scholarly journals A novel terpene synthase controls differences in anti-aphrodisiac pheromone production between closely related Heliconius butterflies

PLoS Biology ◽  
2021 ◽  
Vol 19 (1) ◽  
pp. e3001022
Author(s):  
Kathy Darragh ◽  
Anna Orteu ◽  
Daniella Black ◽  
Kelsey J. R. P. Byers ◽  
Daiane Szczerbowski ◽  
...  
Keyword(s):  
Floral Scent ◽  
Terpene Synthase ◽  
Terpene Synthases ◽  
Common Component ◽  
Evolutionary Origins ◽  
Mapping Gene ◽  
Functional Analyses ◽  
Heliconius Cydno ◽  
Aphrodisiac Pheromone

Plants and insects often use the same compounds for chemical communication, but not much is known about the genetics of convergent evolution of chemical signals. The terpene (E)-β-ocimene is a common component of floral scent and is also used by the butterfly Heliconius melpomene as an anti-aphrodisiac pheromone. While the biosynthesis of terpenes has been described in plants and microorganisms, few terpene synthases (TPSs) have been identified in insects. Here, we study the recent divergence of 2 species, H. melpomene and Heliconius cydno, which differ in the presence of (E)-β-ocimene; combining linkage mapping, gene expression, and functional analyses, we identify 2 novel TPSs. Furthermore, we demonstrate that one, HmelOS, is able to synthesise (E)-β-ocimene in vitro. We find no evidence for TPS activity in HcydOS (HmelOS ortholog of H. cydno), suggesting that the loss of (E)-β-ocimene in this species is the result of coding, not regulatory, differences. The TPS enzymes we discovered are unrelated to previously described plant and insect TPSs, demonstrating that chemical convergence has independent evolutionary origins.

scholarly journals A novel terpene synthase produces an anti-aphrodisiac pheromone in the butterfly Heliconius melpomene

2019 ◽  
Author(s):  
Kathy Darragh ◽  
Anna Orteu ◽  
Kelsey J. R. P. Byers ◽  
Daiane Szczerbowski ◽  
Ian A. Warren ◽  
...  
Keyword(s):  
De Novo ◽  
Floral Scent ◽  
Ips Pini ◽  
Evolutionary Origin ◽  
Insect Species ◽  
Terpene Synthase ◽  
Terpene Synthases ◽  
Common Component ◽  
Heliconius Melpomene ◽  
Aphrodisiac Pheromone

AbstractTerpenes, a group of structurally diverse compounds, are the biggest class of secondary metabolites. While the biosynthesis of terpenes by enzymes known as terpene synthases (TPSs) has been described in plants and microorganisms, few TPSs have been identified in insects, despite the presence of terpenes in multiple insect species. Indeed, in many insect species, it remains unclear whether terpenes are sequestered from plants or biosynthesised de novo. No homologs of plant TPSs have been found in insect genomes, though insect TPSs with an independent evolutionary origin have been found in Hemiptera and Coleoptera. In the butterfly Heliconius melpomene, the monoterpene (E)-β-ocimene acts as an anti-aphrodisiac pheromone, where it is transferred during mating from males to females to avoid re-mating by deterring males. To date only one insect monoterpene synthase has been described, in Ips pini (Coleoptera), and is a multifunctional TPS and isoprenyl diphosphate synthase (IDS). Here, we combine linkage mapping and expression studies to identify candidate genes involved in the biosynthesis of (E)-β-ocimene. We confirm that H. melpomene has two enzymes that exhibit TPS activity, and one of these, HMEL037106g1 is able to synthesise (E)-β-ocimene in vitro. Unlike the enzyme in Ips pini, these enzymes only exhibit residual IDS activity, suggesting they are more specialised TPSs, akin to those found in plants. Phylogenetic analysis shows that these enzymes are unrelated to previously described plant and insect TPSs. The distinct evolutionary origin of TPSs in Lepidoptera suggests that they have evolved multiple times in insects.Significance statementTerpenes are a diverse class of natural compounds, used by both plants and animals for a variety of functions, including chemical communication. In insects it is often unclear whether they are synthesised de novo or sequestered from plants. Some plants and insects have converged to use the same compounds. For instance, (E)-β-ocimene is a common component of floral scent and is also used by the butterfly Heliconius melpomene as an anti-aphrodisiac pheromone. We describe two novel terpene synthases, one of which synthesises (E)-β-ocimene in H. melpomene, unrelated not only to plant enzymes but also other recently identified insect terpene synthases. This provides the first evidence that the ability to synthesise terpenes has arisen multiple times independently within the insects.

