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1.
Nature ; 583(7816): 415-420, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32555456

RESUMO

Animals coexist in commensal, pathogenic or mutualistic relationships with complex communities of diverse organisms, including microorganisms1. Some bacteria produce bioactive neurotransmitters that have previously been proposed to modulate nervous system activity and behaviours of their hosts2,3. However, the mechanistic basis of this microbiota-brain signalling and its physiological relevance are largely unknown. Here we show that in Caenorhabditis elegans, the neuromodulator tyramine produced by commensal Providencia bacteria, which colonize the gut, bypasses the requirement for host tyramine biosynthesis and manipulates a host sensory decision. Bacterially produced tyramine is probably converted to octopamine by the host tyramine ß-hydroxylase enzyme. Octopamine, in turn, targets the OCTR-1 octopamine receptor on ASH nociceptive neurons to modulate an aversive olfactory response. We identify the genes that are required for tyramine biosynthesis in Providencia, and show that these genes are necessary for the modulation of host behaviour. We further find that C. elegans colonized by Providencia preferentially select these bacteria in food choice assays, and that this selection bias requires bacterially produced tyramine and host octopamine signalling. Our results demonstrate that a neurotransmitter produced by gut bacteria mimics the functions of the cognate host molecule to override host control of a sensory decision, and thereby promotes fitness of both the host and the microorganism.


Assuntos
Caenorhabditis elegans/microbiologia , Caenorhabditis elegans/fisiologia , Comportamento Alimentar/fisiologia , Intestinos/microbiologia , Neurotransmissores/metabolismo , Providencia/metabolismo , Olfato/fisiologia , Animais , Aprendizagem da Esquiva/efeitos dos fármacos , Caenorhabditis elegans/efeitos dos fármacos , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Microbioma Gastrointestinal/fisiologia , Metabolômica , Mutação , Octanóis/farmacologia , Octopamina/biossíntese , Octopamina/metabolismo , Providencia/enzimologia , Providencia/fisiologia , Receptores de Amina Biogênica/metabolismo , Receptores Acoplados a Proteínas-G/metabolismo , Células Receptoras Sensoriais/metabolismo , Olfato/efeitos dos fármacos , Tiramina/biossíntese , Tiramina/metabolismo , Tirosina Descarboxilase/deficiência , Tirosina Descarboxilase/genética
2.
PLoS One ; 15(4): e0231976, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32324789

RESUMO

The two alkaloids gramine and hordenine have been known for playing a role in the allelopathic ability in barley (Hordeum vulgare L.). These allelochemicals can be both found in leaves and roots in some barley cultivars whereas in others one seems to exclude the other. In this study eighteen accessions of barley from the Middle-East area, one accession from Tibet and the modern spring cultivar Barke, already used as parental donor in a nested associated mapping (NAM) population, were screened for their gramine, hordenine and N-methyltyramine (the direct precursor of hordenine) content in leaves, roots and exudates. Moreover, the toxicity of the three allelochemicals on root growth inhibition on lettuce (Lactuca sativa L.) was evaluated. Results of this study showed the preferential production of gramine and hordenine in leaves and roots, respectively, in the nineteen barley accessions. On the other hand, in the modern barley cultivar Barke, the highest content of hordenine in roots and the general lack of gramine suggests a favored biosynthesis of the former. Gramine was not detected in the root exudates. In additions, different metabolomic profiles were observed in wild relatives compared to modern barley genotypes. The results also showed the phytotoxic effects of the three compounds on root growth of lettuce seedlings, with a reduction in root length and an increase of root surface area and diameter. In conclusion, this study highlighted the impact of the domestication effects on the production and distribution of the two allelopathic alkaloids gramine and hordenine in barley.


Assuntos
Alelopatia , Genótipo , Hordeum/genética , Hordeum/metabolismo , Filogenia , Hordeum/crescimento & desenvolvimento , Alcaloides Indólicos/metabolismo , Metabolômica , Especificidade da Espécie , Tiramina/análogos & derivados , Tiramina/metabolismo
3.
Life Sci ; 253: 117696, 2020 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-32334013

