A chromosome level reference genome of Diviner's sage (Salvia divinorum) provides insight into salvinorin A biosynthesis.

Background: Diviner's sage (Salvia divinorum; Lamiaceae) is the source of the powerful hallucinogen salvinorin A (SalA). This neoclerodane diterpenoid is an agonist of the human Κ-opioid receptor with potential medical applications in the treatment of chronic pain, addiction, and post-traumatic stress disorder. Only two steps of the approximately twelve step biosynthetic sequence leading to SalA have been resolved to date.

A novel copy number variant in the murine Cdh23 gene gives rise to profound deafness and vestibular dysfunction.

Hearing loss is the most common congenital sensory deficit worldwide and exhibits high genetic heterogeneity, making molecular diagnoses elusive for most individuals. Detecting novel mutations that contribute to hearing loss is crucial to providing accurate personalized diagnoses, tailored interventions, and improving prognosis. Copy number variants (CNVs) are structural mutations that are understudied, potential contributors to hearing loss.

Plastid ancestors lacked a complete Entner-Doudoroff pathway, limiting plants to glycolysis and the pentose phosphate pathway.

The Entner-Doudoroff (ED) pathway provides an alternative to glycolysis. It converts 6-phosphogluconate (6-PG) to glyceraldehyde-3-phosphate and pyruvate in two steps consisting of a dehydratase (EDD) and an aldolase (EDA). Here, we investigate its distribution and significance in higher plants and determine the ED pathway is restricted to prokaryotes due to the absence of EDD genes in eukaryotes.

Arabidopsis TGA256 Transcription Factors Suppress Salicylic-Acid-Induced Sucrose Starvation.

Salicylic acid (SA) is produced by plants in response to pathogen infection. SA binds the NONEXPRESSOR OF PATHOGENESIS-RELATED GENES (NPR) family of receptors to regulate both positive (NPR1) and negative (NPR3/4) plant immune responses by interacting with the clade II TGACG (TGA) motif-binding transcription factors (TGA2, TGA5, and TGA6). Here, we report that the principal metabolome-level response to SA treatment in Arabidopsis is a reduction in sucrose and other free sugars.

Improving the Documentation of Penicillin Allergy Labels Among Pediatric Inpatients.

Background And Objectives: Penicillin allergy is the most common medication allergy, and the penicillin allergy label is commonly over-applied without adequate reaction history inquiry or documentation. Because penicillin allergy labels are often applied in childhood and carried into adulthood, we sought to increase the completeness of reaction history documentation from 20% to 70% for pediatric hospital medicine patients and from 20% to 50% for all other pediatric inpatients within 12 months. As a secondary outcome, we also aimed to increase the proportion of delabeling unnecessary penicillin labels to 20% for all pediatric inpatients.

Assessing the Benefits and Costs of the Hydrogen Cyanide Antiherbivore Defense in .

Understanding the evolution of plant defenses against herbivores requires identifying the benefits and costs of defense. Here, we tested the hypothesis that the benefits and costs of hydrogen cyanide (HCN) defense against herbivory on white clover () are temperature dependent. We first tested how temperature affected HCN production in vitro, and then examined how temperature influenced the efficacy of HCN defense of against a generalist slug () herbivore using no-choice and choice feeding trial assays.

Correction: Differential Subplastidial Localization and Turnover of Enzymes Involved in Isoprenoid Biosynthesis in Chloroplasts.

[This corrects the article DOI: 10.1371/journal.pone.

Bark Beetle Attack History Does Not Influence the Induction of Terpene and Phenolic Defenses in Mature Norway Spruce () Trees by the Bark Beetle-Associated Fungus .

Terpenes and phenolics are important constitutive and inducible conifer defenses against bark beetles and their associated fungi. In this study, the inducible defenses of mature Norway spruce () trees with different histories of attack by the spruce bark beetle, were tested by inoculation with the -associated fungus . We compared trees that had been under previous attack with those under current attack and those that had no record of attack.

An Arabidopsis GCMS chemical ionization technique to quantify adaptive responses in central metabolism.

