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1.
Anticancer Res ; 41(11): 5481-5488, 2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-34732418

RESUMO

BACKGROUND/AIM: Aldehyde dehydrogenases (ALDHs) are considered as markers for normal and cancer stem cells (CSC) and are involved in cell metabolism, proliferation, differentiation, stemness, and retinoic acid (RA) biosynthesis. The aim of the present study was to identify the ALDH isoforms that are associated with the CSC phenotype in non-small cell lung and hepatocellular carcinomas. MATERIALS AND METHODS: We utilized lung (A549) and hepatocellular (HepG2) cancer cells and generated tumor spheres to isolate the CSC sub-population. RESULTS: The CSC enrichment was confirmed by the up-regulation of various CSC-related genes. Comparative qPCR analysis indicated the up-regulation of several ALDH isoforms in A549 and HepG2 spheres. Interestingly, cyclin D1 and Akt, down-stream targets of the RA signaling pathway, were also shown to be significantly up-regulated in both sphere populations. CONCLUSION: Specific ALDH isoforms appear to be important mediators for the acquisition of an CSC phenotype and thus, are potential promising targets for CSC-based therapeutic approaches in lung and hepatocellular carcinomas.


Assuntos
Aldeído Desidrogenase/metabolismo , Carcinoma Hepatocelular/enzimologia , Neoplasias Hepáticas/enzimologia , Neoplasias Pulmonares/enzimologia , Células-Tronco Neoplásicas/enzimologia , Células A549 , Aldeído Desidrogenase/genética , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patologia , Ciclina D1/genética , Ciclina D1/metabolismo , Regulação Enzimológica da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Células Hep G2 , Humanos , Isoenzimas , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patologia , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Células-Tronco Neoplásicas/patologia , Fenótipo , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais , Esferoides Celulares
2.
Medicina (Kaunas) ; 57(10)2021 Oct 13.
Artigo em Inglês | MEDLINE | ID: mdl-34684132

RESUMO

Background and objective:N-acetyltransferases 1 and 2 (NAT1 and NAT2) genes have polymorphisms in accordance with slow and rapid acetylator phenotypes with a role in the development of head and neck cancers (HNCs). Herein, we aimed to evaluate the association of NAT1 and NAT2 polymorphisms with susceptibility to HNCs in an updated meta-analysis. Materials and methods: A search was comprehensively performed in four databases (Web of Science, Scopus, PubMed/Medline, and Cochrane Library until 8 July 2021). The effect sizes, odds ratio (OR) along with 95% confidence interval (CI) were computed. Trial sequential analysis (TSA), publication bias and sensitivity analysis were conducted. Results: Twenty-eight articles including eight studies reporting NAT1 polymorphism and twenty-five studies reporting NAT2 polymorphism were involved in the meta-analysis. The results showed that individuals with slow acetylators of NAT2 polymorphism are at higher risk for HNC OR: 1.22 (95% CI: 1.02, 1.46; p = 0.03). On subgroup analysis, ethnicity, control source, and genotyping methods were found to be significant factors in the association of NAT2 polymorphism with the HNC risk. TSA identified that the amount of information was not large enough and that more studies are needed to establish associations. Conclusions: Slow acetylators in NAT2 polymorphism were related to a high risk of HNC. However, there was no relationship between NAT1 polymorphism and the risk of HNC.


Assuntos
Arilamina N-Acetiltransferase , Neoplasias de Cabeça e Pescoço , Acetiltransferases/genética , Arilamina N-Acetiltransferase/genética , Predisposição Genética para Doença , Neoplasias de Cabeça e Pescoço/genética , Humanos , Isoenzimas/genética , Polimorfismo Genético
3.
Plant Sci ; 312: 111031, 2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-34620435