RESUMO

AIMS: We have previously demonstrated that p-tyramine (TYR), an endogenous trace amine-associated receptor 1 agonist, passage across neuronal membranes involves a transporter exhibiting the pharmacological profile of Organic Cation Transporter 2 (OCT2). Since TYR is also a constituent of foodstuffs and produced by the intestinal microbiota, here we have investigated whether similar processes are involved in the passage of 100 nM TYR across apical and basolateral membranes of the Caco-2 human intestinal epithelial cell line. MATERIALS AND METHODS: [3H]TYR transport across apical and basolateral membranes of Caco-2 cell monolayers was measured in the presence of inhibitors of TYR metabolizing enzymes. Cellular, apical, and basolateral compartments were collected at various timepoints, TYR concentrations calculated, and transport properties pharmacologically characterized. KEY FINDINGS: Apical transport resulted in equimolar accumulation of TYR within cells. Pentamidine (OCT1/OCT2 inhibitor) decreased apical transport (P = 0.001) while atropine (OCT1 inhibitor) had no effect, suggesting apical transport involved OCT2. In contrast, basolateral transport resulted in 500-1000 nM cellular concentrations (P < 0.0001) indicating the presence of an active transporter. Replacement of Na+ on an equimolar basis with choline resulted in loss of TYR transport (P = 0.017). Unexpectedly, this active transport was also atropine-sensitive (P = 0.020). Kinetic analysis of the active transporter revealed Vmax = 43.0 nM/s with a Kt = 33.1 nM. SIGNIFICANCE: We have demonstrated for the first time that TYR is transported across Caco-2 apical membranes via facilitated diffusion by OCT2, whereas transport across basolateral membranes is by a Na+-dependent, atropine-sensitive, active transporter.


Assuntos
Mucosa Intestinal/metabolismo , Transportador 2 de Cátion Orgânico/metabolismo , Sódio/metabolismo , Tiramina/metabolismo , Atropina/metabolismo , Transporte Biológico/fisiologia , Transporte Biológico Ativo/fisiologia , Células CACO-2 , Humanos
4.
J Vis Exp ; (156)2020 02 21.
Artigo em Inglês | MEDLINE | ID: mdl-32150172

RESUMO

Immunostaining is widely used in biomedical research to show the cellular expression pattern of a given protein. Multiplex immunostaining allows labeling using multiple primary antibodies. To minimize antibody cross-reactivity, multiplex immunostaining using indirect staining requires unlabeled primary antibodies from different host species. However, the appropriate combination of different species antibodies is not always available. Here, we describe a method of using unlabeled primary antibodies from the same host species (e.g., in this case both antibodies are from rabbit) for multiplex immunofluorescence on formalin-fixed paraffin-embedded (FFPE) mouse adrenal sections. This method uses the same procedure and reagents used in the antigen retrieval step to strip the activity of the previously stained primary antibody complex. Slides were stained with the first primary antibody using a general immunostaining protocol followed by a binding step with a biotinylated secondary antibody. Then, an avidin-biotin-peroxidase signal development method was used with fluorophore-tyramide as the substrate. The immunoactivity of the first primary antibody complex was stripped through immersion in a microwaved boiling sodium citrate solution for 8 min. The insoluble fluorophore-tyramide deposition remained on the sample, which allowed the slide to be stained with other primary antibodies. Although this method eliminates most false positive signals, some background from antibody cross-reactivity may remain. If the samples are enriched with endogenous biotin, a peroxidase-conjugated secondary antibody may be used to replace the biotinylated secondary antibody to avoid the false positive from recovered endogenous biotin.


Assuntos
Glândulas Suprarrenais/metabolismo , Anticorpos Monoclonais/imunologia , Corantes Fluorescentes/química , Técnicas Imunoenzimáticas/métodos , Micro-Ondas , Tiramina/análogos & derivados , 3-Hidroxiesteroide Desidrogenases/imunologia , Glândulas Suprarrenais/imunologia , Animais , Biotinilação , Sistema Enzimático do Citocromo P-450/imunologia , Imunofluorescência , Humanos , Camundongos , Peroxidase/metabolismo , Coelhos , Coloração e Rotulagem , Tiramina/metabolismo
5.
Insect Biochem Mol Biol ; 120: 103337, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32109588

RESUMO

As the counterparts of noradrenaline and adrenaline in vertebrates, octopamine (OA) regulates multiple physiological and behavioral processes in invertebrate. OA mediates its effects via binding to specific octopamine receptors (OARs). Functional and pharmacological characterization of OARs have been reported in several insects. However, little work was documented in hemipteran insects. We cloned a ß-adrenergic-like OAR (NcOA2B2) from Nephotettix cincticeps. NcOA2B2 shares high similarity with members of the OA2B2 receptor class. Transcript level of NcOA2B2 varied in various tissues and was highly expressed in the leg. After heterologous expression in CHO-K1 cells, NcOA2B2 was dose-dependently activated by OA (EC50 = 2.56 nM) and tyramine (TA) (EC50 = 149 nM). Besides putative octopaminergic agonists, dopaminergic agonists and amitraz and DPMF potently activated NcOA2B2 in a dose-dependent manner. Receptor activity was blocked by potential antagonists and was most efficiently antagonized by asenapine. Phentolamine showed both antagonist and agonist effects on NcOA2B2. Our results offer the important information about molecular and pharmacological characterization of an OAR from N. cincticeps that will provide the basis for forthcoming studies on its roles in physiological processes and behaviors, and facilitate the design of novel insecticides for pest control.


