RESUMO
Fermented seasonings have pleasant flavors that stimulate our appetite. Their flavoring properties change depending on factors such as their materials and fermented conditions. Therefore, a comparative analysis of their flavor is important when evaluating their quality. However, seasonings contain high levels of various matrices such as sugars, proteins, lipids, and ethanol, making it difficult to extract aroma compounds efficiently from them. In this study, we verified a high-efficient and high-throughput volatile flavor analysis of fermented seasonings by solvent-assisted stir bar solid extraction (SA-SBSE) with reverse extraction. We applied SA-SBSE to Japanese fermented seasonings, soy sauce, miso (fermented beans), and mirin (sweet rice wine) and compared their profiles with those from other common extraction methods, headspace gas-solid-phase microextraction (HS-SPME), liquid extraction with solvent-assisted flavor evaporation (LE-SAFE), and conventional SBSE (C-SBSE). The aroma properties and profiles of extracts from SA-SBSE were close to those of the original sample, being similar to that of LE-SAFE. In addition, potent aroma compounds in each sample were extracted by SA-SBSE and LE-SAFE, which were far superior to those by C-SBSE. For quantification, SA-SBSE extracts showed a good standard curve by the standard addition method. We could quantify maltol, one of the most common potent aroma compounds in all samples, for various commercial samples by such high-throughput analysis.
Assuntos
Etanol , Microextração em Fase Sólida , Cromatografia Gasosa-Espectrometria de Massas/métodos , Japão , Microextração em Fase Sólida/métodos , Solventes , Etanol/análise , Compostos Orgânicos , Odorantes/análiseRESUMO
Alternative splicing in the 3'UTR of mammalian genes plays a crucial role in diverse biological processes, including cell differentiation and development. SAM68 is a key splicing regulator that controls the diversity of 3'UTR isoforms through alternative last exon (ALE) selection. However, the tissue/cell type-specific mechanisms underlying the splicing control at the 3' end and its functional significance remain unclear. Here, we show that SAM68 regulates ALE splicing in a dose-dependent manner and the neuronal splicing is differentially regulated depending on the characteristics of the target transcript. Specifically, we found that SAM68 regulates interleukin-1 receptor-associated protein splicing through the interaction with U1 small nuclear ribonucleoprotein. In contrast, the ALE splicing of protocadherin-15 (Pcdh15), a gene implicated in several neuropsychiatric disorders, is independent of U1 small nuclear ribonucleoprotein but modulated by the calcium/calmodulin-dependent protein kinase signaling pathway. We found that the aberrant ALE selection of Pcdh15 led to a conversion from a membrane-bound to a soluble isoform and consequently disrupted its localization into excitatory and inhibitory synapses. Notably, the neuronal expression of the soluble form of PCDH15 preferentially affected the number of inhibitory synapses. Moreover, the soluble form of PCDH15 interacted physically with α-neurexins and further disrupted neuroligin-2-induced inhibitory synapses in artificial synapse formation assays. Our findings provide novel insights into the role of neuron-specific alternative 3'UTR isoform selections in synapse development.
RESUMO
Skeletal muscle atrophy caused by various conditions including aging, nerve damage, and steroid administration, is a serious health problem worldwide. We recently reported that neuron-derived neurotrophic factor (NDNF) functions as a muscle-derived secreted factor, also known as myokine, which exerts protective actions on endothelial cell and cardiomyocyte function. Here, we investigated whether NDNF regulates skeletal muscle atrophy induced by steroid administration and sciatic denervation. NDNF-knockout (KO) mice and age-matched wild-type (WT) mice were subjected to continuous dexamethasone (DEX) treatment or sciatic denervation. NDNF-KO mice exhibited decreased gastrocnemius muscle weight and reduced cross sectional area of myocyte fiber after DEX treatment or sciatic denervation compared with WT mice. Administration of an adenoviral vector expressing NDNF (Ad-NDNF) or recombinant NDNF protein to gastrocnemius muscle of WT mice increased gastrocnemius muscle weight after DEX treatment. NDNF-KO mice showed increased expression of ubiquitin E3-ligases, including atrogin-1 and MuRF-1, in gastrocnemius muscle after DEX treatment, whereas Ad-NDNF reduced expression of atrogin-1 and MuRF-1 in gastrocnemius muscle of WT mice after DEX treatment. Pretreatment of cultured C2C12 myocytes with NDNF protein reversed reduced myotube diameter and increased expression of atrogin-1 and MuRF-1 after DEX stimulation. Treatment of C2C12 myocytes increased Akt phosphorylation. Pretreatment of C2C12 myotubes with the PI3-kinase/Akt inhibitor reversed NDNF-induced increase in myotube fiber diameter after DEX treatment. In conclusion, our findings indicated that NDNF prevents skeletal muscle atrophy in vivo and in vitro through reduction of ubiquitin E3-ligases expression, suggesting that NDNF could be a novel therapeutic target of muscle atrophy.