RESUMO

Glutamate dehydrogenase (GDH) is a central enzyme in nitrogen metabolism, assimilating ammonia into glutamine or deaminating glutamate into α-oxoglutarate. Tea (Camellia sinensis L.) plants assimilate ammonium efficiently, but the role of CsGDH in ammonium assimilation remains unclear. We confirmed that tea has three GDH isogenes: CsGDH1-3. Bioinformatic analysis showed that CsGDH1 encodes the ß-GDH subunit, CsGDH2/3 encode the α-GDH subunit, and their proteins all feature an NADH-specific motif. CsGDH1 is mainly expressed in mature leaves and roots, CsGDH3 is mainly expressed in new shoots and roots, and CsGDH2 has the highest expression level in flowers compared to the other five tissues. Expression patterns of CsGDHs and glutamine synthetase isogenes (CsGSs) under different ammonium concentrations suggested that CsGDHs cooperate with CsGSs to assimilate ammonium, especially under high ammonium conditions. Inhibition of GS and its isogenes resulted in significant induction of CsGDH3 in roots and CsGDH2 in leaves, indicating their potential roles in ammonium assimilation. Moreover, CsGDHs transcripts were highly abundant in chlorotic tea leaves, in constrast to those of CsGSs, suggesting that CsGDHs play a vital role in ammonium assimilation in chlorotic tea mutant. Altogether, our circumstantial evidence that CsGDHs cooperate with CsGSs in ammonium assimilation provides a basis for unveiling their functions in tea plants.


Assuntos
Compostos de Amônio/metabolismo , Camellia sinensis/genética , Camellia sinensis/metabolismo , Glutamato Desidrogenase/genética , Glutamato Desidrogenase/metabolismo , Glutamato-Amônia Ligase/genética , Glutamato-Amônia Ligase/metabolismo , Produtos Agrícolas/genética , Produtos Agrícolas/metabolismo , Flores/metabolismo , Regulação da Expressão Gênica de Plantas , Genes de Plantas , Isoenzimas/genética , Isoenzimas/metabolismo , Folhas de Planta/metabolismo , Raízes de Plantas/metabolismo , Brotos de Planta/metabolismo
4.
Int J Mol Sci ; 22(19)2021 Sep 30.
Artigo em Inglês | MEDLINE | ID: mdl-34638951

RESUMO

The protein phosphorylation of the membrane-bound mitochondrial proteins has become of interest from the point of view of its regulatory role of the function of the respiratory chain, opening of the mitochondrial permeability transition pore (mPTP), and initiation of apoptosis. Earlier, we noticed that upon phosphorylation of proteins in some proteins, the degree of their phosphorylation increases with the opening of mPTP. Two isoforms of myelin basic protein and cyclic nucleotide phosphodiesterase were identified in rat brain non-synaptic mitochondria and it was concluded that they are involved in mPTP regulation. In the present study, using the mass spectrometry method, the phosphorylated protein was identified as Calpain 3 in rat brain non-synaptic mitochondria. In the present study, the phosphoprotein Calpain-3 (p94) (CAPN3) was identified in the rat brain mitochondria as a phosphorylated truncated form of p60-62 kDa by two-dimensional electrophoresis and mass spectrometry. We showed that the calpain inhibitor, calpeptin, was able to suppress the Ca2+ efflux from mitochondria, preventing the opening of mPTP. It was found that phosphorylated truncated CALP3 with a molecular weight of 60-62 contains p-Tyr, which indicates the possible involvement of protein tyrosine phosphatase in this process.


Assuntos
Encéfalo/metabolismo , Sinalização do Cálcio/efeitos dos fármacos , Calpaína/metabolismo , Isoenzimas/metabolismo , Mitocôndrias/metabolismo , Membranas Mitocondriais/metabolismo , Poro de Transição de Permeabilidade Mitocondrial/metabolismo , Proteínas Musculares/metabolismo , Animais , Apoptose/efeitos dos fármacos , Cálcio/metabolismo , Calpaína/antagonistas & inibidores , Calpaína/química , Inibidores de Cisteína Proteinase/farmacologia , Dipeptídeos/farmacologia , Isoenzimas/antagonistas & inibidores , Isoenzimas/química , Leupeptinas/farmacologia , Masculino , Peso Molecular , Proteínas Musculares/antagonistas & inibidores , Proteínas Musculares/química , Fosforilação , Transporte Proteico , Ratos
5.
BMC Plant Biol ; 21(1): 479, 2021 Oct 21.
Artigo em Inglês | MEDLINE | ID: mdl-34674662