Assuntos
Regulação da Expressão Gênica , Hemípteros/genética , Proteínas de Insetos/genética , Receptores de Amina Biogênica/genética , Sequência de Aminoácidos , Animais , AMP Cíclico/metabolismo , Dopamina/metabolismo , Hemípteros/metabolismo , Proteínas de Insetos/química , Proteínas de Insetos/metabolismo , Octopamina/metabolismo , Filogenia , Receptores de Amina Biogênica/química , Receptores de Amina Biogênica/metabolismo , Alinhamento de Sequência , Tiramina/metabolismo
6.
J Biochem ; 167(1): 49-54, 2020 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-31647557

RESUMO

The isotope effects approach was used to elucidate the mechanism of oxidative deamination of 3'-halotyramines, catalyzed by monoamine oxidase A (EC 1.4.3.4). The numerical values of kinetic isotope effect (KIE) and solvent isotope effect (SIE) were established using a non-competitive spectrophotometric technique. Based upon KIE and SIE values, some of the mechanistic details of investigated reaction were discussed.


Assuntos
Monoaminoxidase/metabolismo , Tiramina/metabolismo , Biocatálise , Radioisótopos de Flúor , Humanos , Isótopos de Iodo , Cinética , Oxirredução , Solventes/química
7.
J Pharm Pharm Sci ; 22(1): 585-592, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31804922

RESUMO

PURPOSE: The effects of selective serotonin reuptake inhibitors (SSRIs) such as fluoxetine and paroxetine on dopamine formation from p-tyramine, mediated by cytochrome P450 (CYP) 2D6.2 (Arg296Cys, Ser486Thr) and CYP2D6.10 (Pro34Ser, Ser486Thr), were compared with their effects on CYP2D6.1 (wild type)-mediated dopamine formation, to investigate the influence of a CYP2D6 polymorphism on neuroactive amine metabolism in the brain. METHODS: The Michaelis constants (Km) and maximal velocity (Vmax) values of dopamine formation mediated by CYP2D6.1, CYP2D6.2, and CYP2D6.10 (expressed in recombinant Escherichia coli), and inhibition constants (Ki) of the SSRIs toward dopamine formation catalyzed by the CYP2D6 variants were estimated. RESULTS: The Km values for CYP2D6.2 and CYP2D6.10 decreased at lower fluoxetine concentrations, while the Vmax values for all CYP2D6 variants increased, indicating that fluoxetine stimulated dopamine formation. Conversely, paroxetine competitively inhibited dopamine formation mediated by CYP2D6.1, CYP2D6.2, and CYP2D6.10 with Ki values of 0.47, 1.33, and 31.3 µM, respectively. CONCLUSIONS: These results suggest that the inhibition/stimulation of CYP2D6-mediated dopamine formation by these SSRIs would be affected by CYP2D6 polymorphisms in the brain.


Assuntos
Citocromo P-450 CYP2D6/metabolismo , Dopamina/biossíntese , Fluoxetina/farmacologia , Paroxetina/farmacologia , Inibidores de Captação de Serotonina/farmacologia , Tiramina/metabolismo , Dopamina/análise , Fluoxetina/química , Humanos , Estrutura Molecular , Paroxetina/química , Inibidores de Captação de Serotonina/química
8.
Sci Rep ; 9(1): 16275, 2019 11 07.
Artigo em Inglês | MEDLINE | ID: mdl-31700013

