosynthesis were mainly regulated in the S2 stage. In addition, the genes involved in terpenoid skeleton biosynthesis was also considerably enriched in the S2 stage, according to the results of gene set enrichment analysis (GSEA) of unigenes. Inhibition of the TOR signaling pathway may affect the biosynthesis of terpenoid signaling molecules, inhibit gibberellin’s biosynthesis, and promote abscisic acid’s biosynthesis. This study has discussed the effect of interfering with the TOR pathway on terpenoid biosynthesis in S. miltiorrhiza from the perspective of omics and provides new insight into the interaction between the terpenoid biosynthesis pathway and the growth-defense trade-off of medicinal plants.

The PcbZIP44 transcription factor inhibits patchoulol synthase gene expression and negatively regulates patchoulol biosynthesis in Pogostemon cablin

Patchouli alcohol (patchoulol), a sesquiterpenoid specifically synthesized by herbal plant Pogostemon cablin, has a variety of pharmacological and biological activities and is widely utilized in the medical and cosmetic industries. However, there are few studies on the transcriptional modulation of patchoulol biosynthesis. Some studies have reported that basic leucine zipper (bZIP) transcription factors (TFs) participate in the biosynthesis of plant terpenoids, but there is no report that bZIP TFs are involved in the modulation of patchoulol biosynthesis. This study was conducted to explain the regulation of patchoulol biosynthesis by PcbZIP44, a bZIP TF identified in patchouli for the first time. PcbZIP44 was highly expressed in roots and stems, and its encoded protein was nuclear localized revealed by protoplast subcellular localization experiment. According to the results of the transient dual-luciferase assay and yeast one-hybrid (Y1H) assays, PcbZIP44 can bind to the PcPTS (patchoulol synthase) gene promoter to repress its activity. Overexpression of the PcbZIP44 gene significantly reduced the content of patchoulol and resulted in a significant down-regulation of the PcPTS gene at the transcriptional level. Correspondingly, virus-induced PcbZIP44 gene silencing (VIGS) significantly increased the content of patchoulol and resulted in a significant up-regulation of the PcPTS gene at the transcriptional level. Therefore, these results suggested that PcbZIP44 negatively regulates patchoulol biosynthesis by inhibiting the PcPTS gene in P. cablin.

The transcription factor LaMYC4 from lavender regulates volatile Terpenoid biosynthesis

BackgroundThe basic helix-loop-helix (bHLH) transcription factors (TFs), as one of the largest families of TFs, are essential regulators of plant terpenoid biosynthesis and response to stresses. Lavender has more than 75 volatile terpenoids, yet few TFs have been identified to be involved in the terpenoid biosynthesis.ResultsBased on RNA-Seq, reverse transcription-quantitative polymerase chain reaction, and transgenic technology, this study characterized the stress-responsive transcription factor LaMYC4 regulates terpenoid biosynthesis. Methyl jasmonate (MeJA) treatment increased volatile terpenoid emission, and the differentially expressed gene LaMYC4 was isolated. LaMYC4 expression level was higher in leaf than in other tissues. The expression of LaMYC4 decreased during flower development. The promoter of LaMYC4 contained hormone and stress-responsive regulatory elements and was responsive to various treatments, including UV, MeJA treatment, drought, low temperature, Pseudomonas syringae infection, and NaCl treatment. LaMYC4 overexpression increased the levels of sesquiterpenoids, including caryophyllenes, in Arabidopsis and tobacco plants. Furthermore, the expression of crucial node genes involved in terpenoid biosynthesis and glandular trichome number and size increased in transgenic tobacco.ConclusionsWe have shown that the stress-responsive MYC TF LaMYC4 from ‘Jingxun 2’ lavender regulates volatile terpenoid synthesis. This study is the first to describe the cloning of LaMYC4, and the results help understand the role of LaMYC4 in terpenoid biosynthesis.