RESUMO

Starch branching enzymes (SBEs) are key determinants of the structure and amount of the starch in plant organs, and as such, they have the capacity to influence plant growth, developmental, and fitness processes, and in addition, the industrial end-use of starch. However, little is known about the role of SBEs in determining starch structure-function relations in economically important horticultural crops such as fruit and leafy greens, many of which accumulate starch transiently. Further, a full understanding of the biological function of these types of starches is lacking. Because of this gap in knowledge, this minireview aims to provide an overview of SBEs in horticultural crops, to investigate the potential role of starch in determining postharvest quality. A systematic examination of SBE sequences in 43 diverse horticultural species, identified SBE1, 2 and 3 isoforms in all species examined except apple, olive, and Brassicaceae, which lacked SBE1, but had a duplicated SBE2. Among our findings after a comprehensive and critical review of published data, was that as apple, banana, and tomato fruits ripens, the ratio of the highly digestible amylopectin component of starch increases relative to the more digestion-resistant amylose fraction, with parallel increases in SBE2 transcription, fruit sugar content, and decreases in starch. It is tempting to speculate that during the ripening of these fruit when starch degradation occurs, there are rearrangements made to the structure of starch possibly via branching enzymes to increase starch digestibility to sugars. We propose that based on the known action of SBEs, and these observations, SBEs may affect produce quality, and shelf-life directly through starch accumulation, and indirectly, by altering sugar availability. Further studies where SBE activity is fine-tuned in these crops, can enrich our understanding of the role of starch across species and may improve horticulture postharvest quality.


Assuntos
Enzima Ramificadora de 1,4-alfa-Glucana/genética , Produtos Agrícolas/enzimologia , Isoenzimas , Amido/metabolismo , Enzima Ramificadora de 1,4-alfa-Glucana/metabolismo , Motivos de Aminoácidos , Amilopectina/metabolismo , Amilose/metabolismo , Produtos Agrícolas/genética , Produtos Agrícolas/normas , Grão Comestível , Armazenamento de Alimentos , Frutas , Horticultura , Filogenia , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Tubérculos , Açúcares/metabolismo , Verduras
6.
Appl Microbiol Biotechnol ; 105(21-22): 8059-8072, 2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-34622336

RESUMO

Branched-chain amino acid aminotransferase (BCAT) catalyzes bidirectional transamination in the cell between branched-chain amino acids (BCAAs; valine, leucine, and isoleucine) and branched-chain α-keto acids (BCKAs; α-ketoisovalerate, α-ketoisocaproate, and α-keto-ß-methylvalerate). Eukaryotic cells contain two types of paralogous BCATs: mitochondrial BCAT (BCATm) and cytosolic BCAT (BCATc). Both isozymes have identical enzymatic functions, so they have long been considered to perform similar physiological functions in the cells. However, many studies have gradually revealed the differences in physiological functions and regulatory mechanisms between them. In this article, we present overviews of BCATm and BCATc in both yeast and human. We also introduce BCAT variants found natively or constructed artificially, which could have significant implications for research into the relationship between the primary structures and protein functions of BCATs. KEY POINTS: • BCAT catalyzes bidirectional transamination in the cell between BCAAs and BCKAs. • BCATm and BCATc are different in the metabolic roles and regulatory mechanisms. • BCAT variants offer insight into a relationship between the structure and function.


Assuntos
Isoenzimas , Saccharomyces cerevisiae , Aminoácidos de Cadeia Ramificada , Citosol , Humanos , Isoenzimas/genética , Saccharomyces cerevisiae/genética , Transaminases/genética
7.
Nat Commun ; 12(1): 5963, 2021 10 13.
Artigo em Inglês | MEDLINE | ID: mdl-34645814

RESUMO

P4 ATPases are lipid flippases that are phylogenetically grouped into P4A, P4B and P4C clades. The P4A ATPases are heterodimers composed of a catalytic α-subunit and accessory ß-subunit, and the structures of several heterodimeric flippases have been reported. The S. cerevisiae Neo1 and its orthologs represent the P4B ATPases, which function as monomeric flippases without a ß-subunit. It has been unclear whether monomeric flippases retain the architecture and transport mechanism of the dimeric flippases. Here we report the structure of a P4B ATPase, Neo1, in its E1-ATP, E2P-transition, and E2P states. The structure reveals a conserved architecture as well as highly similar functional intermediate states relative to dimeric flippases. Consistently, structure-guided mutagenesis of residues in the proposed substrate translocation path disrupted Neo1's ability to establish membrane asymmetry. These observations indicate that evolutionarily distant P4 ATPases use a structurally conserved mechanism for substrate transport.