Assuntos
Dexametasona/toxicidade , Músculo Esquelético/efeitos dos fármacos , Atrofia Muscular/prevenção & controle , Fatores de Crescimento Neural/farmacologia , Neurônios/efeitos dos fármacos , Substâncias Protetoras/metabolismo , Animais , Anti-Inflamatórios/toxicidade , Feminino , Regulação da Expressão Gênica , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Músculo Esquelético/metabolismo , Músculo Esquelético/patologia , Atrofia Muscular/induzido quimicamente , Atrofia Muscular/metabolismo , Atrofia Muscular/patologia , Neurônios/metabolismo , Neurônios/patologia , FosforilaçãoRESUMO
The mammalian brain contains multiple types of neuronal cells with complex assemblies and distinct structural and functional properties encoded by divergent gene programs. There is increasing evidence that alternative splicing (AS) plays fundamental roles in transcriptomic diversity and specifying synaptic properties of each neuronal cell type. However, the mechanisms underlying AS regulation and whether it controls synapse formation across GABAergic interneurons have not been fully elucidated. Here we show the differential expression levels of Sam68-like molecule 2 (SLM2), a major splicing regulator of neurexin (NRX), in GABAergic neuronal subtypes and its contribution to GABAergic synapse specification. Cortical SLM2 is strongly expressed not only in excitatory neurons but also in a subpopulation of GABAergic interneurons, especially in VIP-positive neurons that are originated from late-born caudal ganglionic eminence (GE)- derived cells. Using artificial synapse formation assay, we found that GE containing cortices form a strong synapse with LRRTM2, a trans-synaptic receptor of the alternatively spliced segment 4 (AS4)(-) of NRX. SLM2 knock-down reduced the NRX AS4(-) isoform expression and hence weaken LRRTM2-induced synapse formation. The addition of NRX AS4(-) was sufficient to rescue the synaptic formation by LRRTM2 in SLM2 knock-down neurons. Thus, our findings suggest a novel function of SLM2 in modifying network formation of a specific population of GABAergic interneurons and contribute to a better understanding of the roles AS plays in regulating synapse specificity and neuronal molecular diversity.
Assuntos
Processamento Alternativo , Neurônios GABAérgicos , Animais , Interneurônios , Mamíferos , Neurogênese , Sinapses/fisiologiaRESUMO
Green leaf volatiles (GLVs), the common constituents of herbivore-infested plant volatiles (HIPVs), play an important role in plant defense and function as chemical cues to communicate with other individuals in nature. Reportedly, in addition to endogenous GLVs, the absorbance of airborne GLVs emitted by infested neighboring plants also play a major role in plant defense. For example, the exclusive accumulation of (Z)-3-hexenyl vicianoside in the HIPV-exposed tomato plants occurs by the glycosylation of airborne (Z)-3-hexenol (Z3HOL); however, it is unclear how plants process the other absorbed GLVs. This study demonstrates that tomato plants dominantly accumulated GLV-glycosides after exposure to green leaf alcohols [Z3HOL, (E)-2-hexenol, and n-hexanol] using non-targeted LC-MS analysis. Three types of green leaf alcohols were independently glycosylated without isomerization or saturation/desaturation. Airborne green leaf aldehydes and esters were also glycosylated, probably through converting aldehydes and esters into alcohols. Further, we validated these findings in Arabidopsis mutants- (Z)-3-hexenal (Z3HAL) reductase (chr) mutant that inhibits the conversion of Z3HAL to Z3HOL and the acetyl-CoA:(Z)-3-hexen-1-ol acetyltransferase (chat) mutant that impairs the conversion of Z3HOL to (Z)-3-hexenyl acetate. Exposure of the chr and chat mutants to Z3HAL accumulated lower and higher amounts of glycosides than their corresponding wild types (Col-0 and Ler), respectively. These findings suggest that plants process the exogenous GLVs by the reductase(s) and the esterase(s), and a part of the processed GLVs contribute to glycoside accumulation. Overall, the study provides insights into the understanding of the communication of the plants within their ecosystem, which could help develop strategies to protect the crops and maintain a balanced ecosystem.
