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BACKGROUND: Fucoxanthin, a marine xanthophyll carotenoid, has been shown to exert beneficial health effects. Cell-based and animal-based experimental studies have shown that fucoxanthin has the potential to mitigate eczema symptoms. Hence, we sought to assess whether fucoxanthinol 3-arachidate, a fucoxanthin metabolite, measured in maternal serum at birth is associated with eczema development during early childhood. METHODS: Data from the 1989/1990 Isle of Wight birth cohort were analyzed. We focused on data obtained from the 1, 2, and 4 years follow-ups. Fucoxanthinol 3-arachidate was measured in maternal serum at the child's birth as abundance relative to the reference lipids. Eczema was ascertained according to parent-reported clinical history and characteristic morphology and distribution. Log-binomial regression models were used to estimate adjusted risk ratios (aRR) and their 95% confidence intervals (CI). RESULTS: A total of 592 subjects (49.2% males and 50.8% females) were included in the current analysis. Associations between fucoxanthinol 3-arachidate levels and eczema risk during the first 4 years of life (longitudinal analysis) were evaluated using four modeling approaches, which showed higher fucoxanthinol 3-arachidate levels were associated with reduced eczema risk: (i) aRRper 10 unit increase = 0.88, 95% CI: 0.76-1.03; (ii) aRR>0 vs. =0 = 0.67, 0.45-0.99; (iii) aRR≥2.3 vs. <2.3 = 0.66, 0.44-0.98; and (iv) aRRtertile 3 vs. tertile 1 = 0.65, 0.42-0.99. CONCLUSION: Our findings suggest that increased fucoxanthinol 3-arachidate levels measured in maternal serum at the child's birth is associated with reduced eczema risk during the first 4 years of the offspring life.
Assuntos
Eczema , Xantofilas , Masculino , Feminino , Animais , Pré-Escolar , Humanos , Estudos de Coortes , Xantofilas/metabolismo , Eczema/epidemiologiaRESUMO
The role of metabolites, nutrients, and toxins (MNTs) in sera at the end of pregnancy and of their association with offspring respiratory and allergic disorders is underexplored. Untargeted approaches detecting a variety of compounds, known and unknown, are limited. In this cohort study, we first aimed at discovering associations of MNTs in grandmaternal (F0) serum with asthma, immunoglobulin E, skin prick tests, exhaled nitric oxide, and lung function parameters in their parental (F1) offspring. Second, for replication, we tested the identified associations of MNTs with disorders in their grandchildren (F2-offspring) based on F2 cord serum. The statistical analyses were sex-stratified. Using liquid chromatography/high-resolution mass spectrometry in F0, we detected signals for 2286 negative-ion lipids, 59 positive-ion lipids, and 6331 polar MNTs. Nine MNTs (one unknown MNT) discovered in F0-F1 and replicated in F2 showed higher risks of respiratory/allergic outcomes. Twelve MNTs (four unknowns) constituted a potential protection in F1 and F2. We recognized MNTs not yet considered candidates for respiratory/allergic outcomes: a phthalate plasticizer, an antihistamine, a bile acid metabolite, tryptophan metabolites, a hemiterpenoid glycoside, triacylglycerols, hypoxanthine, and polyphenol syringic acid. The findings suggest that MNTs are aspirants for clinical trials to prevent adverse respiratory/allergic outcomes.
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Plants make a variety of specialized metabolites that can mediate interactions with animals, microbes, and competitor plants. Understanding how plants synthesize these compounds enables studies of their biological roles by manipulating their synthesis in vivo as well as producing them in vitro. Acylsugars are a group of protective metabolites that accumulate in the trichomes of many Solanaceae family plants. Acylinositol biosynthesis is of interest because it appears to be restricted to a subgroup of species within the Solanum genus. Previous work characterized a triacylinositol acetyltransferase involved in acylinositol biosynthesis in the Andean fruit plant Solanum quitoense (lulo or naranjilla). We characterized three additional S. quitoense trichome expressed enzymes and found that virus-induced gene silencing of each caused changes in acylinositol accumulation. pH was shown to influence the stability and rearrangement of the product of ASAT1H and could potentially play a role in acylinositol biosynthesis. Surprisingly, the in vitro triacylinositol products of these enzymes are distinct from those that accumulate in planta. This suggests that additional enzymes are required in acylinositol biosynthesis. These characterized S. quitoense enzymes, nonetheless, provide opportunities to test the biological impact and properties of these triacylinositols in vitro.