Assuntos
Adenosina Trifosfatases/química , Lisofosfolipídeos/química , Proteínas de Membrana Transportadoras/química , Fosfatidiletanolaminas/química , Fosfatidilserinas/química , Proteínas de Transferência de Fosfolipídeos/química , Proteínas de Saccharomyces cerevisiae/química , Saccharomyces cerevisiae/enzimologia , Adenosina Trifosfatases/genética , Adenosina Trifosfatases/metabolismo , Sequência de Aminoácidos , Sítios de Ligação , Membrana Celular/química , Membrana Celular/enzimologia , Clonagem Molecular , Microscopia Crioeletrônica , Expressão Gênica , Vetores Genéticos/química , Vetores Genéticos/metabolismo , Humanos , Interações Hidrofóbicas e Hidrofílicas , Isoenzimas/química , Isoenzimas/genética , Isoenzimas/metabolismo , Lisofosfolipídeos/metabolismo , Proteínas de Membrana Transportadoras/genética , Proteínas de Membrana Transportadoras/metabolismo , Modelos Moleculares , Fosfatidiletanolaminas/metabolismo , Fosfatidilserinas/metabolismo , Proteínas de Transferência de Fosfolipídeos/genética , Proteínas de Transferência de Fosfolipídeos/metabolismo , Ligação Proteica , Conformação Proteica em alfa-Hélice , Conformação Proteica em Folha beta , Domínios e Motivos de Interação entre Proteínas , Multimerização Proteica , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Especificidade por Substrato
8.
Nat Commun ; 12(1): 5418, 2021 09 14.
Artigo em Inglês | MEDLINE | ID: mdl-34521839

RESUMO

Advances in genomics have revealed many of the genetic underpinnings of human disease, but exposomics methods are currently inadequate to obtain a similar level of understanding of environmental contributions to human disease. Exposomics methods are limited by low abundance of xenobiotic metabolites and lack of authentic standards, which precludes identification using solely mass spectrometry-based criteria. Here, we develop and validate a method for enzymatic generation of xenobiotic metabolites for use with high-resolution mass spectrometry (HRMS) for chemical identification. Generated xenobiotic metabolites were used to confirm identities of respective metabolites in mice and human samples based upon accurate mass, retention time and co-occurrence with related xenobiotic metabolites. The results establish a generally applicable enzyme-based identification (EBI) for mass spectrometry identification of xenobiotic metabolites and could complement existing criteria for chemical identification.


Assuntos
Sistema Enzimático do Citocromo P-450/metabolismo , Espectrometria de Massas/métodos , Microssomos Hepáticos/enzimologia , Xenobióticos/metabolismo , Animais , Sistema Enzimático do Citocromo P-450/genética , Expressão Gênica , Humanos , Isoenzimas/genética , Isoenzimas/metabolismo , Marcação por Isótopo , Fígado/enzimologia , Desentoxicação Metabólica Fase I/genética , Desintoxicação Metabólica Fase II/genética , Camundongos
9.
Science ; 374(6567): eabj3624, 2021 10 29.
Artigo em Inglês | MEDLINE | ID: mdl-34581622
10.
Drugs R D ; 21(4): 385-397, 2021 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-34542871