scholarly journals Differential gene expression associated with a floral scent polymorphism in the evening primrose Oenothera harringtonii (Onagraceae)

2021 ◽  
Author(s):  
Lindsey L. Bechen ◽  
Matthew G. Johnson ◽  
Geoffrey T. Broadhead ◽  
Rachel A. Levin ◽  
Rick P. Overson ◽  
...  
Keyword(s):  
Gene Expression ◽  
Differential Gene Expression ◽  
Floral Scent ◽  
Terpene Synthase ◽  
Rna Seq ◽  
Loss Of Function ◽  
Terpene Biosynthesis ◽  
Terpene Synthases ◽  
Evening Primrose ◽  
Differential Gene

AbstractBackgroundPlant volatiles play an important role in both plant-pollinator and plant-herbivore interactions. Intraspecific polymorphisms in volatile production are ubiquitous, but studies that explore underlying differential gene expression are rare. Oenothera harringtonii populations are polymorphic in floral emission of the monoterpene (R)-(-)-linalool; some plants emit (R)-(-)-linalool (linalool+ plants) while others do not (linalool-plants). However, the genes associated with differential production of this floral volatile in Oenothera are unknown. We used RNA-Seq to broadly characterize differential gene expression involved in (R)-(-)-linalool biosynthesis. To identify genes that may be associated with the polymorphism for this trait, we used RNA-Seq to compare gene expression in six different Oenothera harringtonii tissues from each of three linalool+ and linalool-plants.ResultsThree clusters of differentially expressed genes were enriched for terpene synthase activity: two were characterized by tissue-specific upregulation and one by upregulation only in plants with flowers that produce (R)-(-)-linalool. A molecular phylogeny of all terpene synthases identified two putative (R)-(-)-linalool synthase transcripts in Oenothera harringtonii, a single allele of which is found exclusively in linalool+ plants.ConclusionsBy using a naturally occurring polymorphism and comparing different tissues, we were able to identify genes putatively involved in the biosynthesis of (R)-(-)-linalool. Expression of these genes in linalool-plants suggests a regulatory polymorphism, rather than a population-specific loss-of-function allele. Additional terpene biosynthesis-related genes that are up-regulated in plants that emit (R)-(-)-linalool may be associated with herbivore defense, suggesting a potential economy of scale between plant reproduction and defense.

scholarly journals Composition and Biosynthesis of Scent Compounds from Sterile Flowers of an Ornamental Plant Clematis florida cv. ‘Kaiser’

Molecules ◽  
2020 ◽  
Vol 25 (7) ◽  
pp. 1711
Author(s):  
Yifan Jiang ◽  
Renjuan Qian ◽  
Wanbo Zhang ◽  
Guo Wei ◽  
Xiaohua Ma ◽  
...  
Keyword(s):  
Floral Scent ◽  
Ornamental Plant ◽  
Terpene Synthase ◽  
Enzyme Assays ◽  
Total Volatile ◽  
Terpene Biosynthesis ◽  
Geranyl Diphosphate ◽  
Multiple Products ◽  
Key Genes

