Lauren A. E. Erland

The biosynthetic origin of irregular monoterpenes in Lavandula: isolation and biochemical characterization of a novel cis prenyl diphosphate synthase gene - lavandulyl diphosphate synthase

Background: Lavandula accumulate irregular monoterpenes of unknown biosynthetic origin. Results: We cloned a cis-prenyl diphosphate synthase (cis-PDPS) that produces precursor for irregular monoterpenes in lavenders. Conclusion: Unlike other plants that utilize trans-PDPSs, Lavandula employ a cis-PDPS to initiate the biosynthesis of irregular monoterpenes. Significance: This is the first report of the involvement of a cis-PDPS in irregular monoterpene biosynthesis. Lavender essential oils are constituted predominantly of regular monoterpenes, for example linalool, 1,8-cineole, and camphor. However, they also contain irregular monoterpenes including lavandulol and lavandulyl acetate. Although the majority of genes responsible for the production of regular monoterpenes in lavenders are now known, enzymes (including lavandulyl diphosphate synthase (LPPS)) catalyzing the biosynthesis of irregular monoterpenes in these plants have not been described. Here, we report the isolation and functional characterization of a novel cis-prenyl diphosphate synthase cDNA, termed Lavandula x intermedia lavandulyl diphosphate synthase (LiLPPS), through a homology-based cloning strategy. The LiLPPS ORF, encoding for a 305-amino acid long protein, was expressed in Escherichia coli, and the recombinant protein was purified by nickel-nitrilotriacetic acid affinity chromatography. The approximately 34.5-kDa bacterially produced protein specifically catalyzed the head-to-middle condensation of two dimethylallyl diphosphate units to LPP in vitro with apparent Km and kcat values of 208 ± 12 μm and 0.1 s−1, respectively. LiLPPS is a homodimeric enzyme with a sigmoidal saturation curve and Hill coefficient of 2.7, suggesting a positive co-operative interaction among its catalytic sites. LiLPPS could be used to modulate the production of lavandulol and its derivatives in plants through metabolic engineering.

A new balancing act: The many roles of melatonin and serotonin in plant growth and development

Melatonin and serotonin are indoleamines first identified as neurotransmitters in vertebrates; they have now been found to be ubiquitously present across all forms of life. Both melatonin and serotonin were discovered in plants several years after their discovery in mammals, but their presence has now been confirmed in almost all plant families. The mechanisms of action of melatonin and serotonin are still poorly defined. Melatonin and serotonin possess important roles in plant growth and development, including functions in chronoregulation and modulation of reproductive development, control of root and shoot organogenesis, maintenance of plant tissues, delay of senescence, and responses to biotic and abiotic stresses. This review focuses on the roles of melatonin and serotonin as a novel class of plant growth regulators. Their roles in reproductive and vegetative plant growth will be examined including an overview of current hypotheses and knowledge regarding their mechanisms of action in specific responses.

Serotonin: An ancient molecule and an important regulator of plant processes

Serotonin is an ancient indoleamine that was presumably part of the life cycle of the first prokaryotic life forms on Earth millions of years ago where it functioned as a powerful antioxidant to combat the increasingly oxygen rich atmosphere. First identified as a neurotransmitter signaling molecule in mammals, it is ubiquitous across all forms of life. Serotonin was discovered in plants many years after its discovery in mammals; however, it has now been confirmed in almost all plant families, where it plays important roles in plant growth and development, including functions in energy acquisition, seasonal cycles, modulation of reproductive development, control of root and shoot organogenesis, maintenance of plant tissues, delay of senescence, and responses to biotic and abiotic stresses. Despite its widespread presence and activity, there are many questions which remain unanswered about the role of serotonin in plants including the mode of signaling and receptor identity as well as the mechanisms of action of this important molecule. This review provides an overview of the role of serotonin in plant life and their ability to adapt.

