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1.
Chemosphere ; 239: 124747, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31514003

RESUMO

BACKGROUNDS: Polychlorinated biphenyls are persistent environmental pollutants associated with the onset of non-alcoholic fatty liver disease in humans, but there is limited information on the underlying mechanism. In the present study, we investigated the alterations in gene expression profiles in normal human liver cells L-02 following exposure to 2, 3, 3', 4, 4', 5 - hexachlorobiphenyl (PCB 156), a potent compound that may induce non-alcoholic fatty liver disease. METHODS: The L-02 cells were exposed to PCB 156 for 72 h and the contents of intracellular triacylglyceride and total cholesterol were subsequently measured. Microarray analysis of mRNAs and long non-coding RNAs (lncRNAs) in the cells was also performed after 3.4 µM PCB 156 treatment. RESULTS: Exposure to PCB 156 (3.4 µM, 72 h) resulted in significant increases of triacylglyceride and total cholesterol concentrations in L-02 cells. Microarray analysis identified 222 differentially expressed mRNAs and 628 differentially expressed lncRNAs. Gene Ontology and pathway analyses associated the differentially expressed mRNAs with metabolic and inflammatory processes. Moreover, lncRNA-mRNA co-expression network revealed 36 network pairs comprising 10 differentially expressed mRNAs and 34 dysregulated lncRNAs. The results of bioinformatics analysis further indicated that dysregulated lncRNA NONHSAT174696, lncRNA NONHSAT179219, and lncRNA NONHSAT161887, as the regulators of EDAR, CYP1B1, and ALDH3A1 respectively, played an important role in the PCB 156-induced lipid metabolism disorder. CONCLUSION: Our findings provide an overview of differentially expressed mRNAs and lncRNAs in L-02 cells exposed to PCB 156, and contribute to the field of polychlorinated biphenyl-induced non-alcoholic fatty liver disease.


Assuntos
Fígado/efeitos dos fármacos , Bifenilos Policlorados/toxicidade , Transcriptoma/efeitos dos fármacos , Aldeído Desidrogenase/genética , Linhagem Celular , Colesterol/metabolismo , Citocromo P-450 CYP1B1/genética , Receptor Edar/genética , Perfilação da Expressão Gênica , Humanos , Fígado/citologia , Fígado/fisiologia , Hepatopatia Gordurosa não Alcoólica/induzido quimicamente , Hepatopatia Gordurosa não Alcoólica/patologia , Análise de Sequência com Séries de Oligonucleotídeos , RNA Longo não Codificante , RNA Mensageiro/metabolismo , Testes de Toxicidade , Triglicerídeos/metabolismo
2.
Fa Yi Xue Za Zhi ; 35(5): 576-580, 2019 Oct.
Artigo em Inglês, Chinês | MEDLINE | ID: mdl-31833292

RESUMO

Abstract: Objective To explore the change rules of blood ethanol and blood acetaldehyde concentration, the impairment of psychomotor functions of different acetaldehyde dehydrogenase (ALDH) 2 genotype individuals after alcohol consumption and the relationship among them. Methods The ALDH2 genotypes in seventy-nine healthy volunteers were obtained by SNaPshotTM method, then divided into ALDH2*1/*1 (wild type) and ALDH2*1/*2 (mutant type) group. After volunteers consumed 1.0 g/kg of alcohol, blood ethanol concentration and blood acetaldehyde concentration at a series of time points before and after alcohol consumption and psychomotor functions, such as, visual selective response time, auditory simple response time and tracking experiment were detected. Biphasic alcohol response questionnaires were collected. Results After alcohol consumption, ALDH2*1/*2 group's blood ethanol and blood acetaldehyde concentration reached the peak earlier than ALDH2*1/*1 group. Its blood acetaldehyde concentration was higher than that of ALDH2*1/*1 group, 1-6 h after alcohol consumption. The psychomotor functions, such as visual selective response time and auditory simple response time in ALDH2*1/*2 group were more significantly impaired than those in ALDH2*1/*1 group after alcohol consumption. There was no statistical significance between the two groups in excitement or sedation reactions (P>0.05). Pearson correlation coefficient test showed that blood acetaldehyde concentration was related with psychomotor function. Conclusion There are significant differences between the psychomotor function of ALDH2 wild type and mutant type individuals after alcohol consumption estimated to be related to the difference in blood acetaldehyde concentration after alcohol consumption.