Clematis florida is a popular ornamental vine species known for diverse colors and shapes of its flowers but not for scent. Here we investigated the composition and biosynthesis of floral scent in ‘Kaiser’, a fragrant cultivar of C. florida that has sterile flowers. Volatile profiling revealed that flowers of ‘Kaiser’ emit more than 20 compounds, with monoterpenes being most abundant. Among the three floral organs, namely sepals, transformed-petals, and ovaries, ovaries had the highest rates of total volatile emission. To determine the molecular mechanism underlying floral scent biosynthesis in ‘Kaiser’, we sequenced a flower transcriptome and searched the transcriptome for terpene synthase genes (TPSs), which are key genes for terpene biosynthesis. Among the TPS genes identified, three were putative intact full-length genes and were designated CfTPS1, CfTPS2, and CfTPS3. Phylogenetic analysis placed CfTPS1, CfTPS2, and CfTPS3 to the TPS-g, TPS-b, and TPS-a subfamily, respectively. Through in vitro enzyme assays with Escherichia coli-expressed recombinant proteins, both CfTPS1 and CfTPS2 were demonstrated to catalyze the conversion of geranyl diphosphate to linalool, the most abundant constituent of C. florida floral scent. In addition, CfTPS1 and CfTPS2 produced the sesquiterpene nerolidol from (E,E)-farnesyl diphosphate. CfTPS3 showed sesquiterpene synthase activity and produced multiple products in vitro. All three CfTPS genes showed higher levels of expression in sepals than those in transformed-petals and ovaries. Our results show that despite being sterile, the flowers of ‘Kaiser’ have normal mechanisms for floral scent biosynthesis that make the flowers fragrant.

scholarly journals Diversity and function of terpene synthases in the production of carrot aroma and flavor compounds

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Andrew Muchlinski ◽  
Mwafaq Ibdah ◽  
Shelby Ellison ◽  
Mossab Yahyaa ◽  
Bhagwat Nawade ◽  
...  
Keyword(s):  
Expression Profiles ◽  
Terpene Synthase ◽  
Terpene Synthases ◽  
Specific Expression ◽  
Metabolite Profiles ◽  
Germacrene D ◽  
Volatile Terpenes ◽  
And Function ◽  
Volatile Terpene

Abstract Carrot (Daucus carota L.) is an important root vegetable crop with high nutritional value, characteristic flavor, and benefits to human health. D. carota tissues produce an essential oil that is rich in volatile terpenes and plays a major role in carrot aroma and flavor. Although terpene composition represents a critical quality attribute of carrots, little is known about the biosynthesis of terpenes in this crop. Here, we functionally characterized 19 terpene synthase (TPS) genes in an orange carrot (genotype DH1) and compared tissue-specific expression profiles and in vitro products of their recombinant proteins with volatile terpene profiles from DH1 and four other colored carrot genotypes. In addition to the previously reported (E)-β-caryophyllene synthase (DcTPS01), we biochemically characterized several TPS proteins with direct correlations to major compounds of carrot flavor and aroma including germacrene D (DcTPS7/11), γ-terpinene (DcTPS30) and α-terpinolene (DcTPS03). Random forest analysis of volatiles from colored carrot cultivars identified nine terpenes that were clearly distinct among the cultivars and likely contribute to differences in sensory quality. Correlation of TPS gene expression and terpene metabolite profiles supported the function of DcTPS01 and DcTPS03 in these cultivars. Our findings provide a roadmap for future breeding efforts to enhance carrot flavor and aroma.

scholarly journals Terpene Synthase-b and Terpene Synthase-e/f Genes Produce Monoterpenes for Phalaenopsis bellina Floral Scent

2021 ◽  
Vol 12 ◽  
Author(s):  
Hsin Huang ◽  
Yi-Wei Kuo ◽  
Yu-Chen Chuang ◽  
Ya-Ping Yang ◽  
Li-Min Huang ◽  
...  
Keyword(s):  
Floral Scent ◽  
Functional Characterization ◽  
Ectopic Expression ◽  
Reproductive Organs ◽  
Olfactory Cues ◽  
Terpene Synthase ◽  
In Planta ◽  
Monoterpene Biosynthesis ◽  
Differential Gene