RESUMO

Tyramine receptor (TyrR) is a biogenic amine G protein-coupled receptor (GPCR) associated with many important physiological functions in insect locomotion, reproduction, and pheromone response. Binding of specific ligands to the TyrR triggers conformational changes, relays the signal to G proteins, and initiates an appropriate cellular response. Here, we monitor the binding effect of agonist compounds, tyramine and amitraz, to a Sitophilus oryzae tyramine receptor (SoTyrR) homology model and their elicited conformational changes. All-atom molecular dynamics (MD) simulations of SoTyrR-ligand complexes have shown varying dynamic behavior, especially at the intracellular loop 3 (IL3) region. Moreover, in contrast to SoTyrR-tyramine, SoTyrR-amitraz and non-liganded SoTyrR shows greater flexibility at IL3 residues and were found to be coupled to the most dominant motion in the receptor. Our results suggest that the conformational changes induced by amitraz are different from the natural ligand tyramine, albeit being both agonists of SoTyrR. This is the first attempt to understand the biophysical implication of amitraz and tyramine binding to the intracellular domains of TyrR. Our data may provide insights into the early effects of ligand binding to the activation process of SoTyrR.


Assuntos
Ligantes , Conformação Molecular , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , Receptores de Amina Biogênica/química , Tiramina/química , Animais , Sítios de Ligação , Besouros , Humanos , Ligação Proteica , Receptores de Amina Biogênica/metabolismo , Tiramina/metabolismo
9.
Horm Mol Biol Clin Investig ; 40(1)2019 Nov 06.
Artigo em Inglês | MEDLINE | ID: mdl-31693494

RESUMO

Background Metabolic syndrome (MetS) is an important contributor to both type 2 diabetes mellitus (T2DM) and atherosclerotic cardiovascular disease (ASCVD). Although MetS affects one third of American adults, its pathogenesis remains to be elucidated. Tyramine, a derivative of tyrosine, has been shown to act as a catecholamine releasing agent in the human body. The aim of this study is to investigate the role of tyramine as an early biomarker for nascent MetS without the confounding of T2DM, ASCVD or smoking. Patients and methods This was an exploratory study of 28 patients with nascent MetS and 20 matched controls carried out in 2018. Metabolites were evaluated from patient's frozen early morning urine samples and were correlated with biomarkers of inflammation and adipokines. They were assayed by the National Institutes of Health (NIH) Western Metabolomics Center using liquid chromatography/mass spectrometry (LC/MS) and standardized to urinary creatinine. All patients had normal hepatic and renal function. Results Tyramine concentrations were significantly reduced in patients with MetS compared to controls, p = 0.0009. In addition, tyramine was significantly inversely correlated with multiple biomarkers of inflammation and cardiometabolic risk factors such as RBP4, monocyte TLR-4 abundance and P38MAPKinase activity, body mass index (BMI) and blood pressure (BP) (both systolic and diastolic). Conclusion In conclusion, low levels of tyramine could contribute to the proinflammatorty state of MetS.


Assuntos
Inflamação/urina , Síndrome Metabólica/urina , Tiramina/urina , Adulto , Idoso , Biomarcadores/metabolismo , Biomarcadores/urina , Feminino , Humanos , Inflamação/diagnóstico , Inflamação/metabolismo , Masculino , Síndrome Metabólica/diagnóstico , Síndrome Metabólica/metabolismo , Metabolômica , Pessoa de Meia-Idade , Tiramina/metabolismo
10.
Curr Opin Insect Sci ; 36: 125-130, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31606580

RESUMO

The biogenic amine octopamine and to some extent its precursor tyramine function as an alerting signal in insects. Octopaminergic/tyraminergic neurons arborize in most parts of the central nervous system and additionally reach almost all peripheral organs, tissues, and muscles. Indeed, octopamine is involved in motivation, arousal, and the initiation of different behaviors reflecting its function as an alerting signal. A well-studied example of octopamine function is feeding behavior in Drosophila. Here, the amine is involved in food search, sugar/bitter sensitivity, food intake, and starvation-induced hyperactivity. Thereby octopamine modulates feeding initiation in response to internal needs and external stimuli. Additionally, it seems that octopamine/tyramine orchestrate behaviors such as locomotion and feeding or flight and song production to adapt the behavioral outcome of an animal to physiological and environmental conditions. There is a possibility that octopamine and tyramine are required in the selection of behaviors in insects.