Assuntos
Acetaldeído/sangue , Consumo de Bebidas Alcoólicas , Aldeído Desidrogenase/genética , Etanol/metabolismo , Polimorfismo Genético/genética , Desempenho Psicomotor/efeitos dos fármacos , Acetaldeído/metabolismo , Consumo de Bebidas Alcoólicas/efeitos adversos , Consumo de Bebidas Alcoólicas/sangue , Aldeído-Desidrogenase Mitocondrial , Aldeído Oxirredutases , Etanol/administração & dosagem , Etanol/sangue , Genótipo , Humanos , Desempenho Psicomotor/fisiologia
3.
Chem Biol Interact ; 314: 108822, 2019 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-31580832

RESUMO

Aldehyde dehydrogenase (ALDH) activity is not only a valuable marker for cancer cells with stem-like features, but also plays a vital role in drug resistance and disease progression in many tumors including melanoma. However, the precise role of ALDH activity in patient prognosis remains unclear. In this study, using the Cancer Genome Atlas (TCGA) RNA-sequencing expression data, we analyzed gene expression of ALDH isozymes in melanoma tumors to define the expression patterns and the prognostic and predictive values of these enzymes. We found that ALDH1A1 and ALDH1A3 had both higher and broader expression ranges in melanoma patients, and that ALDH1A3 expression correlated with better overall survival in metastatic melanoma. Further, stratification of the TCGA cohorts by the mutational subtypes of melanoma specifically revealed that expression of ALDH1A3 correlated with better prognosis in metastatic BRAF-mutant melanoma while expression of ALDH1A1 correlated with better prognosis in BRAF wild-type melanoma. Gene set enrichment analysis (GSEA) of these cohorts identified upregulation in oxidative phosphorylation, adipogenesis, and fatty acid metabolism signaling in ALDH1Alo patients, suggesting BRAF/MEK inhibitor resistance in that subset of patients. On the other hand, GSEA of ALDH1A3hi cohorts revealed upregulation in glycolysis, hypoxia and angiogenesis, suggesting BRAF/MEK inhibitor sensitivity in that subset of patients. Gene expression analysis using pre-treatment tumor samples supports high ALDH1A3 expression before BRAF/MEK inhibitor treatment as predictive of better treatment response in BRAF-mutant melanoma patients. Our study provides evidence that high ALDH1A3 mRNA expression is not only a prognostic marker but also a predictive marker for BRAF/MEK inhibitor treatment response in BRAF-mutant metastatic melanoma patients.


Assuntos
Aldeído Desidrogenase/genética , Aldeído Oxirredutases/genética , Melanoma/patologia , RNA Mensageiro/metabolismo , Idoso , Aldeído Desidrogenase/metabolismo , Aldeído Oxirredutases/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Estimativa de Kaplan-Meier , Masculino , Melanoma/metabolismo , Melanoma/mortalidade , Pessoa de Meia-Idade , Mutação , Prognóstico , Proteínas Proto-Oncogênicas B-raf/genética , Retinal Desidrogenase
4.
Nat Commun ; 10(1): 4068, 2019 09 06.
Artigo em Inglês | MEDLINE | ID: mdl-31492851

RESUMO

The aldehyde dehydrogenase (ALDH) family of metabolic enzymes converts aldehydes to carboxylates. Here, we find that the reductive consequence of ALDH7A1 activity, which generates NADH (nicotinamide adenine dinucleotide, reduced form) from NAD, underlies how ALDH7A1 coordinates a broad inhibition of the intracellular transport pathways. Studying vesicle formation by the Coat Protein I (COPI) complex, we elucidate that NADH generated by ALDH7A1 targets Brefeldin-A ADP-Ribosylated Substrate (BARS) to inhibit COPI vesicle fission. Moreover, defining a physiologic role for the broad transport inhibition exerted by ALDH7A1, we find that it acts to reduce energy consumption during hypoxia and starvation to promote cellular energy homeostasis. These findings advance the understanding of intracellular transport by revealing how the coordination of multiple pathways can be achieved, and also defining circumstances when such coordination is needed, as well as uncovering an unexpected way that NADH acts in cellular energetics.


Assuntos
Oxirredutases do Álcool/metabolismo , Aldeído Desidrogenase/metabolismo , Proteínas de Ligação a DNA/metabolismo , Metabolismo Energético , Homeostase , Espaço Intracelular/metabolismo , Oxirredutases do Álcool/genética , Aldeído Desidrogenase/genética , Transporte Biológico , Vesículas Revestidas pelo Complexo de Proteína do Envoltório/metabolismo , Hipóxia Celular , Proteínas de Ligação a DNA/genética , Células HEK293 , Células HeLa , Humanos , NAD/metabolismo , Transdução de Sinais , Inanição
5.
Adv Exp Med Biol ; 1193: 89-106, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31368099

RESUMO

Heart failure (HF) is a structural or functional cardiac abnormal syndrome characterized with series of symptoms and signs such as breathlessness, fatigue, pulmonary crackles, and peripheral edema. Being a terminal phase of most myocardial lesions, HF has become a leading cause of mobility and mortality worldwide, associated with heavy clinical burden and economic costs affecting over 23 million people [14]. There is an increase to 5.5% with systolic dysfunction and an increase to 36.0% with diastolic dysfunction in people 60 years or older [85]. The costs accompanied with heart failure stand 2-3% of the total healthcare system expenditure in high-income countries and are expected to increase >2-fold in the next 2 decades [34].