RESUMO
Tomato (Solanum lycopersicum) contains α-tomatine, a steroidal glycoalkaloid that contributes to the plant defense against pathogens and herbivores through its bitter taste and toxicity. It accumulates at high levels in all the plant tissues, especially in leaves and immature green fruits, whereas it decreases during fruit ripening through metabolic conversion to the nontoxic esculeoside A, which accumulates in the mature red fruit. This study aimed to identify the gene encoding a C-27 hydroxylase that is a key enzyme in the metabolic conversion of α-tomatine to esculeoside A. The E8 gene, encoding a 2-oxoglutalate-dependent dioxygenase, is well known as an inducible gene in response to ethylene during fruit ripening. The recombinant E8 was found to catalyze the C-27 hydroxylation of lycoperoside C to produce prosapogenin A and is designated as Sl27DOX. The ripe fruit of E8/Sl27DOX-silenced transgenic tomato plants accumulated lycoperoside C and exhibited decreased esculeoside A levels compared with the wild-type (WT) plants. Furthermore, E8/Sl27DOX deletion in tomato accessions resulted in higher lycoperoside C levels in ripe fruits than in WT plants. Thus, E8/Sl27DOX functions as a C-27 hydroxylase of lycoperoside C in the metabolic detoxification of α-tomatine during tomato fruit ripening, and the efficient detoxification by E8/27DOX may provide an advantage in the domestication of cultivated tomatoes.
Assuntos
Frutas/metabolismo , Oxigenases de Função Mista/metabolismo , Proteínas de Plantas/metabolismo , Solanum lycopersicum/metabolismo , Tomatina/análogos & derivados , Frutas/crescimento & desenvolvimento , Solanum lycopersicum/genética , Solanum lycopersicum/crescimento & desenvolvimento , Oxigenases de Função Mista/genética , Filogenia , Proteínas de Plantas/genética , Plantas Geneticamente Modificadas , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Saponinas/metabolismo , Especificidade por Substrato , Tomatina/metabolismoRESUMO
Tomato plants (Solanum lycopersicum) contain steroidal glycoalkaloid α-tomatine, which functions as a chemical barrier to pathogens and predators. α-Tomatine accumulates in all tissues and at particularly high levels in leaves and immature green fruits. The compound is toxic and causes a bitter taste, but its presence decreases through metabolic conversion to nontoxic esculeoside A during fruit ripening. This study identifies the gene encoding a 23-hydroxylase of α-tomatine, which is a key to this process. Some 2-oxoglutarate-dependent dioxygenases were selected as candidates for the metabolic enzyme, and Solyc02g062460, designated Sl23DOX, was found to encode α-tomatine 23-hydroxylase. Biochemical analysis of the recombinant Sl23DOX protein demonstrated that it catalyzes the 23-hydroxylation of α-tomatine and the product spontaneously isomerizes to neorickiioside B, which is an intermediate in α-tomatine metabolism that appears during ripening. Leaves of transgenic tomato plants overexpressing Sl23DOX accumulated not only neorickiioside B but also another intermediate, lycoperoside C (23-O-acetylated neorickiioside B). Furthermore, the ripe fruits of Sl23DOX-silenced transgenic tomato plants contained lower levels of esculeoside A but substantially accumulated α-tomatine. Thus, Sl23DOX functions as α-tomatine 23-hydroxylase during the metabolic processing of toxic α-tomatine in tomato fruit ripening and is a key enzyme in the domestication of cultivated tomatoes.