Assuntos
Comportamento Alimentar/fisiologia , Insetos , Octopamina/metabolismo , Animais , Comportamento Animal/fisiologia , Fenômenos Fisiológicos do Sistema Nervoso , Tiramina/metabolismo
11.
Nature ; 573(7772): 135-138, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31462774

RESUMO

An animal's stress response requires different adaptive strategies depending on the nature and duration of the stressor. Whereas acute stressors, such as predation, induce a rapid and energy-demanding fight-or-flight response, long-term environmental stressors induce the gradual and long-lasting activation of highly conserved cytoprotective processes1-3. In animals across the evolutionary spectrum, continued activation of the fight-or-flight response weakens the animal's resistance to environmental challenges4,5. However, the molecular and cellular mechanisms that regulate the trade-off between the flight response and long-term stressors are poorly understood. Here we show that repeated induction of the flight response in Caenorhabditis elegans shortens lifespan and inhibits conserved cytoprotective mechanisms. The flight response activates neurons that release tyramine, an invertebrate analogue of adrenaline and noradrenaline. Tyramine stimulates the insulin-IGF-1 signalling (IIS) pathway and precludes the induction of stress response genes by activating an adrenergic-like receptor in the intestine. By contrast, long-term environmental stressors, such as heat or oxidative stress, reduce tyramine release and thereby allow the induction of cytoprotective genes. These findings demonstrate that a neural stress hormone supplies a state-dependent neural switch between acute flight and long-term environmental stress responses and provides mechanistic insights into how the flight response impairs cellular defence systems and accelerates ageing.


Assuntos
Caenorhabditis elegans/citologia , Caenorhabditis elegans/fisiologia , Citoproteção , Insulina/metabolismo , Tiramina/metabolismo , Transporte Ativo do Núcleo Celular , Animais , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/metabolismo , Núcleo Celular/metabolismo , Fatores de Transcrição Forkhead/metabolismo , Fator de Crescimento Insulin-Like I/metabolismo , Mucosa Intestinal/metabolismo , Longevidade , Neurônios/metabolismo , Receptores Adrenérgicos/metabolismo , Receptores de Catecolaminas/metabolismo , Transdução de Sinais , Estresse Psicológico
12.
Food Chem ; 298: 125083, 2019 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-31261001

RESUMO

The effects of onion or caraway on changes in the content of biogenic amines were examined in sauerkraut during a fermentation process at 18 °C or 31 °C for 14 days and subsequent storage at 4 °C for 12 weeks. The amines were analysed by high-performance liquid chromatography with pre-column benzoylation. Total biogenic-amine concentration at the end of the fermentation was lower at 31 °C than at 18 °C. However, at this lower temperature, the presence of caraway or onion more significantly (than at 31 °C) reduced the total biogenic-amine content as compared to the control sample without an additive. After 12 weeks of refrigerated storage, concentrations of phenethylamine, tryptamine, and tyramine in the sauerkraut fermented with caraway (and concentrations of putrescine and tryptamine in the sauerkraut fermented with onion) at 31 °C increased as compared to the samples on the last day of fermentation, but did not pose a risk for consumer health.


Assuntos
Aminas Biogênicas/metabolismo , Carum , Alimentos e Bebidas Fermentados , Armazenamento de Alimentos/métodos , Cebolas , Aminas Biogênicas/análise , Brassica , Cromatografia Líquida de Alta Pressão , Temperatura Baixa , Fermentação , Alimentos e Bebidas Fermentados/análise , Humanos , Concentração de Íons de Hidrogênio , Putrescina/análise , Putrescina/metabolismo , Paladar , Tiramina/análise , Tiramina/metabolismo
13.
J Mol Neurosci ; 69(3): 371-379, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31290092

RESUMO

Inhibition of monoamine oxidase (MAO)-A/B can ameliorate depressive- and anxiety-related symptoms via increase of monoamine extracellular levels. However, such inhibition can also instigate hypertensive response following exposure to dietary tyramine (i.e., "the cheese effect"). Novel herbal treatment (NHT) is an herbal formula that has been demonstrated to reduce depressive- and anxiety-like symptoms in pre-clinical studies. The aim of the current study was to examine whether the therapeutic potential of NHT is underlain by inhibition of MAO-A/B and whether NHT poses a risk for tyramine hyper-potentiation. Unpredictable chronic mild stress (UCMS)-exposed mice and naïve mice were treated for 3 weeks with NHT (30 mg/kg; i.p.), the selective serotonin reuptake inhibitor (SSRI) escitalopram (15 mg/kg; i.p.), or saline. Subsequently, MAO-A/B activities in the hypothalamus, striatum, and prefrontal cortex (PFC) were assessed. Exposure to UCMS led to significant increases in both MAO-A and MAO-B activities in the hypothalamus (p < 0.001) and in the PFC (p < 0.01 for MAO-A; p < 0.001 for MAO-B). Neither NHT nor escitalopram had any notable effects. Treatment with NHT was supported as safe in terms of risk for inducing a hypertensive response. The antidepressant- and anxiolytic-like effects of NHT are mediated via pathways other than MAO-A/B inhibition.