Assuntos
Aldeído Desidrogenase/genética , Insuficiência Cardíaca/genética , Custos de Cuidados de Saúde , Insuficiência Cardíaca/economia , Humanos
6.
J Biotechnol ; 303: 1-7, 2019 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-31310781

RESUMO

Phenolic aldehydes from lignocellulose pretreatment harshly inhibit the viability and metabolism of ethanol fermenting strains. Direct conversion of phenolic aldehydes is usually incomplete due to their low water solubility and recalcitrance to bioconversion. Here we consolidated phenolic aldehydes bioconversion and ethanol fermentation in a typical ethanologenic bacterium Zymomonas mobilis by constructing an intracellular oxidative pathway. The gene PP_2680 encoding NAD+-dependent aldehyde dehydrogenase from Pseudomonas putida KT2440 was expressed in Z. mobilis ZM4. The expression significantly improved both aldehyde inhibitor conversion and ethanol fermentability in corn stover hydrolysate. The purified PP_2680 aldehyde dehydrogenase showed strong in vitro oxidative capacity on phenolic aldehydes and its in vivo expression significantly up-regulated the key genes in the ED pathway and the oxidative phosphorylation. This study provided an important concept of simultaneous biodetoxification and fermentation in ethanologenic strains for the improvement of ethanol fermentability.


Assuntos
Aldeído Desidrogenase/metabolismo , Etanol/metabolismo , Zymomonas/crescimento & desenvolvimento , Aldeído Desidrogenase/genética , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Celulose , Fermentação , Regulação Bacteriana da Expressão Gênica , Fosforilação Oxidativa , Pseudomonas putida/enzimologia , Zea mays/química , Zymomonas/enzimologia , Zymomonas/genética
7.
BMC Vet Res ; 15(1): 224, 2019 Jul 02.
Artigo em Inglês | MEDLINE | ID: mdl-31266490

RESUMO

BACKGROUND: As a kind of opportunist pathogen, Staphylococcus xylosus (S. xylosus) can cause mastitis. Antibiotics are widely used for treating infected animals and tylosin is a member of such group. Thus, the continuous use of antibiotics in dairy livestock enterprise will go a long way in increasing tylosin resistance. However, the mechanism of tylosin-resistant S. xylosus is not clear. Here, isobaric tag for relative and absolute quantitation (iTRAQ)-based quantitative proteomics methods was used to find resistance-related proteins. RESULTS: We compared the differential expression of S. xylosus in response to tylosin stress by iTRAQ. A total of 155 proteins (59 up-regulated, 96 down-regulated) with the fold-change of >1.2 or <0.8 (p value ≤0.05) were observed between the S. xylosus treated with 1/2 MIC (0.25 µg/mL) tylosin and the untreated S. xylosus. Bioinformatic analysis revealed that these proteins play important roles in stress-response and transcription. Then, in order to verify the relationship between the above changed proteins and mechanism of tylosin-resistant S. xylosus, we induced the tylosin-resistant S. xylosus, and performed quantitative PCR analysis to verify the changes in the transcription proteins and the stress-response proteins in tylosin-resistant S. xylosus at the mRNA level. The data displayed that ribosomal protein L23 (rplw), thioredoxin(trxA) and Aldehyde dehydrogenase A(aldA-1) are up-regulated in the tylosin-resistant S. xylosus, compared with the tylosin-sensitive strains. CONCLUSION: Our findings demonstrate the important of stress-response and transcription in the tylosin resistance of S. xylosus and provide an insight into the prevention of this resistance, which would aid in finding new medicines .


Assuntos
Proteínas de Bactérias/metabolismo , Farmacorresistência Bacteriana/efeitos dos fármacos , Proteoma/análise , Staphylococcus/efeitos dos fármacos , Tilosina/farmacologia , Aldeído Desidrogenase/genética , Aldeído Desidrogenase/metabolismo , Antibacterianos/farmacologia , Proteínas de Bactérias/isolamento & purificação , Farmacorresistência Bacteriana/genética , Regulação Bacteriana da Expressão Gênica/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Proteômica/métodos , Proteínas Ribossômicas/genética , Proteínas Ribossômicas/metabolismo , Staphylococcus/genética , Staphylococcus/metabolismo , Tiorredoxinas/genética , Tiorredoxinas/metabolismo
8.
Appl Microbiol Biotechnol ; 103(14): 5917-5923, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31111182

RESUMO

Aliphatic medium-chain alkanes, a major component of gasoline, diesel, and jet fuels, are drop-in compatible fuels. Microorganisms with the capacity to produce medium-chain alkanes are promising for the bio-production of drop-in fuel. We found that Klebsiella sp. NBRC100048 has the ability to produce medium-chain alkanes from medium-chain aldehydes. We cloned a gene involved in conversion of aldehydes to alkanes by using a genomic fosmid library derived from Klebsiella sp. NBRC100048. The gene termed orf2991 encodes 506 amino acids and shows 62% sequence homology to the aldehyde dehydrogenase of Escherichia coli, aldB. The finding of orf2991 as a novel alkane-synthesizing enzyme gene similar to E. coli aldehyde dehydrogenase family, which is generally known to catalyze a reaction oxidizing aldehydes to fatty acids, indicated a novel function of aldehyde dehydrogenase. This finding is not only significant academically but allows developing the novel manufacturing methods of alkanes fermentation.