Assuntos
Oxigenases de Função Mista/metabolismo , Solanum lycopersicum/enzimologia , Solanum lycopersicum/metabolismo , Paladar , Tomatina/análogos & derivados , Tomatina/metabolismo , Frutas/metabolismo , Regulação da Expressão Gênica de Plantas , Inativação Metabólica , Solanum lycopersicum/genética , Oxigenases de Função Mista/genética , Folhas de Planta/metabolismo , Plantas Geneticamente Modificadas/metabolismo , Proteínas RecombinantesRESUMO
Neuronal alternative splicing is a core mechanism for functional diversification. We previously found that STAR family proteins (SAM68, SLM1, SLM2) regulate spatiotemporal alternative splicing in the nervous system. However, the whole aspect of alternative splicing programs by STARs remains unclear. Here, we performed a transcriptomic analysis using SAM68 knockout and SAM68/SLM1 double-knockout midbrains. We revealed different alternative splicing activity between SAM68 and SLM1; SAM68 preferentially targets alternative 3' UTR exons. SAM68 knockout causes a long-to-short isoform switch of a number of neuronal targets through the alteration in alternative last exon (ALE) selection or alternative polyadenylation. The altered ALE usage of a novel target, interleukin 1 receptor accessory protein (Il1rap), results in remarkable conversion from a membrane-bound type to a secreted type in Sam68KO brains. Proper ALE selection is necessary for IL1RAP neuronal function. Thus the SAM68-specific splicing program provides a mechanism for neuronal selection of alternative 3' UTR isoforms.
RESUMO
Alternative splicing is crucial for molecular diversification, which greatly contributes to the complexity and specificity of neural functions in the central nervous system (CNS). Neurofascin (NF) is a polymorphic cell surface protein that has a number of splicing isoforms. As the alternative splicing of the neurofascin gene (Nfasc) is developmentally regulated, NF isoforms have distinct functions in immature and mature brains. However, the molecular mechanisms underlying the alternative splicing of Nfasc in neurons are not yet understood. Here, we demonstrate that, alongside developmental regulation, Nfasc alternative splicing is spatially controlled in the mouse brain. We then identified distinct Nfasc splicing patterns at the cell-type level in the cerebellum, with Nfasc186 being expressed in Purkinje cells and absent from granule cells (GCs). Furthermore, we show that high K+-induced depolarization triggers a shift in splicing from Nfasc140 to Nfasc186 in cerebellar GCs. Finally, we identified a neural RNA-binding protein, Rbfox, as a key player in neural NF isoform selection, specifically controlling splicing at exons 26-29. Together, our results show that Nfasc alternative splicing is spatio-temporally and dynamically regulated in cerebellar neurons. Our findings provide profound insight into the mechanisms underlying the functional diversity of neuronal cell-adhesive proteins in the mammalian CNS.
Assuntos
Processamento Alternativo , Moléculas de Adesão Celular/genética , Cerebelo/metabolismo , Regulação da Expressão Gênica , Fatores de Crescimento Neural/genética , Neurônios/metabolismo , Animais , Biomarcadores , Células Cultivadas , Cerebelo/citologia , Vetores Genéticos/genética , Imuno-Histoquímica , Camundongos , Especificidade de Órgãos , Transdução GenéticaRESUMO
Neurexins (NRXs) and neuroligins (NLs) play important roles in synapse specification. The alternatively spliced segment 4 (AS4) of NRX genes (Nrxn) is a critical element in selective trans-synaptic interactions. However, the role of splicing of NRXs and NLs in synapse specification is not fully understood. To investigate the exact role of splice-dependent NRX-NL interaction in the specification of glutamatergic and gamma-aminobutyric acid (GABA)-ergic synapses in the cerebellum, we evaluated the synaptogenic receptor activity of NL1/2/3 isoforms in a neuron-fibroblast co-culture system, in which the Nrxn AS4 segments are manipulated using SLM2, a selective and dominant regulator of AS4 splicing. We show that ectopic SLM2 expression (SLM2 E/E) causes marked skipping of exon 20 of AS4 in cerebellar neuron culture. Whereas NLs can induce VAMP2+ presynaptic contacts from mainly glutamatergic neurons in both uninfected (control) and SLM2 E/E co-cultures, they induce VGAT+ GABAergic contacts in the control culture, but not properly in the SLM2 E/E culture. Furthermore, Nrxn3 is responsible for the NL-induced assembly of GABAergic synapses in co-culture. Importantly, lentivirus-based expression of Nrxn3 containing exon 20 restores the reduced NL-induced GABAergic contacts in the SLM2 E/E co-culture. Therefore, our findings may provide further insights into NRX-NL mediated synapse specification.
