An Update on the Function, Biosynthesis and Regulation of Floral Volatile Terpenoids

Floral volatile terpenoids (FVTs) belong to a group of volatile organic compounds (VOC) that play important roles in attracting pollinators, defending against pathogens and parasites and serving as signals associated with biotic and abiotic stress responses. Although research on FVTs has been increasing, a systematic generalization is lacking. Among flowering plants used mainly for ornamental purposes, a systematic study on the production of FVTs in flowers with characteristic aromas is still limited. This paper reviews the biological functions and biosynthesis of FVTs, which may contribute a foundational aspect for future research. We highlight regulatory mechanisms that control the production of FVTs in ornamental flowers and the intersection of biosynthetic pathways that produce flower fragrance and color. Additionally, we summarize the opportunities and challenges facing FVT research in the whole genome and -omics eras and the possible research directions that will provide a foundation for further innovation and utilization of flowering ornamental plants and their germplasm resources.

Differences in Floral Scent and Petal Reflectance Between Diploid and Tetraploid Chamerion angustifolium

Genome duplication in plants is thought to be a route to speciation due to cytotype incompatibility. However, to reduce cross-pollination between cytotypes in animal-pollinated species, distinctive floral phenotypes, which would allow pollinator-mediated assortative mating between flowers, are also expected. Chamerion angustifolium is a Holarctic species that forms a hybrid zone between diploid and tetraploid populations in the North American Rocky Mountains. Extensive research has shown that these cytotypes differ in many ways, including some floral traits, and that pollinators can discriminate between cytotypes, leading to assortative mating. However, two signals commonly used by insect pollinators have not been measured for this species, namely petal colour and floral scent. Using greenhouse-grown diploids and tetraploids of C. angustifolium from the ploidy hybrid-zone in the North American Rocky Mountains, we show that both floral scent signals and petal reflectance differ between cytotypes. These differences, along with differences in flower size shown previously, could help explain pollinator-mediated assortative mating observed in previous studies. However, these differences in floral phenotypes may vary in importance to pollinators. While the differences in scent included common floral volatiles readily detected by bumblebees, the differences in petal reflectance may not be perceived by bees based on their visual sensitivity across the spectra. Thus, our results suggest that differences in floral volatile emissions are more likely to contribute to pollinator discrimination between cytotypes and highlight the importance of understanding the sensory systems of pollinators when examining floral signals.

Functional Characterization of Hedychium coronarium J. Koenig MYB132 Confers the Potential Role in Floral Aroma Synthesis

The R2R3-MYB transcription factors (TFs) play several key roles in numerous plant biological processes. Hedychium coronarium is an important ornamental plant well-known for its elegant flower shape and abundant aroma type. The floral aroma of H. coronarium is due to the presence of a large amount of terpenes and benzenoids. However, less is known about the role of R2R3-MYB TFs in the regulatory mechanism of floral aroma production in this breed. Herein, we isolate and functionally characterize the R2R3-MYB TF HcMYB132, which is potentially involved in regulating floral aroma synthesis. Sequence alignment analysis revealed that it includes a nuclear localization signal NLS(s) and a 2R, 3R motif signature in the sequences. A subcellular localization assay revealed that HcMYB132 protein localizes to the nucleus. Real-time qPCR assays showed that HcMYB132 is specifically expressed in flowers and its expression pattern correlates with the emission of floral volatile compounds. In HcMYB132-silenced flowers, the levels of floral volatile compounds were significantly reduced, and the expression of key structural volatile synthesis genes was downregulated compared to control. Collectively, these results suggest that HcMYB132 might play a significant role in the regulation of terpenoid biosynthesis in H. coronarium.

Bumblebee electric charge stimulates floral volatile emissions in Petunia integrifolia but not in Antirrhinum majus

The timing of volatile organic compound (VOC) emission by flowering plants often coincides with pollinator foraging activity. Volatile emission is often considered to be paced by environmental variables, such as light intensity, and/or by circadian rhythmicity. The question arises as to what extent pollinators themselves provide information about their presence, in keeping with their long co-evolution with flowering plants. Bumblebees are electrically charged and provide electrical stimulation when visiting plants, as measured via the depolarisation of electric potential in the stem of flowers. Here we test the hypothesis that the electric charge of foraging bumblebees increases the floral volatile emissions of bee pollinated plants. We investigate the change in VOC emissions of two bee-pollinated plants (Petunia integrifolia and Antirrhinum majus) exposed to the electric charge typical of foraging bumblebees. P. integrifolia slightly increases its emissions of a behaviorally and physiologically active compound in response to visits by foraging bumblebees, presenting on average 121 pC of electric charge. We show that for P. integrifolia, strong electrical stimulation (600–700 pC) promotes increased volatile emissions, but this is not found when using weaker electrical charges more representative of flying pollinators (100 pC). Floral volatile emissions of A. majus were not affected by either strong (600–700 pC) or weak electric charges (100 pC). This study opens a new area of research whereby the electrical charge of flying insects may provide information to plants on the presence and phenology of their pollinators. As a form of electroreception, this sensory process would bear adaptive value, enabling plants to better ensure that their attractive chemical messages are released when a potential recipient is present.

