RESUMO
Classically, hormones elicit specific cellular responses by activating dedicated receptors. Nevertheless, the biosynthesis and turnover of many of these hormone molecules also produce chemically related metabolites. These molecules may also possess hormonal activities; therefore, one or more may contribute to the adaptive plasticity of signaling outcomes in host organisms. Here, we show that a catabolite of the plant hormone abscisic acid (ABA), namely phaseic acid (PA), likely emerged in seed plants as a signaling molecule that fine-tunes plant physiology, environmental adaptation, and development. This trait was facilitated by both the emergence-selection of a PA reductase that modulates PA concentrations and by the functional diversification of the ABA receptor family to perceive and respond to PA. Our results suggest that PA serves as a hormone in seed plants through activation of a subset of ABA receptors. This study demonstrates that the co-evolution of hormone metabolism and signaling networks can expand organismal resilience.
Assuntos
Ácido Abscísico/metabolismo , Arabidopsis/metabolismo , Reguladores de Crescimento de Plantas/metabolismo , Sesquiterpenos/metabolismo , Oxirredutases do Álcool/genética , Oxirredutases do Álcool/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , NADP/metabolismo , Transdução de SinaisRESUMO
Individuals can exhibit differences in metabolism that are caused by the interplay of genetic background, nutritional input, microbiota and other environmental factors1-4. It is difficult to connect differences in metabolism to genomic variation and derive underlying molecular mechanisms in humans, owing to differences in diet and lifestyle, among others. Here we use the nematode Caenorhabditis elegans as a model to study inter-individual variation in metabolism. By comparing three wild strains and the commonly used N2 laboratory strain, we find differences in the abundances of both known metabolites and those that have not to our knowledge been previously described. The latter metabolites include conjugates between 3-hydroxypropionate (3HP) and several amino acids (3HP-AAs), which are much higher in abundance in one of the wild strains. 3HP is an intermediate in the propionate shunt pathway, which is activated when flux through the canonical, vitamin-B12-dependent propionate breakdown pathway is perturbed5. We show that increased accumulation of 3HP-AAs is caused by genetic variation in HPHD-1, for which 3HP is a substrate. Our results suggest that the production of 3HP-AAs represents a 'shunt-within-a-shunt' pathway to accommodate a reduction-of-function allele in hphd-1. This study provides a step towards the development of metabolic network models that capture individual-specific differences of metabolism and more closely represent the diversity that is found in entire species.
Assuntos
Caenorhabditis elegans , Redes e Vias Metabólicas , Animais , Humanos , Oxirredutases do Álcool/genética , Oxirredutases do Álcool/metabolismo , Aminoácidos/metabolismo , Caenorhabditis elegans/classificação , Caenorhabditis elegans/enzimologia , Caenorhabditis elegans/genética , Caenorhabditis elegans/metabolismo , Ácido Láctico/análogos & derivados , Ácido Láctico/metabolismo , Redes e Vias Metabólicas/genética , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo , Modelos Animais , Propionatos/metabolismo , Vitamina B 12/metabolismoRESUMO
Retinoic acid (RA), a vitamin A metabolite, regulates transcriptional programs that drive protective or pathogenic immune responses in the intestine, in a manner dependent on RA concentration. Vitamin A is obtained from diet and is metabolized by intestinal epithelial cells (IECs), which operate in intimate association with microbes and immune cells. Here we found that commensal bacteria belonging to class Clostridia modulate RA concentration in the gut by suppressing the expression of retinol dehydrogenase 7 (Rdh7) in IECs. Rdh7 expression and associated RA amounts were lower in the intestinal tissue of conventional mice, as compared to germ-free mice. Deletion of Rdh7 in IECs diminished RA signaling in immune cells, reduced the IL-22-dependent antimicrobial response, and enhanced resistance to colonization by Salmonella Typhimurium. Our findings define a regulatory circuit wherein bacterial regulation of IEC-intrinsic RA synthesis protects microbial communities in the gut from excessive immune activity, achieving a balance that prevents colonization by enteric pathogens.