Assuntos
Alcanos/metabolismo , Proteínas de Bactérias/genética , Klebsiella/genética , Aldeído Desidrogenase/genética , Aldeídos/metabolismo , Proteínas de Bactérias/metabolismo , Biocombustíveis , Clonagem Molecular , Escherichia coli/genética , Biblioteca Genômica , Klebsiella/metabolismo , Engenharia Metabólica , Homologia de Sequência
9.
Chem Biol Interact ; 305: 86-97, 2019 May 25.
Artigo em Inglês | MEDLINE | ID: mdl-30928398

RESUMO

Substrate inhibition by the aldehyde has been observed for decades in NAD(P)+-dependent aldehyde dehydrogenase (ALDH) enzymes, which follow a Bi Bi ordered steady-state kinetic mechanism. In this work, by using theoretical simulations of different possible substrate inhibition mechanisms in monosubstrate and Bi Bi ordered steady-state reactions, we explored the kind and extent of errors arising when estimating the kinetic parameters and determining the kinetic mechanisms if substrate inhibition is intentionally or unintentionally ignored. We found that, in every mechanism, fitting the initial velocity data of apparently non-inhibitory substrate concentrations to a rectangular hyperbola produces important errors, not only in the estimation of Vmax values, which were underestimated as expected, but, surprisingly, even more in the estimation of Km values, which led to overestimation of the Vmax/Km values. We show that the greater errors in Km arises from fitting data that do experience substrate inhibition, although it may not be evident, to a Michaelis-Menten equation, which causes overestimation of the data at low substrate concentrations. Similarly, we show that if substrate inhibition is not fully assessed when inhibitors are evaluated, the estimated inhibition constants will have significant errors, and the type of inhibition could be grossly mistaken. We exemplify these errors with experimental results obtained with the betaine aldehyde dehydrogenase from spinach showing the errors predicted by the theoretical simulations and that these errors are increased in the presence of NADH, which in this enzyme favors aldehyde substrate inhibition. Therefore, we strongly recommend assessing substrate inhibition by the aldehyde in every ALDH kinetic study, particularly when inhibitors are evaluated. The common practices of using an apparently non-inhibitory concentration range of the aldehyde or a single high concentration of the aldehyde or the coenzyme when varying the other to determine true kinetic parameters should be abandoned.


Assuntos
Aldeído Desidrogenase/metabolismo , Aldeídos/metabolismo , Proteínas de Plantas/metabolismo , Proteínas Recombinantes/metabolismo , Aldeído Desidrogenase/antagonistas & inibidores , Aldeído Desidrogenase/genética , Aldeídos/química , Cinética , NAD/metabolismo , Proteínas de Plantas/antagonistas & inibidores , Proteínas de Plantas/genética , Proteínas Recombinantes/genética , Spinacia oleracea/enzimologia , Especificidade por Substrato
10.
Anticancer Res ; 39(4): 1719-1728, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30952711

RESUMO

BACKGROUND: The wingless-type mammary tumour virus integration site 5A (WNT5A) agonist Foxy5 was shown in vitro to affect intracellular signalling implicated in the regulation of colonic cancer stem cells (CSCs). MATERIALS AND METHODS: In order to study whether Foxy5 can modulate CSCs, either HT-29 or Caco-2 human colonic cancer cells, both lacking endogenous WNT5A expression, were inoculated subcutaneously into nude mice. RESULTS: Foxy5 reduced the expression of the stem-cell marker aldehyde dehydrogenase and, interestingly, the specific colon CSC marker double cortin-like kinase 1. Foxy5 also reduced active ß-catenin and the expression of its downstream target Achaete Scute complex homolog 2, a CSC-preserving transcription factor. Foxy5 also reduced cyclo-oxygenase 2 expression, responsible for the formation of the CSC-promoting prostaglandin E2 (PGE2), but increased that of 15-hydroxyprostaglandin dehydrogenase expression, a PGE2-degrading enzyme. Accordingly, Foxy5 impairs both ß-catenin and PGE2 signalling, both of which have been implicated in promoting the niche of colonic CSCs. CONCLUSION: Our data suggest that Foxy5 can complement the traditional adjuvant chemotherapeutic treatment to which CSCs are resistant.