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Plant alkaloids constitute an important class of bioactive chemicals with applications in medicine and agriculture. However, the knowledge gap of the diversity and biosynthesis of phytoalkaloids prevents systematic advances in biotechnology for engineered production of these high-value compounds. In particular, the identification of cytochrome P450s driving the structural diversity of phytoalkaloids has remained challenging. Here, we use a combination of reverse genetics with discovery metabolomics and multivariate statistical analysis followed by in planta transient assays to investigate alkaloid diversity and functionally characterize two candidate cytochrome P450s genes from Atropa belladonna without a priori knowledge of their functions or information regarding the identities of key pathway intermediates. This approach uncovered a largely unexplored root localized alkaloid sub-network that relies on pseudotropine as precursor. The two cytochrome P450s catalyze N-demethylation and ring-hydroxylation reactions within the early steps in the biosynthesis of diverse N-demethylated modified tropane alkaloids.
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Alcaloides , Tropanos , Alcaloides/química , Sistema Enzimático do Citocromo P-450/genética , Metabolômica , Tropanos/metabolismoRESUMO
Acylsugars are defensive, trichome-synthesized sugar esters produced in plants across the Solanaceae (nightshade) family. Although assembled from simple metabolites and synthesized by a relatively short core biosynthetic pathway, tremendous within- and across-species acylsugar structural variation is documented across the family. To advance our understanding of the diversity and the synthesis of acylsugars within the Nicotiana genus, trichome extracts were profiled across the genus coupled with transcriptomics-guided enzyme discovery and in vivo and in vitro analysis. Differences in the types of sugar cores, numbers of acylations, and acyl chain structures contributed to over 300 unique annotated acylsugars throughout Nicotiana. Placement of acyl chain length into a phylogenetic context revealed that an unsaturated acyl chain type was detected in a few closely related species. A comparative transcriptomics approach identified trichome-enriched Nicotiana acuminata acylsugar biosynthetic candidate enzymes. More than 25 acylsugar variants could be produced in a single enzyme assay with four N. acuminata acylsugar acyltransferases (NacASAT1-4) together with structurally diverse acyl-CoAs and sucrose. Liquid chromatography coupled with mass spectrometry screening of in vitro products revealed the ability of these enzymes to make acylsugars not present in Nicotiana plant extracts. In vitro acylsugar production also provided insights into acyltransferase acyl donor promiscuity and acyl acceptor specificity as well as regiospecificity of some ASATs. This study suggests that promiscuous Nicotiana acyltransferases can be used as synthetic biology tools to produce novel and potentially useful metabolites.
Assuntos
Aciltransferases , Tricomas , Aciltransferases/genética , Aciltransferases/metabolismo , Carboidratos , Filogenia , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Açúcares/metabolismo , Biologia Sintética , Nicotiana/genética , Nicotiana/metabolismo , Tricomas/metabolismoRESUMO
Acetaminophen is used by nearly two-thirds of pregnant women. Although considered safe, studies have demonstrated associations between prenatal acetaminophen use and adverse health outcomes in offspring. Since DNA methylation (DNAm) at birth may act as an early indicator of later health, assessments on whether DNAm of newborns is associated with gestational acetaminophen use or its metabolites are needed. Using data from three consecutive generations of the Isle of Wight cohort (F0-grandmothers, F1-mothers, and F2-offspring) we investigated associations between acetaminophen metabolites in F0 serum at delivery with epigenome-wide DNAm in F1 (Guthrie cards) and between acetaminophen use of F1 and F2-cord-serum levels with F2 cord blood DNAm. In epigenome-wide screening, we eliminated non-informative DNAm sites followed by linear regression of informative sites. Based on repeated pregnancies, indication bias analyses tested whether acetaminophen indicated maternal diseases or has a risk in its own right. Considering that individuals with similar intake process acetaminophen differently, metabolites were clustered to distinguish metabolic exposures. Finally, metabolite clusters from F1-maternal and F2-cord sera were tested for their associations with newborn DNAm (F1 and F2). Twenty-one differential DNAm sites in cord blood were associated with reported maternal acetaminophen intake in the F2 generation. For 11 of these cytosine-phosphate-guanine (CpG) sites, an indication bias was excluded and five were replicated in F2 with metabolite clusters. In addition, metabolite clusters showed associations with 25 CpGs in the F0-F1 discovery analysis, of which five CpGs were replicated in the F2-generation. Our results suggest that prenatal acetaminophen use, measured as metabolites, may influence DNAm in newborns.
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Plants synthesize myriad phylogenetically restricted specialized (aka "secondary") metabolites with diverse structures. Metabolism of acylated sugar esters in epidermal glandular secreting trichomes across the Solanaceae (nightshade) family is ideal for investigating the mechanisms of evolutionary metabolic diversification. We developed methods to structurally analyze acylhexose mixtures by 2D NMR, which led to the insight that the Old World species black nightshade (Solanum nigrum) accumulates acylglucoses and acylinositols in the same tissue. Detailed in vitro biochemistry, cross-validated by in vivo virus-induced gene silencing, revealed two unique features of the four-step acylglucose biosynthetic pathway: A trichome-expressed, neofunctionalized invertase-like enzyme, SnASFF1, converts BAHD-produced acylsucroses to acylglucoses, which, in turn, are substrates for the acylglucose acyltransferase, SnAGAT1. This biosynthetic pathway evolved independently from that recently described in the wild tomato Solanum pennellii, reinforcing that acylsugar biosynthesis is evolutionarily dynamic with independent examples of primary metabolic enzyme cooption and additional variation in BAHD acyltransferases.