Assuntos
Disbiose/metabolismo , Células Epiteliais/metabolismo , Interleucinas/metabolismo , Mucosa Intestinal/metabolismo , Tretinoína/metabolismo , Oxirredutases do Álcool/genética , Oxirredutases do Álcool/metabolismo , Animais , Bactérias/classificação , Bactérias/genética , Disbiose/microbiologia , Células Epiteliais/microbiologia , Interações entre Hospedeiro e Microrganismos , Mucosa Intestinal/citologia , Mucosa Intestinal/microbiologia , Linfócitos/metabolismo , Linfócitos/microbiologia , Camundongos Endogâmicos C57BL , Camundongos Knockout , Microbiota/genética , Microbiota/fisiologia , RNA Ribossômico 16S/genética , Salmonella typhimurium/genética , Salmonella typhimurium/fisiologia , Simbiose , Interleucina 22RESUMO
Reactive carbonyl and oxygen species (RCS/ROS), often generated as metabolic byproducts, particularly under conditions of pathology, can cause direct damage to proteins, lipids, and nucleic acids. Glyoxal oxidases (Gloxs) oxidize aldehydes to carboxylic acids, generating hydrogen peroxide (H2O2). Although best characterized for their roles in lignin degradation, Glox in plant fungal pathogens are known to contribute to virulence, however, the mechanism underlying such effects are unclear. Here, we show that Glox in the insect pathogenic fungus, Metarhizium acridum, is highly expressed in mycelia and during formation of infection structures (appressoria), with the enzyme localizing to the cell membrane. MaGlox targeted gene disruption mutants showed RCS and ROS accumulation, resulting in cell toxicity, induction of apoptosis and increased autophagy, inhibiting normal fungal growth and development. The ability of the MaGlox mutant to scavenge RCS was significantly reduced, and the mutant exhibited increased susceptibility to aldehydes, oxidative and cell wall perturbing agents but not toward osmotic stress, with altered cell wall contents. The ΔMaGlox mutant was impaired in its ability to penetrate the host cuticle and evade host immune defense resulting in attenuated pathogenicity. Overexpression of MaGlox promoted fungal growth and conidial germination, increased tolerance to H2O2, but had little to other phenotypic effects. Transcriptomic analyses revealed downregulation of genes related to cell wall synthesis, conidiation, stress tolerance, and host cuticle penetration in the ΔMaGlox mutant. These findings demonstrate that MaGlox-mediated scavenging of RCS is required for virulence, and contributes to normal fungal growth and development, stress resistance.
Assuntos
Oxirredutases do Álcool , Proteínas Fúngicas , Metarhizium , Virulência , Oxirredutases do Álcool/metabolismo , Oxirredutases do Álcool/genética , Proteínas Fúngicas/metabolismo , Proteínas Fúngicas/genética , Metarhizium/patogenicidade , Metarhizium/genética , Metarhizium/metabolismo , Animais , Espécies Reativas de Oxigênio/metabolismo , Estresse Fisiológico , Estresse OxidativoRESUMO
Microglia and astrocytes play essential roles in the maintenance of homeostasis within the central nervous system, but mechanisms that control the magnitude and duration of responses to infection and injury remain poorly understood. Here, we provide evidence that 5-androsten-3ß,17ß-diol (ADIOL) functions as a selective modulator of estrogen receptor (ER)ß to suppress inflammatory responses of microglia and astrocytes. ADIOL and a subset of synthetic ERß-specific ligands, but not 17ß-estradiol, mediate recruitment of CtBP corepressor complexes to AP-1-dependent promoters, thereby repressing genes that amplify inflammatory responses and activate Th17 T cells. Reduction of ADIOL or ERß expression results in exaggerated inflammatory responses to TLR4 agonists. Conversely, the administration of ADIOL or synthetic ERß-specific ligands that promote CtBP recruitment prevents experimental autoimmune encephalomyelitis in an ERß-dependent manner. These findings provide evidence for an ADIOL/ERß/CtBP-transrepression pathway that regulates inflammatory responses in microglia and can be targeted by selective ERß modulators.