Assuntos
Antineoplásicos/farmacologia , Neoplasias do Colo/tratamento farmacológico , Células-Tronco Neoplásicas/efeitos dos fármacos , Oligopeptídeos/farmacologia , Proteína Wnt-5a/agonistas , Aldeído Desidrogenase/genética , Aldeído Desidrogenase/metabolismo , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Células CACO-2 , Neoplasias do Colo/genética , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Ciclo-Oxigenase 2/metabolismo , Dinoprostona/metabolismo , Feminino , Células HT29 , Humanos , Hidroxiprostaglandina Desidrogenases/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Camundongos Endogâmicos BALB C , Camundongos Nus , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Retinal Desidrogenase , Carga Tumoral/efeitos dos fármacos , Microambiente Tumoral , Via de Sinalização Wnt/efeitos dos fármacos , Proteína Wnt-5a/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto , beta Catenina/metabolismo
11.
Appl Microbiol Biotechnol ; 103(10): 4017-4031, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30927024

RESUMO

High-level biosynthesis of desired metabolites is challenging due to complexity of metabolic networks. Here, we report that platform chemical 3-hydroxypropionic acid (3-HP) can be overproduced through promoter engineering and growth-sustaining cultivation, two parallel strategies relying on RNA polymerases (RNAPs). First, we screened a promoter library and revealed that IPTG-inducible tac promoter was most effective for overexpression of PuuC, an endogenous aldehyde dehydrogenase catalyzing 3-HP biosynthesis in Klebsiella pneumoniae. Next, tandem repetitive tac promoters were harnessed to accommodate adequate RNAPs. When three tandem repetitive tac promoters were recruited to overexpress PuuC, up to 102.61 g/L 3-HP was produced. This is the highest 3-HP titer reported so far. In addition, lactic acid completely vanished during the late stage of fermentation. The backflow of lactic acid to pyruvic acid saves the trouble of downstream separation of lactic acid from 3-HP, which has long been a mission impossible because they are small-molecule isomers. Furthermore, timely removal of acid stress and replenishment of nitrogen source are crucial for 3-HP biosynthesis. A mathematical model shows that RNAPs modulate the tradeoff between bacterial growth and 3-HP formation. Overall, promoter engineering and growth-promoting cultivation jointly leverage RNAPs to maximize 3-HP. This study provides a paradigm for maximizing growth-coupled metabolites.


Assuntos
Vias Biossintéticas/genética , Klebsiella pneumoniae/genética , Klebsiella pneumoniae/metabolismo , Ácido Láctico/análogos & derivados , Engenharia Metabólica/métodos , Regiões Promotoras Genéticas , Aldeído Desidrogenase/genética , Aldeído Desidrogenase/metabolismo , RNA Polimerases Dirigidas por DNA/genética , RNA Polimerases Dirigidas por DNA/metabolismo , Expressão Gênica , Klebsiella pneumoniae/crescimento & desenvolvimento , Ácido Láctico/biossíntese , Modelos Teóricos
12.
Biochimie ; 160: 183-192, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30905733

RESUMO

Constant addition of heavy metal pollutants in soil resulting from anthropogenic activities can prove detrimental to both macro and micro life forms inhabiting the ecosystem. The potential functional roles of eukaryotic microbes in such environment were explored in this study by metatranscriptomics approach. Sized eukaryotic cDNA libraries, library A (<0.5 kb), library B (0.5-1.0 kb), and library C (>1 kb) were constructed from the soil RNA and screened for copper (Cu) tolerance genes by using copper sensitive yeast mutant strain cup1Δ. Screening of the cDNA libraries yielded different clones capable of growing in Cu amended medium. In the present investigation, one of the transcripts PLCc38 from the library C was characterized and tested for its ability to tolerate different heavy metals by using metal sensitive yeast mutants. Sequence analysis PLCc38 showed homology with aldehyde dehydrogenase (ALDH) and capable of tolerating high concentrations of Cu (150-1000 µM). Aldeyde dehydrogenases are stress response enzymes capable of eliminating toxic levels of aldehydes generated due to abiotic environmental stresses. The cDNA PLCc38 also provided tolerance to wide range of Cd (40-100 µM), Zn (10-13 mM) and Co (2-50 mM) concentrations. Oxidative stress tolerance potential of PLCc38 was also confirmed in presence of different concentrations of H2O2. This study proves that PLCc38 is a potent gene associated with metal tolerance which could be used to revegetate heavy metal polluted soil or as a biomarker for detection of metal contamination.