Assuntos
Antidepressivos/uso terapêutico , Corpo Estriado/efeitos dos fármacos , Depressão/tratamento farmacológico , Hipotálamo/efeitos dos fármacos , Monoaminoxidase/análise , Proteínas do Tecido Nervoso/análise , Fitoterapia , Preparações de Plantas/uso terapêutico , Córtex Pré-Frontal/efeitos dos fármacos , Animais , Ansiolíticos/farmacologia , Ansiolíticos/uso terapêutico , Antidepressivos/farmacologia , Citalopram/uso terapêutico , Corpo Estriado/enzimologia , Crataegus , Depressão/etiologia , Avaliação Pré-Clínica de Medicamentos , Hipotálamo/enzimologia , Lilium , Camundongos , Camundongos Endogâmicos ICR , Monoaminoxidase/biossíntese , Córtex Pré-Frontal/enzimologia , Inibidores de Captação de Serotonina/uso terapêutico , Estresse Psicológico/psicologia , Triticum , Tiramina/metabolismo , Ziziphus
14.
Appl Microbiol Biotechnol ; 103(15): 6271-6285, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31161392

RESUMO

Proton magnetic resonance-based metabolomics analysis was performed to determine the global metabolite changes in pathogenic bacterium Pseudomonas aeruginosa PAO1 following exposure to quorum sensing (QS) inhibitor hordenine. Pyocyanin inhibition assay confirmed that hordenine exhibited potent QS inhibitory activity. A total of 40 metabolites were assigned by PMR spectra. Hordenine treatment resulted in the destruction of QS system in P. aeruginosa PAO1 by downregulating the expressions of genes involved in QS. The synthesis of antioxidant enzymes was repressed and the oxidative stress was enhanced due to the dysfunctional QS system of P. aeruginosa PAO1. The enhanced oxidative stress induced by the dysfunctional QS system of P. aeruginosa PAO1 altered the membrane components, enhanced membrane permeability, and disturbed energy metabolism, amino acid metabolism, and nucleotide metabolism, and would ultimately attenuate the pathogenicity of P. aeruginosa PAO1. Hordenine may have promising potential for controlling nosocomial pathogens.


Assuntos
Antibacterianos/metabolismo , Metaboloma , Pseudomonas aeruginosa/química , Pseudomonas aeruginosa/efeitos dos fármacos , Percepção de Quorum/efeitos dos fármacos , Tiramina/análogos & derivados , Regulação Bacteriana da Expressão Gênica/efeitos dos fármacos , Espectroscopia de Ressonância Magnética , Metabolômica , Tiramina/metabolismo
15.
Plant J ; 100(1): 20-37, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31124249

RESUMO

Enzyme promiscuity, a common property of many uridine diphosphate sugar-dependent glycosyltransferases (UGTs) that convert small molecules, significantly hinders the identification of natural substrates and therefore the characterization of the physiological role of enzymes. In this paper we present a simple but effective strategy to identify endogenous substrates of plant UGTs using LC-MS-guided targeted glycoside analysis of transgenic plants. We successfully identified natural substrates of two promiscuous Nicotiana benthamiana UGTs (NbUGT73A24 and NbUGT73A25), orthologues of pathogen-induced tobacco UGT (TOGT) from Nicotiana tabacum, which is involved in the hypersensitive reaction. While in N. tabacum, TOGT glucosylated scopoletin after treatment with salicylate, fungal elicitors and the tobacco mosaic virus, NbUGT73A24 and NbUGT73A25 produced glucosides of phytoalexin N-feruloyl tyramine, which may strengthen cell walls to prevent the intrusion of pathogens, and flavonols after agroinfiltration of the corresponding genes in N. benthamiana. Enzymatic glucosylation of fractions of a physiological aglycone library confirmed the biological substrates of UGTs. In addition, overexpression of both genes in N. benthamiana produced clear lesions on the leaves and led to a significantly reduced content of pathogen-induced plant metabolites such as phenylalanine and tryptophan. Our results revealed some additional biological functions of TOGT enzymes and indicated a multifunctional role of UGTs in plant resistance.