HS–SPME–GC–MS and Electronic Nose Reveal Differences in the Volatile Profiles of Hedychium Flowers

Floral fragrance is one of the most important characteristics of ornamental plants and plays a pivotal role in plant lifespan such as pollinator attraction, pest repelling, and protection against abiotic and biotic stresses. However, the precise determination of floral fragrance is limited. In the present study, the floral volatile compounds of six Hedychium accessions exhibiting from faint to highly fragrant were comparatively analyzed via gas chromatography–mass spectrometry (GC–MS) and Electronic nose (E-nose). A total of 42 volatile compounds were identified through GC–MS analysis, including monoterpenoids (18 compounds), sesquiterpenoids (12), benzenoids/phenylpropanoids (8), fatty acid derivatives (2), and others (2). In Hedychium coronarium ‘ZS’, H. forrestii ‘Gaoling’, H. ‘Jin’, H. ‘Caixia’, and H. ‘Zhaoxia’, monoterpenoids were abundant, while sesquiterpenoids were found in large quantities in H. coccineum ‘KMH’. Hierarchical clustering analysis (HCA) divided the 42 volatile compounds into four different groups (I, II, III, IV), and Spearman correlation analysis showed these compounds to have different degrees of correlation. The E-nose was able to group the different accessions in the principal component analysis (PCA) corresponding to scent intensity. Furthermore, the pattern-recognition findings confirmed that the E-nose data validated the GC–MS results. The partial least squares (PLS) analysis between floral volatile compounds and sensors suggested that specific sensors were highly sensitive to terpenoids. In short, the E-nose is proficient in discriminating Hedychium accessions of different volatile profiles in both quantitative and qualitative aspects, offering an accurate and rapid reference technique for future applications.

Toxicity of Methyl Benzoate and Analogs to Adult Aedes aegypti

ABSTRACT Methyl benzoate is a natural product (floral volatile organic compound) that is currently used as a food flavoring ingredient. This compound has shown to be insecticidal in laboratory studies against agricultural and urban pests, including spotted wing drosophila Drosophila suzukii, brown marmorated stink bug Hyalomorpha halys, the diamondback moth Plutella xylostella, and the common bed bug Cimex lectularius, to name several insect taxa. In this study we topically treated adult Aedes aegypti females with methyl benzoate and analogs and determined their toxicities. We found that among adult females, 4 analogs—butyl benzoate, n-pentyl benzoate, vinyl benzoate, and methyl 3-methoxybenzoate—were more toxic than the parent compound, methyl benzoate.

A sensory bias overrides learned preferences of bumblebees for honest signals in Mimulus guttatus

Insect pollinators readily learn olfactory cues, and this is expected to select for ‘honest signals' that provide reliable information about floral rewards. However, plants might alternatively produce signals that exploit pollinators' sensory biases, thereby relaxing selection for signal honesty. We examined the innate and learned preferences of Bombus impatiens for Mimulus guttatus floral scent phenotypes corresponding to different levels of pollen rewards in the presence and absence of the innately attractive floral volatile compound β-trans-bergamotene. Bees learned to prefer honest signals after foraging on live M. guttatus flowers, but only exhibited this preference when presented floral scent phenotypes that did not include β-trans-bergamotene. Our results suggest that a sensory bias for β-trans-bergamotene overrides the ability of B. impatiens to use honest signals when foraging on M. guttatus . This may represent a deceptive pollination strategy that allows plants to minimize investment in costly rewards without incurring reduced rates of pollinator visitation.