Assuntos
Aldeído Desidrogenase/metabolismo , Biodegradação Ambiental , Cobre/farmacologia , Eucariotos/efeitos dos fármacos , Eucariotos/genética , Solo/química , Transcriptoma , Aldeído Desidrogenase/genética , Aldeído Desidrogenase/isolamento & purificação , Sequência de Aminoácidos , Ecossistema , Perfilação da Expressão Gênica , Metais Pesados , Filogenia , Homologia de Sequência , Microbiologia do Solo , Poluentes do Solo/farmacologia
14.
Chem Biol Interact ; 304: 83-87, 2019 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-30862475

RESUMO

Aldehyde dehydrogenases (ALDHs) comprise one of the most ancient protein superfamilies widely distributed in the three domains of life. Their members have been extensively studied in animals and plants, sorted out in different ALDH protein families and their participation in a broad variety of metabolic pathways has been documented. Paradoxically, no systematic studies comprising ALDHs from bacteria have been performed so far. Among bacteria, the genus Pseudomonas occupies numerous ecological niches, and is one of the most complex bacterial genera with the largest number of known species. For these reasons, we selected Pseudomonas as a paradigm to analyze the diversity of ALDHs in bacteria. With this aim, complete Pseudomonas genome sequences and annotations were retrieved from NCBI's RefSeq genome database. The 258 Pseudomonas strains belong to 46 different species, along with 23 with no species designation. The genomes of these Pseudomonas contain from 3,315 to 6,825 annotated protein coding genes. A total of 6,510 ALDH sequences were found in the selected Pseudomonas, with a median of 24 ALDH-coding genes per strain (by comparison humans possess only 19 different ALDH loci). Pseudomonas saudiphocaensis possesses the lowest number of aldh genes (9), while Pseudomonas pseudoalcaligenes KF707 NBRC110670 possesses the maximum number of aldh genes (49). The ALDHs found in Pseudomonas can be sorted out into 42 protein families, with a predominance of 14 families, which contained 76% of all ALDHs found. In this regard, it is important to note that many Pseudomonas genomes have multiple aldh genes coding for proteins belonging to the same family. Given that all strains contained members of families ALDH4, ALDH5, ALDH6, ALDH14, ALDH18 and ALDH27, we consider these families to be part of the core Pseudomonas genome.


Assuntos
Aldeído Desidrogenase/antagonistas & inibidores , Pseudomonas/efeitos dos fármacos , RNA Interferente Pequeno/farmacologia , Aldeído Desidrogenase/genética , Análise por Conglomerados , Humanos , Metabolômica , Proteômica , Pseudomonas/genética , Retinal Desidrogenase , Células Tumorais Cultivadas
15.
J Biol Chem ; 294(14): 5536-5548, 2019 04 05.
Artigo em Inglês | MEDLINE | ID: mdl-30737277

RESUMO

Ethanol (EtOH) is a teratogen, but its teratogenic mechanisms are not fully understood. The alcohol form of vitamin A (retinol/ROL) can be oxidized to all-trans-retinoic acid (RA), which plays a critical role in stem cell differentiation and development. Using an embryonic stem cell (ESC) model to analyze EtOH's effects on differentiation, we show here that EtOH and acetaldehyde, but not acetate, increase differentiation-associated mRNA levels, and that EtOH decreases pluripotency-related mRNAs. Using reporter assays, ChIP assays, and retinoic acid receptor-γ (RARγ) knockout ESC lines generated by CRISPR/Cas9 and homologous recombination, we demonstrate that EtOH signals via RARγ binding to RA response elements (RAREs) in differentiation-associated gene promoters or enhancers. We also report that EtOH-mediated increases in homeobox A1 (Hoxa1) and cytochrome P450 family 26 subfamily A member 1 (Cyp26a1) transcripts, direct RA target genes, require the expression of the RA-synthesizing enzyme, aldehyde dehydrogenase 1 family member A2 (Aldh1a2), suggesting that EtOH-mediated induction of Hoxa1 and Cyp26a1 requires ROL from the serum. As shown with CRISPR/Cas9 knockout lines, the retinol dehydrogenase gene Rdh10 and a functional RARE in the ROL transporter stimulated by retinoic acid 6 (Stra6) gene are required for EtOH induction of Hoxa1 and Cyp26a1 We conclude that EtOH stimulates stem cell differentiation by increasing the influx and metabolism of ROL for downstream RARγ-dependent transcription. In stem cells, EtOH may shift cell fate decisions to alter developmental outcomes by increasing endogenous ROL/RA signaling via increased Stra6 expression and ROL oxidation.


Assuntos
Diferenciação Celular/efeitos dos fármacos , Etanol/farmacologia , Células-Tronco Embrionárias Murinas/metabolismo , Receptores do Ácido Retinoico/metabolismo , Transdução de Sinais/efeitos dos fármacos , Aldeído Desidrogenase/biossíntese , Aldeído Desidrogenase/genética , Animais , Diferenciação Celular/genética , Proteínas de Homeodomínio/biossíntese , Proteínas de Homeodomínio/genética , Proteínas de Membrana/biossíntese , Proteínas de Membrana/genética , Camundongos , Camundongos Knockout , Células-Tronco Embrionárias Murinas/citologia , Receptores do Ácido Retinoico/genética , Retinal Desidrogenase , Ácido Retinoico 4 Hidroxilase/biossíntese , Ácido Retinoico 4 Hidroxilase/genética , Transdução de Sinais/genética , Fatores de Transcrição/biossíntese , Fatores de Transcrição/genética
16.
Methods Mol Biol ; 1923: 97-112, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30737736