Assuntos
Ácidos Cumáricos/metabolismo , Glucose/metabolismo , Glicosiltransferases/genética , Doenças das Plantas/genética , Proteínas de Plantas/genética , Sesquiterpenos/metabolismo , Tabaco/genética , Tiramina/análogos & derivados , Perfilação da Expressão Gênica/métodos , Regulação da Expressão Gênica de Plantas , Glicosídeos/metabolismo , Glicosilação , Glicosiltransferases/metabolismo , Doenças das Plantas/virologia , Folhas de Planta/genética , Folhas de Planta/metabolismo , Folhas de Planta/virologia , Proteínas de Plantas/metabolismo , Especificidade por Substrato , Tabaco/metabolismo , Tabaco/virologia , Vírus do Mosaico do Tabaco/fisiologia , Tiramina/metabolismo
16.
Food Res Int ; 119: 110-118, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30884638

RESUMO

A traditional Thai fermented pork, nham, is a product popularly consumed in Thailand. Fermentation of the protein-rich product by uncontrolled bacterial community can result in high amounts of hazardous biogenic amines (BA). This study aimed to unveil dynamics of microbial community and its relation to BA accumulation in nham. Three batches of nham were analyzed for pH, lactic acid bacteria population, concentrations of organic acids and BA. Bacterial communities were analyzed by pyrosequencing of 16S rRNA gene amplicons. In all batches, pH dropped to the quality standard of nham (≤4.6) within 3-5 days by production of lactic acid and acetic acid. Initial BA levels varied batch-by-batch and increased with fermentation time. In the highest quality batch, levels of histamine, tyramine, and total BA were within the recommended safety limits (200, 100 and 1000 mg/kg, respectively) throughout the 10-days study. However, in other batches, unsafe levels of tyramine and total BA were found after 5 days of fermentation. The results indicated that over-fermentation and inferior conditions of ingredients increased risk due to high levels of BA. Lactobacillus, Lactococcus, Pediococcus and Weissella were prevalent and comprised >90% of total bacteria during fermentation. Weissella was predominant in the batch with low BA while Lactobacillus and Pediococcus were predominant in the higher BA batches. A negative correlation between Weissella dominance and total BA was observed (r = -0.90, p = .003). A 10% increase in dominance of Weissella was associated with 75-170 mg/kg decrease in total BA. W. hellenica was the species prevalent only in low BA batch. Therefore, W. hellenica isolates were suggested as subjects for future study to develop efficient starter culture securing safety of nham.


Assuntos
Bactérias/classificação , Aminas Biogênicas/biossíntese , Alimentos e Bebidas Fermentados/microbiologia , Produtos da Carne/microbiologia , Microbiota , Carne Vermelha/microbiologia , Animais , Bactérias/genética , Bactérias/isolamento & purificação , Técnicas de Cultura Celular por Lotes , Reatores Biológicos , Fermentação , Microbiologia de Alimentos , Inocuidade dos Alimentos , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Concentração de Íons de Hidrogênio , Metagenoma , Microbiota/genética , RNA Ribossômico 16S/genética , Suínos , Tailândia , Tiramina/metabolismo , Weissella/isolamento & purificação , Weissella/metabolismo
17.
Proc Natl Acad Sci U S A ; 116(9): 3805-3810, 2019 02 26.
Artigo em Inglês | MEDLINE | ID: mdl-30808766

RESUMO

Adrenergic signaling profoundly modulates animal behavior. For example, the invertebrate counterpart of norepinephrine, octopamine, and its biological precursor and functional antagonist, tyramine, adjust motor behavior to different nutritional states. In Drosophila larvae, food deprivation increases locomotor speed via octopamine-mediated structural plasticity of neuromuscular synapses, whereas tyramine reduces locomotor speed, but the underlying cellular and molecular mechanisms remain unknown. We show that tyramine is released into the CNS to reduce motoneuron intrinsic excitability and responses to excitatory cholinergic input, both by tyraminehonoka receptor activation and by downstream decrease of L-type calcium current. This central effect of tyramine on motoneurons is required for the adaptive reduction of locomotor activity after feeding. Similarly, peripheral octopamine action on motoneurons has been reported to be required for increasing locomotion upon starvation. We further show that the level of tyramine-ß-hydroxylase (TBH), the enzyme that converts tyramine into octopamine in aminergic neurons, is increased by food deprivation, thus selecting between antagonistic amine actions on motoneurons. Therefore, octopamine and tyramine provide global but distinctly different mechanisms to regulate motoneuron excitability and behavioral plasticity, and their antagonistic actions are balanced within a dynamic range by nutritional effects on TBH.