Assuntos
Encefalomielite Autoimune Experimental/metabolismo , Receptor beta de Estrogênio/metabolismo , Inflamação/metabolismo , Microglia/metabolismo , Transdução de Sinais , 17-Hidroxiesteroide Desidrogenases/metabolismo , Oxirredutases do Álcool/metabolismo , Androstenodiol/metabolismo , Animais , Astrócitos/metabolismo , Células Cultivadas , Proteínas de Ligação a DNA/metabolismo , Encefalomielite Autoimune Experimental/prevenção & controle , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Doenças Neurodegenerativas/metabolismo , Proteínas Proto-Oncogênicas c-fos/metabolismoRESUMO
The C-terminal binding protein (CtBP) is a transcriptional corepressor that plays critical roles in development, tumorigenesis, and cell fate. CtBP proteins are structurally similar to alpha hydroxyacid dehydrogenases and feature a prominent intrinsically disordered region in the C terminus. In the mammalian system, CtBP proteins lacking the C-terminal domain (CTD) are able to function as transcriptional regulators and oligomerize, putting into question the significance of this unstructured domain for gene regulation. Yet, the presence of an unstructured CTD of â¼100 residues, including some short motifs, is conserved across Bilateria, indicating the importance of maintaining this domain over evolutionary time. To uncover the significance of the CtBP CTD, we functionally tested naturally occurring Drosophila isoforms of CtBP that possess or lack the CTD, namely CtBP(L) and CtBP(S). We used the CRISPRi system to recruit dCas9-CtBP(L) and dCas9-CtBP(S) to endogenous promoters to directly compare their transcriptional impacts in vivo. Interestingly, CtBP(S) was able to significantly repress transcription of the Mpp6 promoter, while CtBP(L) was much weaker, suggesting that the long CTD may modulate CtBP's repression activity. In contrast, in cell culture, the isoforms behaved similarly on a transfected Mpp6 reporter gene. The context-specific differences in activity of these two developmentally regulated isoforms suggests that the CTD may help provide a spectrum of repression activity suitable for developmental programs.
Assuntos
Oxirredutases do Álcool , Proteínas de Drosophila , Regulação da Expressão Gênica , Domínios Proteicos , Proteínas Repressoras , Animais , Oxirredutases do Álcool/genética , Oxirredutases do Álcool/metabolismo , Drosophila/enzimologia , Drosophila/genética , Ligação Proteica , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Proteínas Repressoras/metabolismo , Domínios Proteicos/genética , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Linhagem Celular , Regulação da Expressão Gênica/genéticaRESUMO
l-2-hydroxyglutarate dehydrogenase (L2HGDH) is a mitochondrial membrane-associated metabolic enzyme, which catalyzes the oxidation of l-2-hydroxyglutarate (l-2-HG) to 2-oxoglutarate (2-OG). Mutations in human L2HGDH lead to abnormal accumulation of l-2-HG, which causes a neurometabolic disorder named l-2-hydroxyglutaric aciduria (l-2-HGA). Here, we report the crystal structures of Drosophila melanogaster L2HGDH (dmL2HGDH) in FAD-bound form and in complex with FAD and 2-OG and show that dmL2HGDH exhibits high activity and substrate specificity for l-2-HG. dmL2HGDH consists of an FAD-binding domain and a substrate-binding domain, and the active site is located at the interface of the two domains with 2-OG binding to the re-face of the isoalloxazine moiety of FAD. Mutagenesis and activity assay confirmed the functional roles of key residues involved in the substrate binding and catalytic reaction and showed that most of the mutations of dmL2HGDH equivalent to l-2-HGA-associated mutations of human L2HGDH led to complete loss of the activity. The structural and biochemical data together reveal the molecular basis for the substrate specificity and catalytic mechanism of L2HGDH and provide insights into the functional roles of human L2HGDH mutations in the pathogeneses of l-2-HGA.
Assuntos
Oxirredutases do Álcool , Encefalopatias Metabólicas Congênitas , Drosophila melanogaster , Modelos Moleculares , Animais , Humanos , Oxirredutases do Álcool/química , Oxirredutases do Álcool/metabolismo , Encefalopatias Metabólicas Congênitas/enzimologia , Encefalopatias Metabólicas Congênitas/genética , Encefalopatias Metabólicas Congênitas/fisiopatologia , Drosophila melanogaster/enzimologia , Glutaratos/metabolismo , Mutação , Domínio Catalítico/genética , Especificidade por Substrato/genética , Estrutura Terciária de Proteína , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismoRESUMO
Sickle cell disease (SCD) is a chronic hemolytic and systemic hypoxia condition with constant oxidative stress and significant metabolic alterations. However, little is known about the correlation between metabolic alterations and the pathophysiological symptoms. Here, we report that Nrf2, a master regulator of cellular antioxidant responses, regulates the production of the metabolite l-2-hydroxyglutarate (L2HG) to mediate epigenetic histone hypermethylation for gene expression involved in metabolic, oxidative, and ferroptotic stress responses in SCD. Mechanistically, Nrf2 was found to regulate the expression of L2HG dehydrogenase (L2hgdh) to mediate L2HG production under hypoxia. Gene expression profile analysis indicated that reactive oxygen species (ROS) and ferroptosis responses were the most significantly affected signaling pathways after Nrf2 ablation in SCD. Nrf2 silencing and L2HG supplementation sensitize human sickle erythroid cells to ROS and ferroptosis stress. The absence of Nrf2 and accumulation of L2HG significantly affect histone methylation for chromatin structure modification and reduce the assembly of transcription complexes on downstream target genes to regulate ROS and ferroptosis responses. Furthermore, pharmacological activation of Nrf2 was found to have protective effects against ROS and ferroptosis stress in SCD mice. Our data suggest a novel mechanism by which Nrf2 regulates L2HG levels to mediate SCD severity through ROS and ferroptosis stress responses, suggesting that targeting Nrf2 is a viable therapeutic strategy for ameliorating SCD symptoms.