RESUMO

The methylotrophic yeast Pichia pastoris (Komagataella phaffii) is used as an expression system for recombinant protein production for a variety of applications. It grows rapidly on inexpensive media containing methanol, glucose, glycerol, or ethanol as a sole carbon source. P. pastoris makes many posttranslational modifications and produces recombinant proteins either intracellularly or extracellularly. Because of these properties, P. pastoris has become a highly preferred host organism for biotechnology, pharmaceutical industry, and researchers.Recombinant protein production is usually performed under the control of the promoter of the alcohol oxidase gene I (AOX1). The AOX1 promoter is induced by methanol and repressed by glucose and ethanol. The regulation mechanisms of the AOX1 promoter have been studied in recent years. Another promoter used in recombinant protein production is derived from glyceraldehyde 3-phosphate dehydrogenase (GAP). It is a constitutive promoter. Recent literature showed that newly identified promoters of P. pastoris are promising as well, in addition to pAOX1 and pGAP.In this chapter, the regulation mechanisms of inducible pAOX1 and constitutive pGAP promoters are discussed. In addition, here we present an overview about the novel ADH3 promoter and alternative promoters of P. pastoris.


Assuntos
Pichia/genética , Regiões Promotoras Genéticas , Proteínas Recombinantes/genética , Oxirredutases do Álcool/genética , Aldeído Desidrogenase/genética , Regulação Fúngica da Expressão Gênica , Gliceraldeído 3-Fosfato Desidrogenase (NADP+)/genética , Microrganismos Geneticamente Modificados , Pichia/metabolismo , Engenharia de Proteínas/métodos , Proteínas Recombinantes/metabolismo
17.
Chem Biol Interact ; 303: 22-26, 2019 Apr 25.
Artigo em Inglês | MEDLINE | ID: mdl-30776359

RESUMO

Bioinformatic analyses of salmon (Salmo salar) ALDH amino acid sequences supported the presence of at least 30 ALDH genes, which is more than for any other higher vertebrate and is greater than the 19 human ALDH genes currently reported. These included 8 polyploid ALDH genes and proteins: ALDH1A2 (chromosomes 11 and 26); ALDH1L2 (chromosomes 7 and 17); ALDH2, encoding mitochondrial ALDH2 (chromosomes 2 and 5); ALDH3A2 (chromosomes 4, 9 and 20), for which evidence for 5 genes was obtained; ALDH3B1 (chromosomes 3, 6 and 24); ALDH4A1 (chromosomes 12 and 22); ALDH6A1 (chromosomes 1, 6 and 15); and ALDH18A1 (chromosomes 19 and 28). In contrast, 7 salmon ALDH gene families (ALDH1A1, ALDH1A3, ALDH5, ALDH7, ALDH8, ALDH9 and ALDH16) possessed only one gene family member. Phylogenetic studies of salmon and rainbow trout ALDH3A2 genes and proteins suggested that salmonid gene tetraploidy has occurred in at least 2 distinct stages of ALDH3A2 gene evolution.


Assuntos
Aldeído Desidrogenase/genética , Poliploidia , Aldeído Oxirredutases/genética , Animais , Evolução Molecular , Humanos , Oncorhynchus mykiss , Filogenia , Salmão
18.
Chem Biol Interact ; 302: 149-155, 2019 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-30794800

RESUMO

ALDH1L1 (cytosolic 10-formyltetrahydrofolate dehydrogenase) is the enzyme in folate metabolism commonly downregulated in human cancers. One of the mechanisms of the enzyme downregulation is methylation of the promoter of the ALDH1L1 gene. Recent studies underscored ALDH1L1 as a candidate tumor suppressor and potential marker of aggressive cancers. In agreement with the ALDH1L1 loss in cancer, its re-expression leads to inhibition of proliferation and to apoptosis, but also affects migration and invasion of cancer cells through a specific folate-dependent mechanism involved in invasive phenotype. A growing body of literature evaluated the prognostic value of ALDH1L1 expression for cancer disease, the regulatory role of the enzyme in cellular proliferation, and associated metabolic and signaling cellular responses. Overall, there is a strong indication that the ALDH1L1 silencing provides metabolic advantage for tumor progression at a later stage when unlimited proliferation and enhanced motility become critical processes for the tumor expansion. Whether the ALDH1L1 loss is involved in tumor initiation is still an open question.