Melatonin and serotonin: Mediators in the symphony of plant morphogenesis

Melatonin and serotonin are important signaling and stress mitigating molecules that play important roles across growth and development in plants. Despite many well‐documented responses, a systematic investigation of the entire metabolic pathway (tryptophan, tryptamine, and N‐acetylserotonin) does not exist, leaving many open questions. The objective of this study was to determine the responses of Hypericum perforatum (L.) to melatonin, serotonin, and their metabolic precursors. Two well‐characterized germplasm lines (#4 and 112) created by mutation and a haploid breeding program were compared to wild type to identify specific responses. Germplasm line 4 has lower regenerative and photosynthetic capacity than either wild type or line 112, and there are documented significant differences in the chemistry and physiology of lines 4 and 112. Supplementation of the culture media with tryptophan, tryptamine, N‐acetylserotonin, serotonin, or melatonin partially reversed the regenerative recalcitrance and growth impairment of the germplasm lines. Quantification of phytohormones revealed crosstalk between the indoleamines and related phytohormones including cytokinin, salicylic acid, and abscisic acid. We hypothesize that melatonin and serotonin function in coordination with their metabolites in a cascade of phytochemical responses including multiple pathways and phytohormone networks to direct morphogenesis and protect photosynthesis in H. perforatum.

Melatonin Natural Health Products and Supplements: Presence of Serotonin and Significant Variability of Melatonin Content

STUDY OBJECTIVES Melatonin is an important neurohormone, which mediates circadian rhythms and the sleep cycle. As such, it is a popular and readily available supplement for the treatment and prevention of sleep-related disorders including insomnia and jet lag. This study quantified melatonin in 30 commercial supplements, comprising different brands and forms and screened supplements for the presence of serotonin. METHODS A total of 31 supplements were analyzed by ultraperformance liquid chromatography with electrochemical detection for quantification of melatonin and serotonin. Presence of serotonin was confirmed through analysis by ultraperformance liquid chromatography with mass spectrometry detection. RESULTS Melatonin content was found to range from -83% to +478% of the labelled content. Additionally, lot-to-lot variable within a particular product varied by as much as 465%. This variability did not appear to be correlated with manufacturer or product type. Furthermore, serotonin (5-hydroxytryptamine), a related indoleamine and controlled substance used in the treatment of several neurological disorders, was identified in eight of the supplements at levels of 1 to 75 μg. CONCLUSIONS Melatonin content did not meet label within a 10% margin of the label claim in more than 71% of supplements and an additional 26% were found to contain serotonin. It is important that clinicians and patients have confidence in the quality of supplements used in the treatment of sleep disorders. To address this, manufacturers require increased controls to ensure melatonin supplements meet both their label claim, and also are free from contaminants, such as serotonin. COMMENTARY A commentary on this article appears in this issue on page 163.

Melatonin in Plants and Plant Culture Systems: Variability, Stability and Efficient Quantification

Despite growing evidence of the importance of melatonin and serotonin in the plant life, there is still much debate over the stability of melatonin, with extraction and analysis methods varying greatly from lab to lab with respect to time, temperature, light levels, extraction solvents, and mechanical disruption. The variability in methodology has created conflicting results that confound the comparison of studies to determine the role of melatonin in plant physiology. We here describe a fully validated method for the quantification of melatonin, serotonin and their biosynthetic precursors: tryptophan, tryptamine and N-acetylserotonin by liquid chromatography single quadrupole mass spectrometry (LC-MS) in diverse plant species and tissues. This method can be performed on a simple and inexpensive platform, and is both rapid and simple to implement. The method has excellent reproducibility and acceptable sensitivity with percent relative standard deviation (%RSD) in all matrices between 1 and 10% and recovery values of 82–113% for all analytes. Instrument detection limits were 24.4 ng/mL, 6.10 ng/mL, 1.52 ng/mL, 6.10 ng/mL, and 95.3 pg/mL, for serotonin, tryptophan, tryptamine, N-acetylserotonin and melatonin respectively. Method detection limits were 1.62 μg/g, 0.407 μg/g, 0.101 μg/g, 0.407 μg/g, and 6.17 ng/g respectively. The optimized method was then utilized to examine the issue of variable stability of melatonin in plant tissue culture systems. Media composition (Murashige and Skoog, Driver and Kuniyuki walnut or Lloyd and McCowns woody plant medium) and light (16 h photoperiod or dark) were found to have no effect on melatonin or serotonin content. A Youden trial suggested temperature as a major factor leading to degradation of melatonin. Both melatonin and serotonin appeared to be stable across the first 10 days in media, melatonin losses reached a mean minimum degradation at 28 days of approximately 90%; serotonin reached a mean minimum value of approximately 60% at 28 days. These results suggest that melatonin and serotonin show considerable stability in plant systems and these indoleamines and related compounds can be used for investigations that span over 3 weeks.