Assuntos
Anemia Falciforme , Cromatina , Epigênese Genética , Ferroptose , Glutaratos , Fator 2 Relacionado a NF-E2 , Ferroptose/genética , Glutaratos/metabolismo , Fator 2 Relacionado a NF-E2/genética , Fator 2 Relacionado a NF-E2/metabolismo , Anemia Falciforme/genética , Anemia Falciforme/metabolismo , Anemia Falciforme/patologia , Cromatina/metabolismo , Metilação , Oxirredutases do Álcool/metabolismo , Animais , Camundongos , Espécies Reativas de Oxigênio/metabolismo , Transcrição Gênica , Perfilação da Expressão GênicaRESUMO
Although genome-wide hypomethylation is a hallmark of many cancers, roles for active DNA demethylation during tumorigenesis are unknown. Here, loss of the APC tumor suppressor gene causes upregulation of a DNA demethylase system and the concomitant hypomethylation of key intestinal cell fating genes. Notably, this hypomethylation maintained zebrafish intestinal cells in an undifferentiated state that was released upon knockdown of demethylase components. Mechanistically, the demethylase genes are directly activated by Pou5f1 and Cebpß and are indirectly repressed by retinoic acid, which antagonizes Pou5f1 and Cebpß. Apc mutants lack retinoic acid as a result of the transcriptional repression of retinol dehydrogenase l1 via a complex that includes Lef1, Groucho2, Ctbp1, Lsd1, and Corest. Our findings imply a model wherein APC controls intestinal cell fating through a switch in DNA methylation dynamics. Wild-type APC and retinoic acid downregulate demethylase components, thereby promoting DNA methylation of key genes and helping progenitors commit to differentiation.
Assuntos
Proteína da Polipose Adenomatosa do Colo/metabolismo , Polipose Adenomatosa do Colo/metabolismo , Metilação de DNA , Intestinos/embriologia , Peixe-Zebra/embriologia , Polipose Adenomatosa do Colo/patologia , Oxirredutases do Álcool/metabolismo , Animais , Encéfalo/citologia , Proteína beta Intensificadora de Ligação a CCAAT/metabolismo , Linhagem Celular Tumoral , Proliferação de Células , Proteínas Correpressoras/metabolismo , Neoplasias do Colo/metabolismo , Humanos , Mucosa Intestinal/metabolismo , Intestinos/citologia , Fator 3 de Transcrição de Octâmero/metabolismo , Fatores de Transcrição/metabolismo , Transcrição Gênica , Tretinoína/metabolismoRESUMO
One of the most abundant sources of organic carbon in the ocean is glycolate, the secretion of which by marine phytoplankton results in an estimated annual flux of one petagram of glycolate in marine environments1. Although it is generally accepted that glycolate is oxidized to glyoxylate by marine bacteria2-4, the further fate of this C2 metabolite is not well understood. Here we show that ubiquitous marine Proteobacteria are able to assimilate glyoxylate via the ß-hydroxyaspartate cycle (BHAC) that was originally proposed 56 years ago5. We elucidate the biochemistry of the BHAC and describe the structure of its key enzymes, including a previously unknown primary imine reductase. Overall, the BHAC enables the direct production of oxaloacetate from glyoxylate through only four enzymatic steps, representing-to our knowledge-the most efficient glyoxylate assimilation route described to date. Analysis of marine metagenomes shows that the BHAC is globally distributed and on average 20-fold more abundant than the glycerate pathway, the only other known pathway for net glyoxylate assimilation. In a field study of a phytoplankton bloom, we show that glycolate is present in high nanomolar concentrations and taken up by prokaryotes at rates that allow a full turnover of the glycolate pool within one week. During the bloom, genes that encode BHAC key enzymes are present in up to 1.5% of the bacterial community and actively transcribed, supporting the role of the BHAC in glycolate assimilation and suggesting a previously undescribed trophic interaction between autotrophic phytoplankton and heterotrophic bacterioplankton.