Assuntos
Processamento Alternativo , Moléculas de Adesão Celular Neuronais/metabolismo , Cerebelo/citologia , Proteínas de Membrana/metabolismo , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Proteínas de Ligação a RNA/metabolismo , Animais , Proteínas de Ligação ao Cálcio , Moléculas de Adesão Celular Neuronais/genética , Células Cultivadas , Cerebelo/metabolismo , Técnicas de Cocultura , Células HEK293 , Humanos , Camundongos Endogâmicos ICR , Moléculas de Adesão de Célula Nervosa/genética , Moléculas de Adesão de Célula Nervosa/metabolismo , Neurônios/citologia , Neurônios/metabolismo , Proteínas de Ligação a RNA/genética , Sinapses/metabolismo , Ácido gama-Aminobutírico/metabolismoRESUMO
Volatile terpenes are ones of the characteristic aromas of Japanese pepper (Zanthoxylum piperitum). It has been hypothesized that the specialized epithelial cells surrounding the secretory cavities of Japanese pepper fruits and leaves are responsible for the synthesis of monoterpenes and sesquiterpenes, which are generally produced by terpene synthases (TPSs); however, direct evidence for the formation of terpenes in Japanese pepper remains elusive. Here we report that monoterpenes and sesquiterpenes accumulate inside the secretory cavities of Japanese pepper leaves, but not in other parts of leaf tissues that do not include secretory cavities. We have obtained cDNAs for ZpTPS1 and ZpTPS2, which are responsible for biosynthesis of the sesquiterpenes ß-caryophyllene and germacrene D, respectively, in Japanese pepper. In addition, we also identified a cDNA for the monoterpene synthase ZpTPS3. Expression of ZpTPS3 in Escherichia coli in addition to Agrobacterium-mediated transient ZpTPS3 expression in Nicotiana benthamiana demonstrated the catalytic activity of ZpTPS3 to form ß-phellandrene as the major product. In situ hybridization in Japanese pepper leaf tissue revealed that ZpTPS3 transcript specifically accumulated in the epithelial cells surrounding secretory cavities. Expression of ZpTPS3 in epithelial cells was only detectable during early stages of cavity development, whereas the formation of volatile terpenes occurred at a constant rate throughout the expansion of secretory cavities. Our studies have improved the understanding of the currently uncharacterized processes controlling volatile terpene biosynthesis in Japanese pepper leaves.
RESUMO
Various commercial tomato juices with different flavors are available at markets worldwide. To clarify the marker compounds related to the flavor characteristics of tomato juice, we analyzed 15 pure commercial tomato juices by a combination of volatile profiling and sensory evaluation. The correlations among volatiles and the relationship between volatiles and sensory descriptors were elucidated by multivariate analyses. Consequently, the tomato juices made from fresh market tomatoes (including the popular Japanese tomato variety "Momotaro") were clearly separated from other juices made from processing tomatoes, by both the volatile composition and sensory profiles. cis-3-Hexenol, hexanal, and apocarotenoids negatively contributed to the juices from fresh market tomatoes, whereas Strecker aldehydes and furfural showed positive contributions to the juices. Accordingly, the sensory characteristics of juices from fresh market tomatoes were related to cooked and fruity flavors but not to green or fresh notes.