Expression of genes encoding terpenoid biosynthesis enzymes during leaf development of Eucalyptus camaldulensis

To reveal the regulation mechanism of terpenoid biosynthesis in the leaves of Eucalyptus camaldulensis, the content of volatiles in eucalyptus leaves and the transcriptome databases of young and mature leaves were analyzed. The results showed that E. camaldulensis contains 92 and 89 kinds of volatile substances in the young and mature leaves, respectively. Among them, the content of 1,8-cineole, β-pinene, and other substances was significantly different in young and mature leaves. A total of 99 802 unigenes were obtained from the transcriptome database of young and mature leaves of E. camaldulensis and 18 441 genes displayed obviously differential expressions during both developmental stages. There were 6 982 up-regulated unigenes and 11 461 down-regulated unigenes in the young leaf stage compared to the mature leaf stage. The key genes for terpenoid biosynthesis, including limonene synthase-10, limonene synthase-11, myrcene synthase-1, α- pinene synthase-2, and 1,8-cineole synthase-2, were selected for further analysis to explore the mechanism of gene regulation and genetic transformation. The expressions of key genes were validated by RT-qPCR, and their expressions were consistent with RNA-seq data. WRKY, MYB, NAC, and bHLH transcription factors (TFs) displayed important regulatory effects on the above key genes. Thus, a regulatory network model of terpenoid biosynthesis was constructed using target genes and TFs during leaf development in E. camaldulensis. These results provide theoretical evidence for understanding the terpenoid biosynthesis in plants and reference for terpenoids utilization by genetic engineering methods in E. camaldulensis.

A systematic review of 1-Deoxy-D-xylulose-5-phosphate synthase in terpenoid biosynthesis in plants

A systematic review of 1-Deoxy-D-xylulose-5-phosphate synthase in terpenoid biosynthesis in plants

A Cryptic Plant Terpene Cyclase Producing Unconventional 18- and 14-Membered Macrocyclic C25 and C20 Terpenoids with Immunosuppressive Activity.

A versatile terpene synthase (LcTPS2) producing unconventional macrocyclic terpenoids was characterized from Leucosceptrum canum. Engineered Escherichia coli and Nicotiana benthamiana expressing LcTPS2 produced six 18-/14-membered sesterterpenoids including five new ones and two 14-membered diterpenoids. These products represent the first macrocyclic sesterterpenoids from plants and the largest sesterterpenoid ring system identified to date. Two variants F516A and F516G producing approximately 3.3- and 2.5-fold, respectively, more sesterterpenoids than the wild-type enzyme were engineered. Both 18- and 14-membered ring sesterterpenoids displayed significant inhibitory activity on the IL-2 and IFN-γ production of T cells probably via inhibition of the MAPK pathway. The findings will contribute to the development of efficient biocatalysts to create bioactive macrocyclic sesterterpenoids, and also herald a new potential in the well-trodden territory of plant terpenoid biosynthesis.

A Cryptic Plant Terpene Cyclase Producing Unconventional 18‐ and 14‐Membered Macrocyclic C25 and C20 Terpenoids with Immunosuppressive Activity

AbstractA versatile terpene synthase (LcTPS2) producing unconventional macrocyclic terpenoids was characterized from Leucosceptrum canum. Engineered Escherichia coli and Nicotiana benthamiana expressing LcTPS2 produced six 18‐/14‐membered sesterterpenoids including five new ones and two 14‐membered diterpenoids. These products represent the first macrocyclic sesterterpenoids from plants and the largest sesterterpenoid ring system identified to date. Two variants F516A and F516G producing approximately 3.3‐ and 2.5‐fold, respectively, more sesterterpenoids than the wild‐type enzyme were engineered. Both 18‐ and 14‐membered ring sesterterpenoids displayed significant inhibitory activity on the IL‐2 and IFN‐γ production of T cells probably via inhibition of the MAPK pathway. The findings will contribute to the development of efficient biocatalysts to create bioactive macrocyclic sesterterpenoids, and also herald a new potential in the well‐trodden territory of plant terpenoid biosynthesis.

Endophytes: the novel sources for plant terpenoid biosynthesis.