We present a methodology to survey central metabolism in 13CO2-labeled Arabidopsis (Arabidopsis thaliana) rosettes by ammonia positive chemical ionization-gas chromatography-mass spectrometry. This technique preserves the molecular ion cluster of methyloxime/trimethylsilyl-derivatized analytes up to 1 kDa, providing unambiguous nominal mass assignment of >200 central metabolites and 13C incorporation rates into a subset of 111 from the tricarboxylic acid (TCA) cycle, photorespiratory pathway, amino acid metabolism, shikimate pathway, and lipid and sugar metabolism. In short-term labeling assays, we observed plateau labeling of ∼35% for intermediates of the photorespiratory cycle except for glyoxylate, which reached only ∼4% labeling and was also present at molar concentrations several fold lower than other photorespiratory intermediates.

Soapbark Triterpenes: Quillaja brasiliensis Cell Culture Sapogenin and Free Sterol Analysis by GCMS.

Triterpene saponins of the genus Quillaja (Quillajaceae) are known for their immunoadjuvant, hypocholesterolemic, and anti-inflammatory activity. Plant cell cultures are useful for the study of saponin metabolism and industrial production of these bioactive compounds. While structurally related phytosterols are primary metabolites essential to growth and development, saponins are responsive to pathogen and abiotic stress, fulfilling roles in plant specialized metabolism.

Design and fabrication of an improved dynamic flow cuvette for CO labeling in Arabidopsis plants.

Background: Stable isotope labeling is a non-invasive, sensitive means of monitoring metabolic flux in plants. The most physiologically meaningful information is obtained from experiments that take advantage of the natural photosynthetic carbon assimilation pathway to introduce a traceable marker with minimal effects on the physiology of the organism. The fundamental substrate in isotopic labeling experiments is CO, which can reveal the earliest events in carbon assimilation and realistically portray downstream metabolism when administered under conditions suitable for making kinetic inferences.

CryoSIM: super-resolution 3D structured illumination cryogenic fluorescence microscopy for correlated ultrastructural imaging.

Rapid cryopreservation of biological specimens is the gold standard for visualizing cellular structures in their true structural context. However, current commercial cryo-fluorescence microscopes are limited to low resolutions. To fill this gap, we have developed cryoSIM, a microscope for 3D super-resolution fluorescence cryo-imaging for correlation with cryo-electron microscopy or cryo-soft X-ray tomography.

Cytosolic geraniol and citronellol biosynthesis require a Nudix hydrolase in rose-scented geranium (Pelargonium graveolens).

Geraniol, citronellol and their esters are high-value acyclic monoterpenes used in food technology, perfumery and cosmetics. A major source of these compounds is the essential oil of rose-scented geraniums of the genus Pelargonium. We provide evidence that their biosynthesis mainly takes place in the cytosol of glandular trichomes via geranyl monophosphate (GP) through the action of a Nudix hydrolase.

Isotope ratio-based quantification of carbon assimilation highlights the role of plastidial isoprenoid precursor availability in photosynthesis.

Background: We report a method to estimate carbon assimilation based on isotope ratio-mass spectrometry (IRMS) of CO labeled plant tissue. Photosynthetic carbon assimilation is the principal experimental observable which integrates important aspects of primary plant metabolism. It is traditionally measured through gas exchange.

Negative regulation of plastidial isoprenoid pathway by herbivore-induced β-cyclocitral in .

Insect damage to plants is known to up-regulate defense and down-regulate growth processes. While there are frequent reports about up-regulation of defense signaling and production of defense metabolites in response to herbivory, much less is understood about the mechanisms by which growth and carbon assimilation are down-regulated. Here we demonstrate that insect herbivory down-regulates the 2--methyl-D-erythritol-4-phosphate (MEP) pathway in (), a pathway making primarily metabolites for use in photosynthesis.

Implementation of a 4Pi-SMS super-resolution microscope.

The development of single-molecule switching (SMS) fluorescence microscopy (also called single-molecule localization microscopy) over the last decade has enabled researchers to image cell biological structures at unprecedented resolution. Using two opposing objectives in a so-called 4Pi geometry doubles the available numerical aperture, and coupling this with interferometric detection has demonstrated 3D resolution down to 10 nm over entire cellular volumes. The aim of this protocol is to enable interested researchers to establish 4Pi-SMS super-resolution microscopy in their laboratories.

Genetic dissection of climacteric fruit ripening in a melon population segregating for ripening behavior.