Assuntos
Manipulação de Alimentos , Sucos de Frutas e Vegetais/análise , Frutas/química , Solanum lycopersicum/química , Paladar/efeitos dos fármacos , Compostos Orgânicos Voláteis/análise , Compostos Orgânicos Voláteis/farmacologia , Humanos , Compostos Orgânicos Voláteis/químicaRESUMO
Successful feeding of the substrate geranylpyrophosphate (GPP) to monoterpene synthase is critical to the efficient microbial production of monoterpenes. Overexpression of GPP synthases, metabolic channeling from GPP synthase to terpene synthases, and down-tuning of endogenous competitors have been successfully used to increase the production of monoterpene. Nevertheless, the production of monoterpenes has remained considerably lower than that of hemi-/sesqui-terpenoids. We tested whether it is effective to improve the cellular activity of monoterpene synthases. To this end, we developed a high-throughput screening system to monitor for elevated GPP consumption. Through a single round of mutagenesis and screening, we isolated a pinene synthase variant that outperformed the wild-type (parent) enzyme in multiple contexts in Escherichia coli and cyanobacteria. The purified variant exhibited drastically altered metal dependency, enabling to keep the activity in the cytosol that is manganese-deficient. Coexpression of this variant with mevalonate pathway enzymes, isopentenylpyrophosphate isomerase, and GPP synthase yielded 140 mg/L pinene in a flask culture.
Assuntos
Alquil e Aril Transferases/metabolismo , Cianobactérias/metabolismo , Escherichia coli/metabolismo , Isomerases de Ligação Dupla Carbono-Carbono/metabolismo , Citosol/metabolismo , Hemiterpenos , Monoterpenos/metabolismo , Mutagênese/fisiologia , Fosfatos de Poli-Isoprenil/metabolismoRESUMO
Autism spectrum disorders (ASDs) are a heterogeneous group of neurodevelopmental disorders characterized by impairments in social interactions and stereotyped behaviors. Valproic acid (VPA) is frequently used to treat epilepsy and bipolar disorders. When taken during pregnancy, VPA increases the risk of the unborn child to develop an ASD. In rodents, in utero VPA exposure can precipitate behavioral phenotypes related to ASD in the offspring. Therefore, such rodent models may allow for identification of synaptic pathophysiology underlying ASD risk. Here, we systematically probed alterations in synaptic proteins that might contribute to autism-related behavior in the offspring of in utero VPA-exposed mice. Moreover, we tested whether direct VPA exposure of cultured neocortical neurons may recapitulate the molecular alterations seen in vivo. VPA-exposed neurons in culture exhibit a significant increase in the number of glutamatergic synapses accompanied by a significant decrease in the number of GABAergic synapses. This shift in excitatory/inhibitory balance results in substantially increased spontaneous activity in neuronal networks arising from VPA-exposed neurons. Pharmacological experiments demonstrate that the alterations in GABAergic and glutamatergic synaptic proteins and structures are largely caused by inhibition of histone deacetylases. Therefore, our study highlights an epigenetic mechanism underlying the synaptic pathophysiology in this ASD model.
Assuntos
Anticonvulsivantes/farmacologia , Neocórtex/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Efeitos Tardios da Exposição Pré-Natal , Sinapses/efeitos dos fármacos , Ácido Valproico/farmacologia , Animais , Feminino , Camundongos , Camundongos Endogâmicos ICR , Neocórtex/citologia , Proteínas do Tecido Nervoso/metabolismo , Gravidez , Sinapses/metabolismoRESUMO
Alternative pre-mRNA splicing is a fundamental mechanism that generates molecular diversity from a single gene. In the central nervous system (CNS), key neural developmental steps are thought to be controlled by alternative splicing decisions, including the molecular diversity underlying synaptic wiring, plasticity, and remodeling. Significant progress has been made in understanding the molecular mechanisms and functions of alternative pre-mRNA splicing in neurons through studies in invertebrate systems; however, recent studies have begun to uncover the potential role of neuronal alternative splicing in the mammalian CNS. This article provides an overview of recent findings regarding the regulation and function of neuronal alternative splicing. In particular, we focus on the spatio-temporal regulation of neurexin, a synaptic adhesion molecule, by neuronal cell type-specific factors and neuronal activity, which are thought to be especially important for characterizing neural development and function within the mammalian CNS. Notably, there is increasing evidence that implicates the dysregulation of neuronal splicing events in several neurological disorders. Therefore, understanding the detailed mechanisms of neuronal alternative splicing in the mammalian CNS may provide plausible treatment strategies for these diseases.