Terpenoids are natural compounds predominantly present in plants. They have many pharmaceutical and/or nutritional functions, and have been widely applied in medical, food, and cosmetics industries. Recently, terpenoids have been used in the clinical treatment of COVID-19 due to the good antiviral activities. The increasing demand for terpenoids in international markets poses a serious threat to many plant species. For environmentally sustainable development, microbial cell factories have been utilized as the promising platform to produce terpenoids. Nevertheless, the bioproduction of most terpenoids cannot meet commercial requirements due to the low cost-benefit ratio until now. The biosynthetic potential of endophytes has gained attention in recent decades owing to the continual discovery of endophytes capable of synthesizing plant bioactive compounds. Accordingly, endophytes could be alternative sources of terpenoid-producing strains or terpenoid synthetic genes. In this review, we summarized the research progress describing the main and supporting roles of endophytes in terpenoid biosynthesis and biotransformation, and discussed the current problems and challenges which may prevent the further exploitation. This review will improve our understanding of endophyte resources for terpenoid production in industry in the future.Graphical abstract The four main research interests on endophytes for terpenoid production. A: Isolation of terpenoid-producing endophytes; B: The heterologous expression of endophyte-derived terpenoid synthetic genes; C: Endophytes promoting their hosts’ terpenoid production. The blue dashed arrows indicate signal transduction; D: Biotransformation of terpenoids by endophytes or their enzymes. Key points • The mechanisms employed by endophytes in terpenoid synthesis in vivo and in vitro.• Endophytes have the commercial potentials in terpenoid bioproduction and biotransformation.• Synthetic biology and multiomics will improve terpenoid bioproduction in engineered cell factories.

Research advances in cytochrome P450-catalysed pharmaceutical terpenoid biosynthesis in plants.

Terpenoids, the biggest class of plant secondary metabolites, have a wide range of significant physiological roles, while many of them are important natural drugs. Biosynthesis of pharmaceutical terpenoids in plants is a fairly complex process, most of which involves cytochrome P450 (CYP450) monooxygenases. CYP450 enzymes are versatile biocatalysts that play critical roles in terpenoid skeleton modification and structural diversity. Therefore, the discovery and identification of CYP450 genes is significant for elucidating the terpenoid biosynthetic pathway. This review summarizes the progress and cloning strategies relating to CYP450s in pharmaceutical terpenoid biosynthesis of the past decade.

Rose floral scent

The rose is the queen of flowers and is widely used as a garden plant and for the cut flower market. Roses are also used for the production of essential oil for the cosmetic and perfume industries. A lot of botanical roses are scented and use their volatiles to attract pollinators. Fragrances in garden roses are very diverse and scent has always been an important character in the selection process. Breeders have recently tried to introduce new fragrances, for instance reminiscent of fruit or spice odors. But despite their efforts, some roses on the market are not very fragrant, especially the ones selected for the cut flower market. The cause of this lack of scent is not known. In modern roses or Hybrid Tea roses, scent is mainly produced by petals, although stamens can also contribute to the emission of volatiles. Scent compounds are concentrated in rose petal epidermis, probably in small lipid droplets. In spite of numerous chemical studies, the biosynthetic pathways of many rose scent compounds are unknown. We are studying several genes involved in the biosynthesis of scent in rose. For example, we recently characterized the enzymes responsible for the so-called “tea scent” emitted by Chinese roses. Apart from “tea scent”, terpenoids, especially monoterpenes, are also major constituents of rose flowers, mostly responsible for the “typical rose scent”. Generally, terpenoid biosynthesis in plants is achieved by various terpene synthases. However, with a combination of transcriptomic and genetic approaches, our research group recently discovered a terpene synthase-independent pathway. A key enzyme of this pathway is RhNUDX1, belonging to the Nudix protein family. It has geranyl diphosphate diphosphohydrolase activity in vitro. A positive correlation was found between the expression levels of RhNUDX1 gene and the production of the monoterpene geraniol, indicating its essential role in scent production in roses.

Clade IVa Basic Helix-Loop-Helix Transcription Factors Form Part of a Conserved Jasmonate Signaling Circuit for the Regulation of Bioactive Plant Terpenoid Biosynthesis.