Assuntos
Oxigenases de Função Mista/genética , Neurônios Motores/metabolismo , Octopamina/genética , Receptores de Amina Biogênica/genética , Tiramina/metabolismo , Animais , Comportamento Animal/fisiologia , Canais de Cálcio Tipo L/genética , Canais de Cálcio Tipo L/metabolismo , Drosophila melanogaster/genética , Drosophila melanogaster/metabolismo , Drosophila melanogaster/fisiologia , Privação de Alimentos/fisiologia , Larva/metabolismo , Larva/fisiologia , Locomoção/genética , Locomoção/fisiologia , Oxigenases de Função Mista/metabolismo , Neurônios Motores/fisiologia , Estado Nutricional/genética , Estado Nutricional/fisiologia , Octopamina/metabolismo , Receptores de Amina Biogênica/metabolismo , Sinapses/metabolismo , Sinapses/fisiologia
19.
Food Chem ; 276: 745-753, 2019 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-30409657

RESUMO

The phenethylamine alkaloid hordenine, present in germinated barley, was identified recently as a functionally selective dopamine D2 receptor agonist contributing potentially to the rewarding effects of drinking beer. Here, it was shown that the hordenine precursor N-methyltyramine binds with a similar affinity to the dopamine D2 receptor as hordenine (Ki 31.3 µM) showing also selectivity towards the G protein-mediated pathway over the ß-arrestin pathway. Using a newly developed UHPLC-ESI-MS/MS method to monitor beer production, we demonstrated that hordenine and N-methyltyramine were released continuously from barley malt during mashing and were stable during fermentation and conditioning. The amounts released from different base malt types were in a similar range but tended to be higher from caramel malts. Hordenine and N-methyltyramine concentrations in 24 types of beer varied between 1.05-6.32 and 0.59-4.61 mg/L, respectively. Thus, the human uptake of the alkaloids during beer consumption is in the low milligram range.


Assuntos
Cerveja/análise , Agonistas de Dopamina/análise , Tiramina/análogos & derivados , Animais , Células CHO , Cromatografia Líquida de Alta Pressão , Cricetulus , Agonistas de Dopamina/metabolismo , Fermentação , Análise de Alimentos/métodos , Hordeum/metabolismo , Humanos , Ensaio Radioligante , Receptores de Dopamina D2/agonistas , Receptores de Dopamina D2/genética , Receptores de Dopamina D2/metabolismo , Espectrometria de Massas por Ionização por Electrospray , Espectrometria de Massas em Tandem , Tiramina/análise , Tiramina/metabolismo
20.
J Cell Biochem ; 120(4): 4804-4812, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30390333

RESUMO

Approximately 29 000 men die of prostate cancer (PCa) each year in the United States, and 90% to 100% of them are due to incurable bone metastasis. It is difficult to determine (1) when PCa disseminates in the natural history of the disease; (2) where cancer cell disseminates before becoming overt metastatic lesions; and (3) which tumors are aggressive and which are indolent. Tumor tissue and liquid (blood and bone marrow) biopsies provide important information to answer these questions, but significant limitations exist for immunostaining strategies that assess protein expression in these tissues. Classic immunohistochemistry (IHC) assays can typically assess expression of one or two proteins per tissue section. We have developed a novel immunofluorescence staining protocol to detect a panel of seven proteins on PCa tissue from primary tumor biopsies and metastatic lesion autopsy tissue, as well as cancer cells from liquid biopsies. We used a tyramide-based system to amplify the true signal and optimized the protocol to reduce background signal, thereby boosting the signal-to-noise ratio. Any protein-specific antibody in this protocol can be exchanged for a different validated antibody. This protocol therefore, represents a highly informative and flexible assay that can be used to provide important information about cancer tissue for the purpose of improving detection, diagnosis, and treatment.


Assuntos
Biotina/análogos & derivados , Neoplasias Ósseas/diagnóstico , Imunofluorescência/métodos , Neoplasias da Próstata/diagnóstico , Coloração e Rotulagem/métodos , Tiramina/análogos & derivados , Animais , Antígenos de Superfície/metabolismo , Biotina/metabolismo , Neoplasias Ósseas/metabolismo , Neoplasias Ósseas/secundário , Linhagem Celular Tumoral , Corantes Fluorescentes/metabolismo , Glutamato Carboxipeptidase II/metabolismo , Xenoenxertos , Humanos , Indóis/metabolismo , Calicreínas/metabolismo , Masculino , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Fosfoproteínas/metabolismo , Antígeno Prostático Específico/metabolismo , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Proteínas de Ligação a RNA/metabolismo , Racemases e Epimerases/metabolismo , Receptores Androgênicos/metabolismo , Tiramina/metabolismo
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