Assuntos
Processamento Alternativo , Encéfalo/metabolismo , RNA/genética , Animais , Encéfalo/embriologia , Encéfalo/crescimento & desenvolvimento , Cálcio/metabolismo , Moléculas de Adesão Celular Neuronais/genética , Moléculas de Adesão Celular Neuronais/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Humanos , Plasticidade Neuronal , Neurônios/citologia , Neurônios/metabolismo , RNA/metabolismo , Transdução de SinaisRESUMO
Homeostatic synaptic plasticity, or synaptic scaling, is a mechanism that tunes neuronal transmission to compensate for prolonged, excessive changes in neuronal activity. Both excitatory and inhibitory neurons undergo homeostatic changes based on synaptic transmission strength, which could effectively contribute to a fine-tuning of circuit activity. However, gene regulation that underlies homeostatic synaptic plasticity in GABAergic (GABA, gamma aminobutyric) neurons is still poorly understood. The present study demonstrated activity-dependent dynamic scaling in which NMDA-R (N-methyl-D-aspartic acid receptor) activity regulated the expression of GABA synthetic enzymes: glutamic acid decarboxylase 65 and 67 (GAD65 and GAD67). Results revealed that activity-regulated BDNF (brain-derived neurotrophic factor) release is necessary, but not sufficient, for activity-dependent up-scaling of these GAD isoforms. Bidirectional forms of activity-dependent GAD expression require both BDNF-dependent and BDNF-independent pathways, both triggered by NMDA-R activity. Additional results indicated that these two GAD genes differ in their responsiveness to chronic changes in neuronal activity, which could be partially caused by differential dependence on BDNF. In parallel to activity-dependent bidirectional scaling in GAD expression, the present study further observed that a chronic change in neuronal activity leads to an alteration in neurotransmitter release from GABAergic neurons in a homeostatic, bidirectional fashion. Therefore, the differential expression of GAD65 and 67 during prolonged changes in neuronal activity may be implicated in some aspects of bidirectional homeostatic plasticity within mature GABAergic presynapses.
Assuntos
Fator Neurotrófico Derivado do Encéfalo/fisiologia , Regulação da Expressão Gênica , Glutamato Descarboxilase/biossíntese , Transdução de Sinais/fisiologia , Animais , Benzilaminas/farmacologia , Bicuculina/farmacologia , Butadienos/farmacologia , Sinalização do Cálcio/efeitos dos fármacos , Carbazóis/farmacologia , Células Cultivadas , Córtex Cerebral/citologia , Indução Enzimática/efeitos dos fármacos , Neurônios GABAérgicos/efeitos dos fármacos , Neurônios GABAérgicos/enzimologia , Neurônios GABAérgicos/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Glutamato Descarboxilase/genética , Homeostase , Alcaloides Indólicos/farmacologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos ICR , Nitrilas/farmacologia , Isoformas de Proteínas/biossíntese , Isoformas de Proteínas/genética , Inibidores de Proteínas Quinases/farmacologia , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Receptor trkB/antagonistas & inibidores , Receptor trkB/fisiologia , Receptores de N-Metil-D-Aspartato/efeitos dos fármacos , Receptores de N-Metil-D-Aspartato/fisiologia , Transdução de Sinais/efeitos dos fármacos , Sulfonamidas/farmacologia , Ácido gama-Aminobutírico/metabolismoRESUMO
In many plants, biogenic volatile organic compounds (BVOCs) are produced as specialized metabolites that contribute to the characteristics of each plant. The varieties and composition of BVOCs are chemically diverse by plant species and the circumstances in which the plants grow, and also influenced by herbivory damage and pathogen infection. Plant-produced BVOCs are receptive to many organisms, from microorganisms to human, as both airborne attractants and repellants. In addition, it is known that some BVOCs act as signals to prime a plant for the defense response in plant-to-plant communications. The compositional profiles of BVOCs can, thus, have profound influences in the physiological and ecological aspects of living organisms. Apart from that, some of them are commercially valuable as aroma/flavor compounds for human. Metabolomic technologies have recently revealed new insights in biological systems through metabolic dynamics. Here, the recent advances in metabolomics technologies focusing on plant-produced BVOC analyses are overviewed. Their application markedly improves our knowledge of the role of BVOCs in chemosystematics, ecological influences, and aroma research, as well as being useful to prove the biosynthetic mechanisms of BVOCs.