Assuntos
Organismos Aquáticos/metabolismo , Ácido Aspártico/análogos & derivados , Glicolatos/metabolismo , Redes e Vias Metabólicas , Proteobactérias/metabolismo , Oxirredutases do Álcool/metabolismo , Aldeído Liases/metabolismo , Organismos Aquáticos/enzimologia , Ácido Aspártico/metabolismo , Biocatálise , Glioxilatos/metabolismo , Hidroliases/metabolismo , Cinética , Oxirredutases/metabolismo , Fitoplâncton/enzimologia , Fitoplâncton/metabolismo , Proteobactérias/enzimologia , Transaminases/metabolismoRESUMO
Period circadian regulator 3 (PER3) functions as a tumor suppressor in various cancers. However, the role of PER3 in multiple myeloma (MM) has not been reported yet. Through this study, we aimed to investigate the potential role of PER3 in MM and the underlying mechanisms. RT-qPCR and western blotting were used to determine the mRNA and protein expression levels of PER3. Glyoxylate reductase 1 homolog (GLYR1) was predicted to be a transcription factor of PER3. The binding sites of GLYR1 on the promoter region of PER3 were analyzed using UCSC and confirmed using luciferase and chromatin immunoprecipitation assays. Viability, apoptosis, and metathesis were determined using CCK-8, colony formation, TUNEL, and transwell assays. We found that PER3 expression decreased in MM. Low PER3 levels may predict poor survival rates; PER3 overexpression suppresses the viability and migration of MM cells and promotes apoptosis. Moreover, GLYR1 transcriptionally activates PER3, and the knockdown of PER3 alleviates the effects of GLYR1 and induces its malignant behavior in MM cells. To conclude, GLYR1 upregulates PER3 and suppresses the aggressive behavior of MM cells, suggesting that GLYR1/PER3 signaling may be a potential therapeutic target for MM.
Assuntos
Movimento Celular , Proliferação de Células , Mieloma Múltiplo , Proteínas Circadianas Period , Humanos , Mieloma Múltiplo/genética , Mieloma Múltiplo/metabolismo , Mieloma Múltiplo/patologia , Linhagem Celular Tumoral , Proteínas Circadianas Period/genética , Proteínas Circadianas Period/metabolismo , Oxirredutases do Álcool/metabolismo , Oxirredutases do Álcool/genética , Apoptose , Regulação Neoplásica da Expressão GênicaRESUMO
Theoretical concepts linking the structure, function, and evolution of a protein, while often intuitive, necessitate validation through investigations in real-world systems. Our study empirically explores the evolutionary implications of multiple gene copies in an organism by shedding light on the structure-function modulations observed in Pseudomonas aeruginosa's second copy of ketopantoate reductase (PaKPR2). We demonstrated with two apo structures that the typical active site cleft of the protein transforms into a two-sided pocket where a molecular gate made up of two residues controls the substrate entry site, resulting in its inactivity toward the natural substrate ketopantoate. Strikingly, this structural modification made the protein active against several important α-keto-acid substrates with varied efficiency. Structural constraints at the binding site for this altered functional trait were analyzed with two binary complexes that show the conserved residue microenvironment faces restricted movements due to domain closure. Finally, its mechanistic highlights gathered from a ternary complex structure help in delineating the molecular perspectives behind its kinetic cooperativity toward these broad range of substrates. Detailed structural characteristics of the protein presented here also identified four key amino acid residues responsible for its versatile α-keto-acid reductase activity, which can be further modified to improve its functional properties through protein engineering.
Assuntos
Proteínas de Bactérias , Pseudomonas aeruginosa , Pseudomonas aeruginosa/enzimologia , Pseudomonas aeruginosa/genética , Pseudomonas aeruginosa/metabolismo , Proteínas de Bactérias/metabolismo , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Evolução Molecular , Domínio Catalítico , Especificidade por Substrato , Modelos Moleculares , Oxirredutases do Álcool/química , Oxirredutases do Álcool/metabolismo , Oxirredutases do Álcool/genética , Cristalografia por Raios X , Conformação Proteica , CinéticaRESUMO
Maintenance of metabolic homeostasis is secured by metabolite-sensing systems, which can be overwhelmed by constant macronutrient surplus in obesity. Not only the uptake processes but also the consumption of energy substrates determine the cellular metabolic burden. We herein describe a novel transcriptional system in this context comprised of peroxisome proliferator-activated receptor alpha (PPARα), a master regulator for fatty acid oxidation, and C-terminal binding protein 2 (CtBP2), a metabolite-sensing transcriptional corepressor. CtBP2 interacts with PPARα to repress its activity, and the interaction is enhanced upon binding to malonyl-CoA, a metabolic intermediate increased in tissues in obesity and reported to suppress fatty acid oxidation through inhibition of carnitine palmitoyltransferase 1. In line with our preceding observations that CtBP2 adopts a monomeric configuration upon binding to acyl-CoAs, we determined that mutations in CtBP2 that shift the conformational equilibrium toward monomers increase the interaction between CtBP2 and PPARα. In contrast, metabolic manipulations that reduce malonyl-CoA decreased the formation of the CtBP2-PPARα complex. Consistent with these in vitro findings, we found that the CtBP2-PPARα interaction is accelerated in obese livers while genetic deletion of CtBP2 in the liver causes derepression of PPARα target genes. These findings support our model where CtBP2 exists primarily as a monomer in the metabolic milieu of obesity to repress PPARα, representing a liability in metabolic diseases that can be exploited to develop therapeutic approaches.