Plants produce many bioactive, specialized metabolites to defend themselves when facing various stress situations. Their biosynthesis is directed by a tightly controlled regulatory circuit that is elicited by phytohormones such as jasmonate (JA). The basic helix-loop-helix (bHLH) transcription factors (TFs) bHLH iridoid synthesis 1 (BIS1) and Triterpene Saponin Activating Regulator (TSAR) 1 and 2, from Catharanthus roseus and Medicago truncatula, respectively, all belong to clade IVa of the bHLH protein family and activate distinct terpenoid pathways, thereby mediating monoterpenoid indole alkaloid (MIA) and triterpene saponin (TS) accumulation, respectively, in these two species. In this study, we report that promoters of the genes encoding the enzymes involved in the specific terpenoid pathway of one of these species can be transactivated by the orthologous bHLH factor from the other species through recognition of the same cis-regulatory elements. Accordingly, ectopic expression of CrBIS1 in M. truncatula hairy roots up-regulated the expression of all genes required for soyasaponin production, resulting in strongly increased levels of soyasaponins in the transformed roots. Likewise, transient expression of MtTSAR1 and MtTSAR2 in C. roseus petals led to up-regulation of the genes involved in the iridoid branch of the MIA pathway. Together, our data illustrate the functional similarity of these JA-inducible TFs and indicate that recruitment of defined cis-regulatory elements constitutes an important aspect of the evolution of conserved regulatory modules for the activation of species-specific terpenoid biosynthesis pathways by common signals such as the JA phytohormones.

Structural Analyses of Short-Chain Prenyltransferases Identify an Evolutionarily Conserved GFPPS Clade in Brassicaceae Plants

Terpenoids are the largest and most diverse class of plant-specialized metabolites, which function in diverse physiological processes during plant development. In the biosynthesis of plant terpenoids, short-chain prenyltransferases (SC-PTs), together with terpene synthases (TPSs), play critical roles in determining terpenoid diversity. SC-PTs biosynthesize prenyl pyrophosphates with different chain lengths, and these compounds are the direct precursors of terpenoids. Arabidopsis thaliana possesses a subgroup of SC-PTs whose functions are not clearly known. In this study, we focus on 10 geranylgeranyl pyrophosphate synthase-like [GGPPSL] proteins, which are commonly thought to produce GGPP [C20]. We found that a subset of members of the Arabidopsis GGPPSL gene family have undergone neo-functionalization: GGPPSL6, 7, 9, and 10 mainly have geranylfarnesyl pyrophosphate synthase activity (C25; renamed AtGFPPS1, 2, 3, and 4), and GGPPSL8 produces even longer chain prenyl pyrophosphate (≥C30; renamed polyprenyl pyrophosphate synthase 2, AtPPPS2). By solving the crystal structures of AtGFPPS2, AtPPPS2, and AtGGPPS11, we reveal the product chain-length determination mechanism of SC-PTs and interpret it as a “three floors” model. Using this model, we identified a novel GFPPS clade distributed in Brassicaceae plants and found that the GFPPS gene typically occurs in tandem with a gene encoding a TPS, forming a GFPPS–TPS gene cluster.

Cloning and expression analysis of 4-diphosphocytidyl-2-C-methyl-D-erythritol kinase gene in Cinnamomum camphora

The 4-diphosphocytidyl-2-C-methyl-D-erythritol kinase was the fourth key enzymes in plant terpenoid biosynthesis pathway of methyl erythritol phosphate pathway(MEP). According to the study of Cinnamomum camphora transcriptome data,we abtained the 4-diphosphocytidyl-2-C-methyl-D-erythritol kinase gene using RT-PCR,and named CcCMK1,then deposited it in GeneBank(Accession number： Ku376098).Bioinformatics analysis showed the open reading frame (ORF) of the CcCMK1 was 1 212 bp.The putative protein encoded 403 amino acids,and its molecular weight was 44.46 kDa and theoretically isoelectric point was 4.99.Transmembrane structure analysis showed that there was no transmembrane structure. Signal peptide analysis showed that it was a non secretory protein, and there was no signal peptide. The subcellular localization showed that the chloroplast was located in the chloroplast.Analysis of the expression of CcCMK1 gene in five chemotypes of C. camphora using Real-time PCR showed its expression level was highest in C. longepaniculatum, and the lowest in Borneol camphor.This research provided a basis for characterizing the key enzyme genes of terpenoid biosynthetic pathway in C. camphora.