Orchids are the most species-rich plants and highly interactive with pollinators via visual or olfactory cues. Biosynthesis and emission of volatile organic compounds (VOCs) to the atmosphere facilitate the olfactory cues and ensure successful pollination. Phalaenopsis bellina is a scented orchid with monoterpenes as major VOCs, comprising linalool, geraniol, and their derivatives. Comparative transcriptomics analysis identified four terpene synthase-b (TPS-b) genes and two TPS-e/f genes with differential gene expression between scented and scentless Phalaenopsis species. Here, we confirmed their differential expression between scented and scentless Phalaenopsis orchids and excluded one TPS-b candidate. We analyzed the temporal and spatial expression and functionally characterized these TPSs. Both TPS-b and TPS-e/f genes showed an increased expression on blooming day or 3 days post-anthesis (D + 3) before the optimal emission of floral scent on D + 5, with especially high expression of PbTPS5 and PbTPS10. The TPS-b genes are expressed exclusively in reproductive organs, whereas the TPS-e/f genes are expressed in both reproductive and vegetative organs. In planta functional characterization of both PbTPS5 and PbTPS10 in tobacco and scentless Phalaenopsis plants did not produce terpenoids. Further ectopic expression in scented Phalaenopsis cultivar P. I-Hsin Venus showed that linalool was the main product, with PbTPS10 displaying 3-fold higher activity than PbTPS5. On in vitro enzyme assay with purified recombinant TPS-b proteins ectopically expressed in Escherichia coli, geraniol was the product catalyzed by PbTPS5 and PbTPS9. PbTPS3 was a linalool/(β)-cis-ocimene synthase and PbTPS4 a linalool synthase. In conclusion, both TPS-b and TPS-e/f enzymes orchestrated floral monoterpene biosynthesis in P. bellina.

scholarly journals Genome-Wide Identification and Expression Analysis of Terpene Synthase Genes in Cymbidium faberi

2021 ◽  
Vol 12 ◽  
Author(s):  
Qian-Qian Wang ◽  
Meng-Jia Zhu ◽  
Xia Yu ◽  
Yuan-Yang Bi ◽  
Zhuang Zhou ◽  
...  
Keyword(s):  
Expression Patterns ◽  
Terpene Synthase ◽  
Terpene Synthases ◽  
Specific Expression ◽  
Functional Roles ◽  
Genome Wide ◽  
Reverse Transcription Quantitative Pcr ◽  
Full Flowering ◽  
Qpcr Experiment

Terpene synthases (TPSs) are essential for forming terpenes, which play numerous functional roles in attracting pollinators, defending plants, and moderating the interaction between plants. TPSs have been reported in some orchids, but genome-wide identification of terpenes in Cymbidium faberi is still lacking. In this study, 32 putative TPS genes were classified in C. faberi and divided into three subfamilies (TPS-a, TPS-b, and TPS-e/f). Motif and gene structure analysis revealed that most CfTPS genes had the conserved aspartate-rich DDxxD motif. TPS genes in the TPS-a and TPS-b subfamilies had variations in the RRX8W motif. Most cis-elements of CfTPS genes were found in the phytohormone responsiveness category, and MYC contained most of the numbers associated with MeJA responsiveness. The Ka/Ks ratios of 12/13 CfTPS gene pairs were less than one, indicated that most CfTPS genes have undergone negative selection. The tissue-specific expression patterns showed that 28 genes were expressed in at least one tissue in C. faberi, and TPS genes were most highly expressed in flowers, followed by leaves and pseudobulbs. In addition, four CfTPS genes were selected for the real-time reverse transcription quantitative PCR (RT-qPCR) experiment. The results revealed that CfTPS12, CfTPS18, CfTPS23, and CfTPS28 were mainly expressed in the full flowering stage. CfTPS18 could convert GPP to β-myrcene, geraniol, and α-pinene in vitro. These findings of CfTPS genes of C. faberi may provide valuable information for further studies on TPSs in orchids.

scholarly journals Transcriptomic Insight into Terpenoid Biosynthesis and Functional Characterization of Three Diterpene Synthases in Scutellaria barbata