RESUMO

BACKGROUND AND OBJECTIVE: Fabry disease, an X-linked lysosomal storage disorder characterized by absent or reduced alpha-galactosidase activity, is a lifelong disease that impairs patients' quality of life. Patients with Fabry disease have a considerably shortened lifespan, with mortality being mainly due to renal failure, cardiovascular disease, or cerebrovascular disease. Enzyme replacement therapy with agalsidase alfa has been shown to attenuate the renal, cardiovascular, and neuropathic disease progression associated with Fabry disease. The objective of this study was to investigate the safety of a new animal component-free version of agalsidase alfa. METHODS: A phase III/IV, open-label, single-arm, multicenter safety study was conducted in Canadian patients with Fabry disease between August 2011 and September 2017 as a regulatory requirement to assess the safety of agalsidase alfa produced using an animal component-free bioreactor process. Eligible patients had a documented diagnosis of Fabry disease and satisfied current Canadian guidelines for receiving enzyme replacement therapy for Fabry disease. Following treatment with animal component-free bioreactor-processed agalsidase alfa, treatment-emergent adverse events were monitored, and post hoc analyses of infusion-related reactions by antidrug antibody and neutralizing antibody statuses were conducted. The data were analyzed using descriptive statistics. RESULTS: A total of 167 patients (mean [standard deviation] age, 48.9 [14.8] years), including six pediatric patients (< 18 years of age), received at least one full or partial infusion of agalsidase alfa animal component-free. Fewer than 5% of treatment-emergent adverse events (212/4446) observed in 40 patients were reported as infusion-related reactions. Antidrug antibody and neutralizing antibody status did not affect the proportion of patients with infusion-related reactions. No clinically significant changes in vital signs were observed in patients over the course of the study. CONCLUSIONS: Long-term treatment with bioreactor-produced agalsidase alfa animal component-free did not reveal new safety signals in this population of Canadian patients with Fabry disease. The treatment-emergent adverse event profile was consistent with the clinical manifestations of the disease and the known safety profile of roller bottle-produced agalsidase alfa. CLINICAL TRIAL REGISTRATION: ClinicalTrials.gov identifier NCT01298141.


Assuntos
Doença de Fabry , alfa-Galactosidase , Animais , Reatores Biológicos , Canadá , Criança , Terapia de Reposição de Enzimas , Doença de Fabry/tratamento farmacológico , Humanos , Isoenzimas , Pessoa de Meia-Idade , Qualidade de Vida , Proteínas Recombinantes/uso terapêutico , Resultado do Tratamento , alfa-Galactosidase/efeitos adversos
11.
J Biol Chem ; 297(4): 101201, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-34537244

RESUMO

Different forms of photoreceptor degeneration cause blindness. Retinal degeneration-3 protein (RD3) deficiency in photoreceptors leads to recessive congenital blindness. We proposed that aberrant activation of the retinal membrane guanylyl cyclase (RetGC) by its calcium-sensor proteins (guanylyl cyclase-activating protein [GCAP]) causes this retinal degeneration and that RD3 protects photoreceptors by preventing such activation. We here present in vivo evidence that RD3 protects photoreceptors by suppressing activation of both RetGC1 and RetGC2 isozymes. We further suggested that insufficient inhibition of RetGC by RD3 could contribute to some dominant forms of retinal degeneration. The R838S substitution in RetGC1 that causes autosomal-dominant cone-rod dystrophy 6, not only impedes deceleration of RetGC1 activity by Ca2+GCAPs but also elevates this isozyme's resistance to inhibition by RD3. We found that RD3 prolongs the survival of photoreceptors in transgenic mice harboring human R838S RetGC1 (R838S+). Overexpression of GFP-tagged human RD3 did not improve the calcium sensitivity of cGMP production in R838S+ retinas but slowed the progression of retinal blindness and photoreceptor degeneration. Fluorescence of the GFP-tagged RD3 in the retina only partially overlapped with immunofluorescence of RetGC1 or GCAP1, indicating that RD3 separates from the enzyme before the RetGC1:GCAP1 complex is formed in the photoreceptor outer segment. Most importantly, our in vivo results indicate that, in addition to the abnormal Ca2+ sensitivity of R838S RetGC1 in the outer segment, the mutated RetGC1 becomes resistant to inhibition by RD3 in a different cellular compartment(s) and suggest that RD3 overexpression could be utilized to reduce the severity of cone-rod dystrophy 6 pathology.


Assuntos
Guanilato Ciclase/metabolismo , Proteínas Nucleares/metabolismo , Células Fotorreceptoras de Vertebrados/metabolismo , Receptores de Superfície Celular/metabolismo , Animais , Guanilato Ciclase/genética , Proteínas Ativadoras de Guanilato Ciclase/genética , Proteínas Ativadoras de Guanilato Ciclase/metabolismo , Células HEK293 , Humanos , Isoenzimas/genética , Isoenzimas/metabolismo , Camundongos , Camundongos Knockout , Mutação , Proteínas Nucleares/genética , Receptores de Superfície Celular/genética , Retinite Pigmentosa/genética , Retinite Pigmentosa/metabolismo
12.
Nat Commun ; 12(1): 5285, 2021 09 06.
Artigo em Inglês | MEDLINE | ID: mdl-34489442