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Acylsugars constitute an abundant class of pest- and pathogen-protective Solanaceae family plant-specialized metabolites produced in secretory glandular trichomes. Solanum pennellii produces copious triacylated sucrose and glucose esters, and the core biosynthetic pathway producing these compounds was previously characterized. We performed untargeted metabolomic analysis of S. pennellii surface metabolites from accessions spanning the species range, which indicated geographic trends in the acylsugar profile and revealed two compound classes previously undescribed from this species, tetraacylglucoses and flavonoid aglycones. A combination of ultrahigh-performance liquid chromatography-high resolution mass spectrometry (UHPLC-HR-MS) and NMR spectroscopy identified variations in the number, length, and branching pattern of acyl chains, and the proportion of sugar cores in acylsugars among accessions. The new dimensions of acylsugar variation revealed by this analysis further indicate variation in the biosynthetic and degradative pathways responsible for acylsugar accumulation. These findings provide a starting point for deeper investigation of acylsugar biosynthesis, an understanding of which can be exploited through crop breeding or metabolic engineering strategies to improve the endogenous defenses of crop plants.
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Nicotine is a major constituent of cigarette smoke. Its primary metabolite in maternal and cord sera, cotinine, is considered a biomarker of prenatal smoking. Nicotine and cotinine half-lives are decreased in pregnancy due to their increased rate of metabolism and conversion to downstream metabolites such as norcotinine and 3-hydroxycotinine. Hence, downstream metabolites of nicotine may provide informative biomarkers of prenatal smoking. In this study of three generations (F0-mothers, F1-offspring who became mothers, and F2-offspring), we present a biochemical assessment of prenatal smoking exposure based on maternal and cord sera levels of nicotine, cotinine, norcotinine, and 3-hydroxycotinine. As potential markers of early effects of prenatal smoking, associations with differential DNA methylation (DNAm) in the F1- and F2-offspring were assessed. All metabolites in maternal and cord sera were associated with self-reported prenatal smoking, except for nicotine. We compared maternal self-report of smoking in pregnancy to biochemical evidence of prenatal smoking exposure. Self-report of F0-mothers of F1 in 1989-1990 had more accuracy identifying prenatal smoking related to maternal metabolites in maternal serum (sensitivity = 94.6%, specificity = 86.9%) compared to self-reports of F1-mothers of F2 (2010-2016) associated with cord serum markers (sensitivity = 66.7%, specificity = 78.8%). Nicotine levels in sera showed no significant association with any DNAm site previously linked to maternal smoking. Its downstream metabolites, however, were associated with DNAm sites located on the MYO1G, AHRR, and GFI1 genes. In conclusion, cotinine, norcotinine, and 3-hydroxycotinine in maternal and cord sera provide informative biomarkers and should be considered when assessing prenatal smoking. The observed association of offspring DNAm with metabolites, except for nicotine, may imply that the toxic effects of prenatal nicotine exposure are exerted by downstream metabolites, rather than nicotine. If differential DNA methylation on the MYO1G, AHRR, and GFI1 genes transmit adverse effects of prenatal nicotine exposure to the child, there is a need to investigate whether preventing changes in DNA methylation by reducing the metabolic rate of nicotine and conversion to harmful metabolites may protect exposed children.
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Metilação de DNA , Nicotina , Efeitos Tardios da Exposição Pré-Natal , Fumar , Biomarcadores , Criança , Cotinina/análise , Feminino , Humanos , Recém-Nascido , Masculino , Exposição Materna , Nicotina/análise , Gravidez , Efeitos Tardios da Exposição Pré-Natal/genética , Fumar/efeitos adversosRESUMO
Prior work documented use of γ-phosphate modified ATP analogs to label DNA using T4 polynucleotide kinases (T4PNK), although applications have been limited. To fully characterize kinase-catalyzed labeling of nucleic acids, we explored use of ATP-biotin as a cosubstrate with T4PNK. T4PNK accepted ATP-biotin to 5'-label single stranded DNA. However, T4PNK-mediated labeling of double stranded substrates was low yielding. In addition, the phosphoramidate bond connecting the biotin group to the DNA was unstable. These results suggest that kinase-catalyzed biotinylation will be useful with single stranded DNA substrates and mild reaction conditions. By revealing the scope and limitations of kinase-catalyzed biotinylation, these studies provide a foundation for future development and application of kinase-catalyzed labeling to DNA-based biological studies.