Assuntos
Oxirredutases do Álcool , Proteínas Correpressoras , Obesidade , PPAR alfa , Humanos , Ácidos Graxos/metabolismo , Fígado/metabolismo , Obesidade/genética , Obesidade/metabolismo , PPAR alfa/genética , PPAR alfa/metabolismo , Oxirredutases do Álcool/metabolismo , Proteínas Correpressoras/metabolismo , Regulação AlostéricaRESUMO
Vanillyl alcohol oxidases (VAOs) belong to the 4-phenol oxidases family and are found predominantly in lignin-degrading ascomycetes. Systematical investigation of the enzyme family at the sequence level resulted in discovery and characterization of the second recombinantly produced VAO member, DcVAO, from Diplodia corticola. Remarkably high activities for 2,6-substituted substrates like 4-allyl-2,6-dimethoxy-phenol (3.5 ± 0.02 U mg-1) or 4-(hydroxymethyl)-2,6-dimethoxyphenol (6.3 ± 0.5 U mg-1) were observed, which could be attributed to a Phe to Ala exchange in the catalytic center. In order to rationalize this rare substrate preference among VAOs, we resurrected and characterized three ancestral enzymes and performed mutagenesis analyses. The results indicate that a Cys/Glu exchange was required to retain activity for É£-hydroxylations and shifted the acceptance towards benzyl ethers (up to 4.0 ± 0.1 U mg-1). Our findings contribute to the understanding of the functionality of VAO enzyme group, and with DcVAO, we add a new enzyme to the repertoire of ether cleaving biocatalysts.
Assuntos
Oxirredutases do Álcool , Ascomicetos , Biocatálise , Oxirredutases do Álcool/química , Oxirredutases do Álcool/genética , Oxirredutases do Álcool/metabolismo , Ascomicetos/enzimologia , Fenóis/química , Fenóis/metabolismo , Especificidade por Substrato , Hidroxilação , Éteres/química , Éteres/metabolismoRESUMO
Sensing of environmental challenges, such as mechanical injury, by a single plant tissue results in the activation of systemic signaling, which attunes the plant's physiology and morphology for better survival and reproduction. As key signals, both calcium ions (Ca2+ ) and hydrogen peroxide (H2 O2 ) interplay with each other to mediate plant systemic signaling. However, the mechanisms underlying Ca2+ -H2 O2 crosstalk are not fully revealed. Our previous study showed that the interaction between glycolate oxidase and catalase, key enzymes of photorespiration, serves as a molecular switch (GC switch) to dynamically modulate photorespiratory H2 O2 fluctuations via metabolic channeling. In this study, we further demonstrate that local wounding induces a rapid shift of the GC switch to a more interactive state in systemic leaves, resulting in a sharp decrease in peroxisomal H2 O2 levels, in contrast to a simultaneous outburst of the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase-derived apoplastic H2 O2 . Moreover, the systemic response of the two processes depends on the transmission of Ca2+ signaling, mediated by glutamate-receptor-like Ca2+ channels 3.3 and 3.6. Mechanistically, by direct binding and/or indirect mediation by some potential biochemical sensors, peroxisomal Ca2+ regulates the GC switch states in situ, leading to changes in H2 O2 levels. Our findings provide new insights into the functions of photorespiratory H2 O2 in plant systemic acclimation and an optimized systemic H2 O2 signaling via spatiotemporal interplay between the GC switch and NADPH oxidases.