Assuntos
Aldeído Desidrogenase/genética , Neoplasias/patologia , Aldeído Desidrogenase/antagonistas & inibidores , Aldeído Desidrogenase/metabolismo , Apoptose , Metilação de DNA , Progressão da Doença , Ácido Fólico/metabolismo , Humanos , Neoplasias/metabolismo , Oxirredutases atuantes sobre Doadores de Grupo CH-NH , Interferência de RNA , Transdução de Sinais , Proteína Supressora de Tumor p53/metabolismo
19.
mBio ; 10(1)2019 01 22.
Artigo em Inglês | MEDLINE | ID: mdl-30670620

RESUMO

Butanol production by Clostridium acetobutylicum is accompanied by coproduction of acetone and ethanol, which reduces the yield of butanol and increases the production cost. Here, we report development of several clostridial aldehyde/alcohol dehydrogenase (AAD) variants showing increased butanol selectivity by a series of design and analysis procedures, including random mutagenesis, substrate specificity feature analysis, and structure-based butanol selectivity design. The butanol/ethanol ratios (B/E ratios) were dramatically increased to 17.47 and 15.91 g butanol/g ethanol for AADF716L and AADN655H, respectively, which are 5.8-fold and 5.3-fold higher than the ratios obtained with the wild-type AAD. The much-increased B/E ratio obtained was due to the dramatic reduction in ethanol production (0.59 ± 0.01 g/liter) that resulted from engineering the substrate binding chamber and the active site of AAD. This protein design strategy can be applied generally for engineering enzymes to alter substrate selectivity.IMPORTANCE Renewable biofuel represents one of the answers to solving the energy crisis and climate change problems. Butanol produced naturally by clostridia has superior liquid fuel characteristics and thus has the potential to replace gasoline. Due to the lack of efficient genetic manipulation tools, however, clostridial strain improvement has been slower than improvement of other microorganisms. Furthermore, fermentation coproducing various by-products requires costly downstream processing for butanol purification. Here, we report the results of enzyme engineering of aldehyde/alcohol dehydrogenase (AAD) to increase butanol selectivity. A metabolically engineered Clostridium acetobutylicum strain expressing the engineered aldehyde/alcohol dehydrogenase gene was capable of producing butanol at a high level of selectivity.


Assuntos
Álcool Desidrogenase/metabolismo , Aldeído Desidrogenase/metabolismo , Butanóis/metabolismo , Clostridium acetobutylicum/enzimologia , Clostridium acetobutylicum/metabolismo , Engenharia Metabólica , Acetona/metabolismo , Álcool Desidrogenase/química , Álcool Desidrogenase/genética , Aldeído Desidrogenase/química , Aldeído Desidrogenase/genética , Domínio Catalítico , Etanol/metabolismo , Fermentação , Simulação de Dinâmica Molecular , Mutagênese Sítio-Dirigida
20.
J Biol Chem ; 294(2): 547-558, 2019 01 11.
Artigo em Inglês | MEDLINE | ID: mdl-30429219

RESUMO

Disruption of circadian rhythms has been implicated in an increased risk for cancer development. The Period2 (Per2) gene encodes one of the major components of the mammalian circadian clock, which plays a key role in controlling the circadian rhythms in physiology and behavior. PER2 has also been reported to suppress the malignant transformation of cells, but its role in the regulation of cancer susceptibility to chemotherapeutic drugs remains unclear. In this study, we found that oncogene-transformed embryonic fibroblasts prepared from Per2-mutant (Per2m/m ) mice, which are susceptible to both spontaneous and radiation-induced tumorigenesis, were resistant against common chemotherapeutic drugs and that this resistance is associated with up-regulation of the aldehyde dehydrogenase 3a1 (Aldh3a1) gene. Co-expression of the oncogenes H-rasV12 and SV40 large T-antigen induced malignant transformation of both WT and Per2m/m cells, but the cytotoxic effects of the chemotherapeutic agents methotrexate, gemcitabine, etoposide, vincristine, and oxaliplatin were significantly alleviated in the oncogene-transformed Per2m/m cells. Although introduction of the two oncogenes increased the expression of Aldh3a1 in both WT and Per2m/m cells, the ALDH3A1 protein levels in the Per2m/m cells were ∼7-fold higher than in WT cells. The elevated ALDH3A1 levels in the oncogene-transformed Per2m/m cells were sufficient to prevent chemotherapeutic drug-induced accumulation of reactive oxygen species. Consequently, shRNA-mediated suppression of Aldh3a1 expression relieved the chemoresistance of the Per2m/m cells. These results suggest a role for mutated PER2 in the development of multiple drug resistance and may inform therapeutic strategies for cancer management.


Assuntos
Aldeído Desidrogenase/genética , Carcinogênese/genética , Resistencia a Medicamentos Antineoplásicos , Proteínas Circadianas Period/genética , Regulação para Cima , Animais , Carcinogênese/efeitos dos fármacos , Células Cultivadas , Relógios Circadianos , Camundongos Endogâmicos ICR , Mutação , Neoplasias/tratamento farmacológico , Neoplasias/etiologia , Neoplasias/genética
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