Melon is as an alternative model to understand fruit ripening due to the coexistence of climacteric and non-climacteric varieties within the same species, allowing the study of the processes that regulate this complex trait with genetic approaches. We phenotyped a population of recombinant inbred lines (RILs), obtained by crossing a climacteric (Védrantais, cantalupensis type) and a non-climcteric variety (Piel de Sapo T111, inodorus type), for traits related to climacteric maturation and ethylene production. Individuals in the RIL population exhibited various combinations of phenotypes that differed in the amount of ethylene produced, the early onset of ethylene production, and other phenotypes associated with ripening.

3D Correlative Cryo-Structured Illumination Fluorescence and Soft X-ray Microscopy Elucidates Reovirus Intracellular Release Pathway.

Imaging of biological matter across resolution scales entails the challenge of preserving the direct and unambiguous correlation of subject features from the macroscopic to the microscopic level. Here, we present a correlative imaging platform developed specifically for imaging cells in 3D under cryogenic conditions by using X-rays and visible light. Rapid cryo-preservation of biological specimens is the current gold standard in sample preparation for ultrastructural analysis in X-ray imaging.

Medically Useful Plant Terpenoids: Biosynthesis, Occurrence, and Mechanism of Action.

Specialized plant terpenoids have found fortuitous uses in medicine due to their evolutionary and biochemical selection for biological activity in animals. However, these highly functionalized natural products are produced through complex biosynthetic pathways for which we have a complete understanding in only a few cases. Here we review some of the most effective and promising plant terpenoids that are currently used in medicine and medical research and provide updates on their biosynthesis, natural occurrence, and mechanism of action in the body.

Distinct metabolic pathways drive monoterpenoid biosynthesis in a natural population of Pelargonium graveolens.

Pelargonium graveolens is a wild predecessor to rose-scented geranium hybrids prized for their essential oils used as fragrances and flavorings. However, little is known about their biosynthesis. Here we present metabolic evidence that at least two distinct monoterpene biosynthetic pathways contribute to their volatile profiles, namely, cyclic p-menthanes such as (-)-isomenthone and acyclic monoterpene alcohols such as geraniol and (-)-citronellol and their derivatives (referred to here as citronelloid monoterpenes).

The plastidial metabolite 2-C-methyl-D-erythritol-2,4-cyclodiphosphate modulates defence responses against aphids.

Feeding by insect herbivores such as caterpillars and aphids induces plant resistance mechanisms that are mediated by the phytohormones jasmonic acid (JA) and salicylic acid (SA). These phytohormonal pathways often crosstalk. Besides phytohormones, methyl-D-erythriol-2,4-cyclodiphosphate (MEcPP), the penultimate metabolite in the methyl-D-erythritol-4-phosphate pathway, has been speculated to regulate transcription of nuclear genes in response to biotic stressors such as aphids.

Nerolidol production in agroinfiltrated tobacco: Impact of protein stability and membrane targeting of strawberry (Fragraria ananassa) NEROLIDOL SYNTHASE1.

The sesquiterpene alcohol nerolidol, synthesized from farnesyl diphosphate (FDP), mediates plant-insect interactions across multiple trophic levels with major implications for pest management in agriculture. We compared nerolidol engineering strategies in tobacco using agroinfiltration to transiently express strawberry (Fragraria ananassa) linalool/nerolidol synthase (FaNES1) either at the endoplasmic reticulum (ER) or in the cytosol as a soluble protein. Using solid phase microextraction and gas chromatography-mass spectrometry (SPME-GCMS), we have determined that FaNES1 directed to the ER via fusion to the transmembrane domain of squalene synthase or hydroxymethylglutaryl - CoA reductase displayed significant improvements in terms of transcript levels, protein accumulation, and volatile production when compared to its cytosolic form.

Non-invasive quantification of ethylene in attached fruit headspace at 1 p.p.b. by gas chromatography-mass spectrometry.

Ethylene is a gaseous plant hormone involved in defense, adaptations to environmental stress and fruit ripening. Its relevance to the latter makes its detection highly useful for physiologists interested in the onset of ripening. Produced as a sharp peak during the respiratory burst, ethylene is biologically active at tens of nl L .

Differential Subplastidial Localization and Turnover of Enzymes Involved in Isoprenoid Biosynthesis in Chloroplasts.

Plastidial isoprenoids are a diverse group of metabolites with roles in photosynthesis, growth regulation, and interaction with the environment. The methylerythritol 4-phosphate (MEP) pathway produces the metabolic precursors of all types of plastidial isoprenoids. Proteomics studies in Arabidopsis thaliana have shown that all the enzymes of the MEP pathway are localized in the plastid stroma.