Assuntos
Oxirredutases do Álcool , Plantas , Catalase/metabolismo , Plantas/metabolismo , Oxirredutases do Álcool/metabolismo , Receptores de Glutamato , Peróxido de Hidrogênio/metabolismoRESUMO
BACKGROUND: Purple non-heading Chinese cabbage [Brassica campestris (syn. Brassica rapa) ssp. chinensis] has become popular because of its richness in anthocyanin. However, anthocyanin only accumulates in the upper epidermis of leaves. Further studies are needed to investigate the molecular mechanisms underlying the specific accumulation of it. RESULTS: In this study, we used the laser capture frozen section method (LCM) to divide purple (ZBC) and green (LBC) non-heading Chinese cabbage leaves into upper and lower epidermis parts (Pup represents the purple upper epidermis, Plow represents the purple lower epidermis, Gup represents the green upper epidermis, Glow represents the green lower epidermis). Through transcriptome sequencing, we found that the DIHYDROFLAVONOL 4-REDUCTASE-encoding gene BcDFR, is strongly expressed in Pup but hardly in others (Plow, Gup, Glow). Further, a deletion and insertion in the promoter of BcDFR in LBC were found, which may interfere with BcDFR expression. Subsequent analysis of gene structure and conserved structural domains showed that BcDFR is highly conserved in Brassica species. The predicted protein-protein interaction network of BcDFR suggests that it interacts with almost all functional proteins in the anthocyanin biosynthesis pathway. Finally, the results of the tobacco transient expression also demonstrated that BcDFR promotes the synthesis and accumulation of anthocyanin. CONCLUSIONS: BcDFR is specifically highly expressed on the upper epidermis of purple non-heading Chinese cabbage leaves and regulates anthocyanin biosynthesis and accumulation. Our study provides new insights into the functional analysis and transcriptional regulatory network of anthocyanin-related genes in purple non-heading Chinese cabbage.
Assuntos
Antocianinas , Brassica , Proteínas de Plantas , Antocianinas/biossíntese , Brassica/genética , Brassica/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Transcriptoma , Microdissecção e Captura a Laser , Regulação da Expressão Gênica de Plantas , Perfilação da Expressão Gênica , Oxirredutases do Álcool/genética , Oxirredutases do Álcool/metabolismo , Folhas de Planta/genética , Folhas de Planta/metabolismo , RNA-Seq , Regiões Promotoras GenéticasRESUMO
Evolution of sequence-specific transcription factors clearly drives lineage-specific innovations, but less is known about how changes in the central transcriptional machinery may contribute to evolutionary transformations. In particular, transcriptional regulators are rich in intrinsically disordered regions that appear to be magnets for evolutionary innovation. The C-terminal Binding Protein (CtBP) is a transcriptional corepressor derived from an ancestral lineage of alpha hydroxyacid dehydrogenases; it is found in mammals and invertebrates, and features a core NAD-binding domain as well as an unstructured C-terminus (CTD) of unknown function. CtBP can act on promoters and enhancers to repress transcription through chromatin-linked mechanisms. Our comparative phylogenetic study shows that CtBP is a bilaterian innovation whose CTD of about 100 residues is present in almost all orthologs. CtBP CTDs contain conserved blocks of residues and retain a predicted disordered property, despite having variations in the primary sequence. Interestingly, the structure of the C-terminus has undergone radical transformation independently in certain lineages including flatworms and nematodes. Also contributing to CTD diversity is the production of myriad alternative RNA splicing products, including the production of "short" tailless forms of CtBP in Drosophila. Additional diversity stems from multiple gene duplications in vertebrates, where up to five CtBP orthologs have been observed. Vertebrate lineages show fewer major modifications in the unstructured CTD, possibly because gene regulatory constraints of the vertebrate body plan place specific constraints on this domain. Our study highlights the rich regulatory potential of this previously unstudied domain of a central transcriptional regulator.