RESUMO

The mammalian DNA methylome is formed by two antagonizing processes, methylation by DNA methyltransferases (DNMT) and demethylation by ten-eleven translocation (TET) dioxygenases. Although the dynamics of either methylation or demethylation have been intensively studied in the past decade, the direct effects of their interaction on gene expression remain elusive. Here, we quantify the concurrence of DNA methylation and demethylation by the percentage of unmethylated CpGs within a partially methylated read from bisulfite sequencing. After verifying 'methylation concurrence' by its strong association with the co-localization of DNMT and TET enzymes, we observe that methylation concurrence is strongly correlated with gene expression. Notably, elevated methylation concurrence in tumors is associated with the repression of 40~60% of tumor suppressor genes, which cannot be explained by promoter hypermethylation alone. Furthermore, methylation concurrence can be used to stratify large undermethylated regions with negligible differences in average methylation into two subgroups with distinct chromatin accessibility and gene regulation patterns. Together, methylation concurrence represents a unique methylation metric important for transcription regulation and is distinct from conventional metrics, such as average methylation and methylation variation.


Assuntos
Metilação de DNA , Metilases de Modificação do DNA/genética , Proteínas de Ligação a DNA/genética , Epigênese Genética , Genoma , Neoplasias/genética , Proteínas Proto-Oncogênicas/genética , Transcrição Genética , Animais , Cromatina/química , Cromatina/metabolismo , Ilhas de CpG , DNA/genética , DNA/metabolismo , Metilases de Modificação do DNA/metabolismo , Proteínas de Ligação a DNA/metabolismo , Ontologia Genética , Histonas/genética , Histonas/metabolismo , Humanos , Isoenzimas/genética , Isoenzimas/metabolismo , Camundongos , Anotação de Sequência Molecular , Células-Tronco Embrionárias Murinas/citologia , Células-Tronco Embrionárias Murinas/metabolismo , Neoplasias/metabolismo , Neoplasias/patologia , Proteínas Proto-Oncogênicas/metabolismo , Linfócitos T/citologia , Linfócitos T/metabolismo
13.
Cells ; 10(9)2021 09 04.
Artigo em Inglês | MEDLINE | ID: mdl-34571964

RESUMO

Oxidative stress within the vascular endothelium, due to excess generation of reactive oxygen species (ROS), is thought to be fundamental to the initiation and progression of the cardiovascular complications of type 2 diabetes mellitus. The term ROS encompasses a variety of chemical species including superoxide anion (O2•-), hydroxyl radical (OH-) and hydrogen peroxide (H2O2). While constitutive generation of low concentrations of ROS are indispensable for normal cellular function, excess O2•- can result in irreversible tissue damage. Excess ROS generation is catalysed by xanthine oxidase, uncoupled nitric oxide synthases, the mitochondrial electron transport chain and the nicotinamide adenine dinucleotide phosphate (NADPH) oxidases. Amongst enzymatic sources of O2•- the Nox2 isoform of NADPH oxidase is thought to be critical to the oxidative stress found in type 2 diabetes mellitus. In contrast, the transcriptionally regulated Nox4 isoform, which generates H2O2, may fulfil a protective role and contribute to normal glucose homeostasis. This review describes the key roles of Nox2 and Nox4, as well as Nox1 and Nox5, in glucose homeostasis, endothelial function and oxidative stress, with a key focus on how they are regulated in health, and dysregulated in type 2 diabetes mellitus.


Assuntos
Glicemia/metabolismo , Diabetes Mellitus Tipo 2/enzimologia , Angiopatias Diabéticas/enzimologia , Células Endoteliais/enzimologia , NADPH Oxidases/metabolismo , Estresse Oxidativo , Espécies Reativas de Oxigênio/metabolismo , Animais , Biomarcadores/sangue , Diabetes Mellitus Tipo 2/sangue , Diabetes Mellitus Tipo 2/patologia , Angiopatias Diabéticas/sangue , Angiopatias Diabéticas/patologia , Células Endoteliais/patologia , Homeostase , Humanos , Isoenzimas , Transdução de Sinais
14.
World J Gastroenterol ; 27(28): 4653-4666, 2021 Jul 28.
Artigo em Inglês | MEDLINE | ID: mdl-34366627