RESUMO
A metabolome--the collection of comprehensive quantitative data on metabolites in an organism--has been increasingly utilized for applications such as data-intensive systems biology, disease diagnostics, biomarker discovery, and assessment of food quality. A considerable number of tools and databases have been developed to date for the analysis of data generated by various combinations of chromatography and mass spectrometry. We report here a web portal named KOMICS (The Kazusa Metabolomics Portal), where the tools and databases that we developed are available for free to academic users. KOMICS includes the tools and databases for preprocessing, mining, visualization, and publication of metabolomics data. Improvements in the annotation of unknown metabolites and dissemination of comprehensive metabolomic data are the primary aims behind the development of this portal. For this purpose, PowerGet and FragmentAlign include a manual curation function for the results of metabolite feature alignments. A metadata-specific wiki-based database, Metabolonote, functions as a hub of web resources related to the submitters' work. This feature is expected to increase citation of the submitters' work, thereby promoting data publication. As an example of the practical use of KOMICS, a workflow for a study on Jatropha curcas is presented. The tools and databases available at KOMICS should contribute to enhanced production, interpretation, and utilization of metabolomic Big Data.
Assuntos
Bases de Dados Factuais , Internet , Metaboloma , Metabolômica , Humanos , Software , Biologia de SistemasRESUMO
Plants receive volatile compounds emitted by neighboring plants that are infested by herbivores, and consequently the receiver plants begin to defend against forthcoming herbivory. However, to date, how plants receive volatiles and, consequently, how they fortify their defenses, is largely unknown. In this study, we found that undamaged tomato plants exposed to volatiles emitted by conspecifics infested with common cutworms (exposed plants) became more defensive against the larvae than those exposed to volatiles from uninfested conspecifics (control plants) in a constant airflow system under laboratory conditions. Comprehensive metabolite analyses showed that only the amount of (Z)-3-hexenylvicianoside (HexVic) was higher in exposed than control plants. This compound negatively affected the performance of common cutworms when added to an artificial diet. The aglycon of HexVic, (Z)-3-hexenol, was obtained from neighboring infested plants via the air. The amount of jasmonates (JAs) was not higher in exposed plants, and HexVic biosynthesis was independent of JA signaling. The use of (Z)-3-hexenol from neighboring damaged conspecifics for HexVic biosynthesis in exposed plants was also observed in an experimental field, indicating that (Z)-3-hexenol intake occurred even under fluctuating environmental conditions. Specific use of airborne (Z)-3-hexenol to form HexVic in undamaged tomato plants reveals a previously unidentified mechanism of plant defense.
Assuntos
Hexanóis/metabolismo , Odorantes , Solanum lycopersicum/metabolismo , Solanum lycopersicum/parasitologia , Spodoptera/crescimento & desenvolvimento , Animais , Ciclopentanos/metabolismo , Ciclopentanos/farmacologia , Glicosídeos/metabolismo , Herbivoria/fisiologia , Larva/fisiologia , Solanum lycopersicum/efeitos dos fármacos , Oxilipinas/metabolismo , Oxilipinas/farmacologia , Folhas de Planta/metabolismo , Compostos Orgânicos Voláteis/metabolismoRESUMO
The unique functional properties and molecular identity of neuronal cell populations rely on cell type-specific gene expression programs. Alternative splicing represents a powerful mechanism for expanding the capacity of genomes to generate molecular diversity. Neuronal cells exhibit particularly extensive alternative splicing regulation. We report a highly selective expression of the KH domain-containing splicing regulators SLM1 and SLM2 in the mouse brain. Conditional ablation of SLM1 resulted in a severe defect in the neuronal isoform content of the polymorphic synaptic receptors neurexin-1, -2, and -3. Thus, cell type-specific expression of SLM1 provides a mechanism for shaping the molecular repertoires of synaptic adhesion molecules in neuronal populations in vivo.