Molecules ◽  
2018 ◽  
Vol 23 (11) ◽  
pp. 2952 ◽  
Author(s):  
Huabei Zhang ◽  
Baolong Jin ◽  
Junling Bu ◽  
Juan Guo ◽  
Tong Chen ◽  
...  
Keyword(s):  
Average Length ◽  
Functional Characterization ◽  
Terpene Synthase ◽  
Terpenoid Biosynthesis ◽  
Vitro Assay ◽  
Scutellaria Barbata ◽  
Aerial Parts ◽  
Diterpene Synthase ◽  
Clerodane Diterpenoids

Scutellaria barbata (Lamiaceae) is an important medicinal herb widely used in China, Korea, India, and other Asian countries. Neo-clerodane diterpenoids are the largest known group of Scutellaria diterpenoids and show promising cytotoxic activity against several cancer cell lines. Here, Illumina-based deep transcriptome analysis of flowers, the aerial parts (leaf and stem), and roots of S. barbata was used to explore terpenoid-related genes. In total, 121,958,564 clean RNA-sequence reads were assembled into 88,980 transcripts, with an average length of 1370 nt and N50 length of 2144 nt, indicating high assembly quality. We identified nearly all known terpenoid-related genes (33 genes) involved in biosynthesis of the terpenoid backbone and 14 terpene synthase genes which generate skeletons for different terpenoids. Three full length diterpene synthase genes were functionally identified using an in vitro assay. SbTPS8 and SbTPS9 were identified as normal-CPP and ent-CPP synthase, respectively. SbTPS12 reacts with SbTPS8 to produce miltiradiene. Furthermore, SbTPS12 was proven to be a less promiscuous class I diterpene synthase. These results give a comprehensive understanding of the terpenoid biosynthesis in S. barbata and provide useful information for enhancing the production of bioactive neo-clerodane diterpenoids through genetic engineering.

scholarly journals Transcriptome and proteome of the corm, leaf and flower of Hypoxis hemerocallidea (African potato)

PLoS ONE ◽  
2021 ◽  
Vol 16 (7) ◽  
pp. e0253741
Author(s):  
Mihai-Silviu Tomescu ◽  
Selisha Ann Sooklal ◽  
Thuto Ntsowe ◽  
Previn Naicker ◽  
Barbara Darnhofer ◽  
...  
Keyword(s):  
De Novo ◽  
Differential Expression Analysis ◽  
Terpene Synthase ◽  
Future Research ◽  
Terpene Synthases ◽  
Illumina Hiseq ◽  
Prostate Hyperplasia ◽  
Inflammatory Conditions ◽  
Urinary Infections ◽  
Hypoxis Hemerocallidea

The corm of Hypoxis hemerocallidea, commonly known as the African potato, is used in traditional medicine to treat several medical conditions such as urinary infections, benign prostate hyperplasia, inflammatory conditions and testicular tumours. The metabolites contributing to the medicinal properties of H. hemerocallidea have been identified in several studies and, more recently, the active terpenoids of the plant were profiled. However, the biosynthetic pathways and the enzymes involved in the production of the terpene metabolites in H. hemerocallidea have not been characterised at a transcriptomic or proteomic level. In this study, total RNA extracted from the corm, leaf and flower tissues of H. hemerocallidea was sequenced on the Illumina HiSeq 2500 platform. A total of 143,549 transcripts were assembled de novo using Trinity and 107,131 transcripts were functionally annotated using the nr, GO, COG, KEGG and SWISS-PROT databases. Additionally, the proteome of the three tissues were sequenced using LC-MS/MS, revealing aspects of secondary metabolism and serving as data validation for the transcriptome. Functional annotation led to the identification of numerous terpene synthases such as nerolidol synthase, germacrene D synthase, and cycloartenol synthase amongst others. Annotations also revealed a transcript encoding the terpene synthase phytoalexin momilactone A synthase. Differential expression analysis using edgeR identified 946 transcripts differentially expressed between the three tissues and revealed that the leaf upregulates linalool synthase compared to the corm and the flower tissues. The transcriptome as well as the proteome of Hypoxis hemerocallidea presented here provide a foundation for future research.

scholarly journals Functional analyses of phosphatidylserine/PI(4)P exchangers with diverse lipid species and membrane contexts set unanticipated rules on lipid transfer