Assuntos
Proteínas Repressoras , Fatores de Transcrição , Animais , Proteínas Repressoras/genética , Proteínas Repressoras/química , Filogenia , Fatores de Transcrição/metabolismo , Oxirredutases do Álcool/genética , Oxirredutases do Álcool/metabolismo , Drosophila/metabolismo , Vertebrados/metabolismo , Processamento Alternativo , Proteínas Correpressoras/genética , Proteínas Correpressoras/metabolismo , Ligação Proteica , Fosfoproteínas/genética , Mamíferos/metabolismoRESUMO
Long noncoding RNAs (lncRNAs) have emerged as important molecules and potential new targets for human cancers. This study investigates the function of lncRNA CTBP1 antisense RNA (CTBP1-AS) in prostate cancer (PCa) and explores the entailed molecular mechanism. Aberrantly expressed genes potentially correlated with PCa progression were probed using integrated bioinformatics analyses. A cohort of 68 patients with PCa was included, and their tumor and para-cancerous tissues were collected. CTBP1-AS was highly expressed in PCa tissues and cells and associated with poor patient prognosis. By contrast, tumor protein p63 (TP63) and S100 calcium binding protein A14 (S100A14) were poorly expressed in the PCa tissues and cells. CTBP1-AS did not affect TP63 expression; however it blocked the TP63-mediated transcriptional activation of S100A14, thereby reducing its expression. CTBP1-AS silencing suppressed proliferation, apoptosis resistance, migration, invasion, and tumorigenicity of PCa cell lines, while its overexpression led to inverse results. The malignant phenotype of cells was further weakened by TP63 overexpression but restored following artificial S100A14 silencing. In conclusion, this study demonstrates that CTBP1-AS plays an oncogenic role in PCa by blocking TP63-mediated transcriptional activation of S100A14. This may provide insight into the management of PCa.
Assuntos
Proliferação de Células , Progressão da Doença , Regulação Neoplásica da Expressão Gênica , Neoplasias da Próstata , RNA Longo não Codificante , Fatores de Transcrição , Proteínas Supressoras de Tumor , Animais , Humanos , Masculino , Camundongos , Oxirredutases do Álcool/genética , Oxirredutases do Álcool/metabolismo , Apoptose/genética , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Prognóstico , Neoplasias da Próstata/genética , Neoplasias da Próstata/patologia , Neoplasias da Próstata/metabolismo , RNA Antissenso/genética , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Ativação Transcricional , Proteínas Supressoras de Tumor/genética , Proteínas Supressoras de Tumor/metabolismoRESUMO
Human-borne acetone is a potent marker of lipid metabolism. Here, an enzyme immobilization method for secondary alcohol dehydrogenase (S-ADH), which is suitable for highly sensitive and selective biosensing of acetone, was developed, and then its applicability was demonstrated for spatiotemporal imaging of concentration distribution. After various investigations, S-ADH-immobilized meshes could be prepared with less than 5% variation by cross-linking S-ADH with glutaraldehyde on a cotton mesh at 40 °C for 15 min. Furthermore, high activity was obtained by adjusting the concentration of the coenzyme nicotinamide adenine dinucleotide (NADH) solution added to the S-ADH-immobilized mesh to 500 µM and the solvent to a potassium phosphate buffer solution at pH 6.5. The gas imaging system using the S-ADH-immobilized mesh was able to image the decrease in NADH fluorescence (ex 340 nm, fl 490 nm) caused by the catalytic reaction of S-ADH and the acetone distribution in the concentration range of 0.1-10 ppm-v, including the breath concentration of healthy people at rest. The exhaled breath of two healthy subjects at 6 h of fasting was quantified as 377 and 673 ppb-v, which were consistent with the values quantified by gas chromatography-mass spectrometry.
Assuntos
Acetona , Testes Respiratórios , Enzimas Imobilizadas , Acetona/análise , Acetona/química , Humanos , Enzimas Imobilizadas/química , Enzimas Imobilizadas/metabolismo , Técnicas Biossensoriais , Oxirredutases do Álcool/química , Oxirredutases do Álcool/metabolismo , Gases/química , Gases/análise , Expiração , NAD/análise , NAD/química , NAD/metabolismoRESUMO
Hyperuricemia is a chronic metabolic disease caused by purine metabolism disorder. And several gene loci and transporter proteins that associated with uric acid transport functions have been identified. Retinol Dehydrogenase 12 (RDH12), recognized for its role in safeguarding photoreceptors, and our study investigated the potential impact of Rdh12 mutations on other organs and diseases, particularly hyperuricemia. We assessed Rdh12 mRNA expression levels in various tissues and conducted serum biochemical analyses in Rdh12-/- mice. Compared with the wild type, significant alterations in serum uric acid levels and kidney-related biochemical indicators have been revealed. Then further analysis, including quantitative RT-PCR of gene expression in the liver and kidney, highlighted variations in the expression levels of specific genes linked to hyperuricemia. And renal histology assessment exposed mild pathological lesions in the kidneys of Rdh12-/- mice. In summary, our study suggests that Rdh12 mutations impact not only retinal function but also contribute to hyperuricemia and renal disease phenotypes in mice. Our finding implies that individuals with Rdh12 mutations may be prone to hyperuricemia and gout, emphasizing the significance of preventive measures and regular examinations in daily life.