Post-modification Enzymes Involved in the Biosynthesis of Plant Terpenoids

Terpenoids are called terpenome for their structural diversities. Besides their important roles in plant growth, development and environmental responses, terpenoids have been widely used in medicine and organic chemicals. The biosynthesis of terpenoids in plants can be conceptually divided into three discrete processes: the formation of terpene precursors, the construction of terpene skeletons, followed by the complex post-modifications. After the skeleton construction by terpene synthases (TPS), diverse reactions catalyzed by the responsible post-modification enzymes result in tens of thousands of terpenoids. In this paper, with our preliminary studies on the biosynthesis of Isodon diterpenes, a number of representative post-modification enzymes of plant terpenoids formation reported in recent years are reviewed, which include P450 monooxidases, double-bond reductases, acyltransferases and glycosyltransferases.

Post-modification Enzymes Involved in the Biosynthesis of Plant Terpenoids

Post-modification Enzymes Involved in the Biosynthesis of Plant Terpenoids

Post-modification Enzymes Involved in the Biosynthesis of Plant Terpenoids

Post-modification Enzymes Involved in the Biosynthesis of Plant Terpenoids

Old substrates for new enzymes of terpenoid biosynthesis

Enzymologists have never had it so good. Thanks to advances in gene discovery, driven by high-throughput sequencing of genomes and transcriptomes, and improved heterologous expression of proteins, the functional characterization of new enzymes is generally straightforward: clone the gene, express the protein, assay for activity, and publish the results. However, without knowing the correct substrate to use in the assay such efforts can easily go astray. A paper by Schilmiller et al. (1) published in the latest issue of PNAS shows how one can elegantly avoid this pitfall by integrating genomic, genetic, enzymological, and metabolite-profiling approaches. In this important contribution, the authors report the discovery of a new substrate for enzymes of plant terpenoid biosynthesis.

The Correlation of Evolutionary Rate with Pathway Position in Plant Terpenoid Biosynthesis

Genes are expected to face stronger selective constraint and to evolve more slowly if they encode enzymes upstream as opposed to downstream in metabolic pathways, because upstream genes are more pleiotropic, being required for a wider range of end products. However, few clear examples of this trend in evolutionary rate variation exist. We examined whether genes involved in plant terpenoid biosynthesis exhibit such a pattern, using data for 40 genes from five fully sequenced angiosperms, Oryza, Vitis, Arabidopsis, Populus, and Ricinus. Our results show that d(N)/d(S) does in fact correlate with pathway position along pathways converting glucose to the terpenoid phytohormones abscissic acid, gibberellic acid (GA), and brassinosteroids. Upstream versus downstream rate variation is particularly strong in the GA pathway. In contrast, we found no or little apparent variation in d(N)/d(S) with gene copy number. We also introduce a new measure of pathway position, the Pathway Pleiotropy Index (PPI), which counts groups of enzymes between pathway branch points. We found that this measure is superior to pathway position in explaining variation in d(N)/d(S) along each pathway. Although at least 8 of the 40 genes showed evidence of positive selection, correlations of d(N)/d(S) with PPI remain significant when these genes are removed. Therefore, our results are consistent with the prediction that selective constraint is progressively relaxed along metabolic pathways.

Metabolic Engineering of Terpenoid Biosynthesis in Plants

Metabolic engineering of terpenoids in plants is a fascinating research topic from two main perspectives. On the one hand, the various biological activities of these compounds make their engineering a new tool for improving a considerable number of traits in crops. These include for example enhanced disease resistance, weed control by producing allelopathic compounds, better pest management, production of medicinal compounds, increased value of ornamentals and fruit and improved pollination. On the other hand, the same plants altered in the profile of terpenoids and their precursor pools make a most important contribution to fundamental studies on terpenoid biosynthesis and its regulation. In this review we describe the recent results with terpenoid engineering, showing that engineering of these compounds and their derivatives in plant cells is feasible, although with some requirements and limitations. For example, in terpenoid engineering experiments crucial factors are the subcellular localization of both the precursor pool and the introduced enzymes, the activity of endogenous plant enzymes which modify the introduced terpenoid skeleton, the costs of engineering in terms of effects on other pathways sharing the same precursor pool and the phytotoxicity of the introduced terpenoids. Finally, we will show that transgenic plants altered in their terpenoid profile exert novel biological activities on their environment, for example influencing insect behavior