2021 ◽  
Author(s):  
Souade Ikhlef ◽  
Nicolas-Frédéric Lipp ◽  
Vanessa Delfosse ◽  
Nicolas Fuggetta ◽  
William Bourguet ◽  
...  
Keyword(s):  
Lipid Transfer ◽  
Oxysterol Binding Protein ◽  
Lipid Exchange ◽  
Exchange Processes ◽  
Lipid Species ◽  
Acyl Chains ◽  
Phosphatidylinositol 4 ◽  
Functional Analyses ◽  
Related Proteins

Several members of the oxysterol-binding protein-related proteins (ORPs)/oxysterol-binding homology (Osh) family exchange phosphatidylserine (PS) and phosphatidylinositol 4-phosphate (PI(4)P) at the endoplasmic reticulum/plasma membrane (PM) interface. It is unclear whether they preferentially exchange PS and PI(4)P with specific acyl chains to tune the features of the PM, whether they use phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2) instead of PI(4)P for exchange processes and whether their activity is influenced by the association of PS with sterol in the PM. Here, we measured in vitro how the yeast Osh6p and human ORP8 transported diverse PS and PI(4)P subspecies, including major cellular species, between membranes. We established how their activity is impacted by the length and unsaturation degree of these lipids. Surprisingly, the speed at which they individually transfer these ligands inversely depends on their affinity for them. To be fast, the transfer of high-affinity ligands requires an exchange with a counterligand of equivalent affinity. Besides, we determined that Osh6p and ORP8 cannot use PI(4,5)P2 for intracellular lipid exchange, as they have a low affinity for this lipid compared to PI(4)P, and do not transfer more PS into sterol-rich membranes. This study provides insights into PS/PI(4)P exchangers and sets unanticipated rules on how the activity of lipid transfer proteins relates to their affinity for ligands.

scholarly journals Identification of Sesquiterpene Synthases from Nostoc punctiforme PCC 73102 and Nostoc sp. Strain PCC 7120

2008 ◽  
Vol 190 (18) ◽  
pp. 6084-6096 ◽  
Author(s):  
Sean A. Agger ◽  
Fernando Lopez-Gallego ◽  
Thomas R. Hoye ◽  
Claudia Schmidt-Dannert
Keyword(s):  
Functional Expression ◽  
Terpene Synthase ◽  
Putative Hybrid ◽  
Nostoc Punctiforme ◽  
Terpene Synthases ◽  
E Coli ◽  
Reductase Gene ◽  
Pcc 7120 ◽  
Defense Compound

ABSTRACT Cyanobacteria are a rich source of natural products and are known to produce terpenoids. These bacteria are the major source of the musty-smelling terpenes geosmin and 2-methylisoborneol, which are found in many natural water supplies; however, no terpene synthases have been characterized from these organisms to date. Here, we describe the characterization of three sesquiterpene synthases identified in Nostoc sp. strain PCC 7120 (terpene synthase NS1) and Nostoc punctiforme PCC 73102 (terpene synthases NP1 and NP2). The second terpene synthase in N. punctiforme (NP2) is homologous to fusion-type sesquiterpene synthases from Streptomyces spp. shown to produce geosmin via an intermediate germacradienol. The enzymes were functionally expressed in Escherichia coli, and their terpene products were structurally identified as germacrene A (from NS1), the eudesmadiene 8a-epi-α-selinene (from NP1), and germacradienol (from NP2). The product of NP1, 8a-epi-α-selinene, so far has been isolated only from termites, in which it functions as a defense compound. Terpene synthases NP1 and NS1 are part of an apparent minicluster that includes a P450 and a putative hybrid two-component protein located downstream of the terpene synthases. Coexpression of P450 genes with their adjacent located terpene synthase genes in E. coli demonstrates that the P450 from Nostoc sp. can be functionally expressed in E. coli when coexpressed with a ferredoxin gene and a ferredoxin reductase gene from Nostoc and that the enzyme oxygenates the NS1 terpene product germacrene A. This represents to the best of our knowledge the first example of functional expression of a cyanobacterial P450 in E. coli.

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