Melatonin in plant signalling and behaviour

Melatonin is an indoleamine neurotransmitter that has recently become well established as an important multi-functional signalling molecule in plants. These signals have been found to induce several important physiological responses that may be interpreted as behaviours. The diverse processes in which melatonin has been implicated in plants have expanded far beyond the traditional roles for which it has been implicated in mammals, which include sleep, tropisms and reproduction. These functions, however, appear to also be important melatonin mediated processes in plants, though the mechanisms underlying these functions have yet to be fully elucidated. Mediation or redirection of plant physiological processes induced by melatonin can be summarised as a series of behaviours including, among others: herbivore defence, avoidance of undesirable circumstances or attraction to opportune conditions, problem solving and response to environmental stimulus. As the mechanisms of melatonin action are elucidated, its involvement in plant growth, development and behaviour is likely to expand beyond the aspects discussed in this review and hold promise for applications in diverse fundamental and applied plant sciences including conservation, cryopreservation, morphogenesis, industrial agriculture and natural health products.

Chapter 57 – Lavender (Lavandula angustifolia) Oils

Lavenders are popular aromatic plants in the Lamiaceae family native to the Mediterranean region. Aqueous extracts, essential oils, and dried parts of these plants have long been used in cosmetics, hygiene products, and traditional medicines. In addition, they are used as food additives due to their pleasant flavor and aroma, and their antibacterial, antifungal, insect repellent, insecticidal, and antioxidant properties. With increasing concerns about loss of effectiveness and safety of commercial antibiotics, antifungals, insecticides, and preservatives, botanical products, including lavenders, are coming to the forefront. In this review, the current and potential applications of lavender essential oil in the food sciences are discussed.

Melatonin in plant morphogenesis

Melatonin is a ubiquitously present indoleamine with a vast capacity for modulating the growth and behavior of plants, animals, and microbes. Though melatonin was discovered in plants decades after its discovery in mammals, its presence has now been confirmed in almost all plant families. Despite this, the in vitro and in vivo mechanisms of action of melatonin are still poorly defined. Although there are an increasingly large number of investigations into the roles of melatonin in plants, few take advantage of in vitro culture systems. Melatonin has been found to possess several important roles in plant growth and development, including functions in rhythmic and cyclic processes, such as chronoregulation, seasonal and senescence processes, as well as modulation of reproductive development, control of root and shoot organogenesis, maintenance of plant tissues, and responses to biotic and abiotic stresses. This review highlights the potential for use of melatonin in several in vitro systems, the roles it plays in plant morphogenesis, and the importance of melatonin in communication within and between plants, and how in vitro systems can be exploited to better understand these understudied functions of melatonin.

Bark and wood tissues of American elm exhibit distinct responses to Dutch elm disease

Tolerance to Dutch elm disease (DED) has been linked to the rapid and/or high induction of disease-responsive genes after infection with the fungus Ophiostoma novo-ulmi. Although the fungal infection by O. novo-ulmi primarily takes places in xylem vessels, it is still unclear how xylem contributes to the defense against DED. Taking advantage of the easy separation of wood and bark tissues in young American elm saplings, here we show that most disease-responsive genes exhibited higher expression in wood compared to bark tissues after fungal infection. On the other hand, the stress-related phytohormones were generally more abundant in the bark compared to wood tissues. However, only endogenous levels of jasmonates (JAs), but not salicylic acid (SA) and abscisic acid (ABA) increased in the inoculated tissues. This, along with the upregulation of JA-biosynthesis genes in inoculated bark and core tissues further suggest that phloem and xylem might contribute to the de novo biosynthesis of JA after fungal infection. The comparison between two tolerant elm varieties, ‘Valley Forge’ and ‘Princeton,’ also indicated that tolerance against DED might be mediated by different mechanisms in the xylem. The present study sheds some light on the amplitude and kinetics of defense responses produced in the xylem and phloem in response to DED.