Dynamic histone acetylation in floral volatile synthesis and emission in petunia flowers

Biosynthesis of secondary metabolites relies on primary metabolic pathways to provide precursors, energy, and cofactors, thus requiring coordinated regulation of primary and secondary metabolic networks. However, to date, it remains largely unknown how this coordination is achieved. Using Petunia hybrida flowers, which emit high levels of phenylpropanoid/benzenoid volatile organic compounds (VOCs), we uncovered genome-wide dynamic deposition of histone H3 lysine 9 acetylation (H3K9ac) during anthesis as an underlying mechanism to coordinate primary and secondary metabolic networks. The observed epigenome reprogramming is accompanied by transcriptional activation at gene loci involved in primary metabolic pathways that provide precursor phenylalanine, as well as secondary metabolic pathways to produce volatile compounds. We also observed transcriptional repression among genes involved in alternative phenylpropanoid branches that compete for metabolic precursors. We show that GNAT family histone acetyltransferase(s) (HATs) are required for the expression of genes involved in VOC biosynthesis and emission, by using chemical inhibitors of HATs, and by knocking down a specific HAT gene, ELP3, through transient RNAi. Together, our study supports that regulatory mechanisms at chromatin level may play an essential role in activating primary and secondary metabolic pathways to regulate VOC synthesis in petunia flowers.

Dynamic histone acetylation in floral volatile synthesis and emission in petunia flowers

ABSTRACTBiosynthesis of secondary metabolites relies on primary metabolic pathways to provide precursors, energy, and cofactors, thus requiring coordinated regulation of primary and secondary metabolic networks. However, to date it remains largely unknown how this coordination is achieved. Using Petunia hybrida flowers, which emit high levels of phenylpropanoid/benzenoid volatile organic compounds (VOCs), we uncovered genome-wide dynamic deposition of histone H3 lysine 9 acetylation (H3K9ac) during anthesis as an underlying mechanism to coordinate primary and secondary metabolic networks. The observed epigenome reprogramming is accompanied by transcriptional activation, at gene loci involved in primary metabolic pathways that provide precursor phenylalanine, as well as secondary metabolic pathways to produce volatile compounds. We also observed transcriptional repression among genes involved in alternative phenylpropanoid branches that compete for metabolic precursors. We show that GNAT family histone acetyltransferase(s) (HATs) are required for the expression of genes involved in VOC biosynthesis and emission, by using chemical inhibitors of HATs, and by knocking down a specific HAT, ELP3, through transient RNAi. Together, our study supports that chromatin level regulatory mechanisms may play an essential role in activating primary and secondary metabolic pathways to regulate VOC synthesis in petunia flowers.HIGHLIGHTOur study shows that posttranslational modification of histones is essential for regulating the biosynthesis and emission of floral scent compounds, thus providing insights into chromatin level regulation of secondary metabolism.

Flower Production, Headspace Volatiles, Pollen Nutrients, and Florivory in Tanacetum vulgare Chemotypes

Floral volatiles and reward traits are major drivers for the behavior of mutualistic as well as antagonistic flower visitors, i.e., pollinators and florivores. These floral traits differ tremendously between species, but intraspecific differences and their consequences on organism interactions remain largely unknown. Floral volatile compounds, such as terpenoids, function as cues to advertise rewards to pollinators, but should at the same time also repel florivores. The reward composition, e.g., protein and lipid contents in pollen, differs between individuals of distinct plant families. Whether the nutritional value of rewards within the same plant species is linked to their chemotypes, which differ in their pattern of specialized metabolites, has yet not been investigated. In the present study, we compared Tanacetum vulgare plants of five terpenoid chemotypes with regard to flower production, floral headspace volatiles, pollen macronutrient and terpenoid content, and floral attractiveness to florivorous beetles. Our analyses revealed remarkable differences between the chemotypes in the amount and diameter of flower heads, duration of bloom period, and pollen nutritional quality. The floral headspace composition of pollen-producing mature flowers, but not of premature flowers, was correlated to that of pollen and leaves in the same plant individual. For two chemotypes, florivorous beetles discriminated between the scent of mature and premature flower heads and preferred the latter. In semi-field experiments, the abundance of florivorous beetles and flower tissue miners differed between T. vulgare chemotypes. Moreover, the scent environment affected the choice and beetles were more abundant in homogenous plots composed of one single chemotype than in plots with different neighboring chemotypes. In conclusion, flower production, floral metabolic composition and pollen quality varied to a remarkable extend within the species T. vulgare, and the attractiveness of floral scent differed also intra-individually with floral ontogeny. We found evidence for a trade-off between pollen lipid content and pollen amount on a per-plant-level. Our study highlights that chemotypes which are more susceptible to florivory are less attacked when they grow in the neighborhood of other chemotypes and thus gain a benefit from high overall chemodiversity.