RESUMO

Gastric cancer accounts for the majority cancer-related deaths worldwide. Although various methods have considerably improved the screening, diagnosis, and treatment of gastric cancer, its incidence is still high in Asia, and the 5-year survival rate of advanced gastric cancer patients is only 10%-20%. Therefore, more effective drugs and better screening strategies are needed for reducing the incidence and mortality of gastric cancer. Cyclooxygenase-2 (COX-2) is considered to be the key inducible enzyme in prostaglandins (PGs) synthesis, which is involved in multiple pathways in the inflammatory response. For example, inflammatory cytokines stimulate innate immune responses via Toll-like receptors and nuclear factor-kappa B to induce COX-2/PGE2 pathway. In these processes, the production of an inflammatory microenvironment promotes the occurrence of gastric cancer. Epidemiological studies have also indicated that non-steroidal anti-inflammatory drugs can reduce the risk of malignant tumors of the digestive system by blocking the effect of COX-2. However, clinical use of COX-2 inhibitors to prevent or treat gastric cancer may be limited because of potential side effects, especially in the cardiovascular system. Given these side effects and low treatment efficacy, new therapeutic approaches and early screening strategies are urgently needed. Some studies have shown that genetic variation in COX-2 also play an important role in carcinogenesis. However, the genetic variation analysis in these studies is incomplete and isolated, pointing out only a few single nucleotide polymorphisms (SNPs) and the risk of gastric cancer, and no comprehensive study covering the whole gene region has been carried out. In addition, copy number variation (CNV) is not mentioned. In this review, we summarize the SNPs in the whole COX-2 gene sequence, including exons, introns, and both the 5' and 3' untranslated regions. Results suggest that COX-2 does not increase its expression through the CNV and the SNPs in COX-2 may serve as the potential marker to establish risk stratification in the general population. This review synthesizes emerging insights of COX-2 as a biomarker in multiple studies, summarizes the association between whole COX-2 sequence variation and susceptibility to gastric cancer, and discusses the future prospect of therapeutic intervention, which will be helpful for early screening and further research to find new approaches to gastric cancer treatment.


Assuntos
Neoplasias Gástricas , Anti-Inflamatórios não Esteroides , Ciclo-Oxigenase 1 , Ciclo-Oxigenase 2/genética , Inibidores de Ciclo-Oxigenase 2 , Variações do Número de Cópias de DNA , Humanos , Inflamação , Isoenzimas , Proteínas de Membrana , Neoplasias Gástricas/tratamento farmacológico , Neoplasias Gástricas/genética , Microambiente Tumoral
15.
Int J Mol Sci ; 22(15)2021 Jul 29.
Artigo em Inglês | MEDLINE | ID: mdl-34360902

RESUMO

Acyl-CoA:lysophosphatidylethanolamine acyltransferases (LPEATs) are known as enzymes utilizing acyl-CoAs and lysophospholipids to produce phosphatidylethanolamine. Recently, it has been discovered that they are also involved in the growth regulation of Arabidopsis thaliana. In our study we investigated expression of each Camelina sativa LPEAT isoform and their behavior in response to temperature changes. In order to conduct a more extensive biochemical evaluation we focused both on LPEAT enzymes present in microsomal fractions from C. sativa plant tissues, and on cloned CsLPEAT isoforms expressed in yeast system. Phylogenetic analyses revealed that CsLPEAT1c and CsLPEAT2c originated from Camelina hispida, whereas other isoforms originated from Camelina neglecta. The expression ratio of all CsLPEAT1 isoforms to all CsLPEAT2 isoforms was higher in seeds than in other tissues. The isoforms also displayed divergent substrate specificities in utilization of LPE; CsLPEAT1 preferred 18:1-LPE, whereas CsLPEAT2 preferred 18:2-LPE. Unlike CsLPEAT1, CsLPEAT2 isoforms were specific towards very-long-chain fatty acids. Above all, we discovered that temperature strongly regulates LPEATs activity and substrate specificity towards different acyl donors, making LPEATs sort of a sensor of external thermal changes. We observed the presented findings not only for LPEAT activity in plant-derived microsomal fractions, but also for yeast-expressed individual CsLPEAT isoforms.


Assuntos
Aciltransferases/metabolismo , Camellia/enzimologia , Camellia/genética , Fosfatidiletanolaminas/metabolismo , Proteínas de Plantas/metabolismo , Sementes/enzimologia , Temperatura , Acil Coenzima A/metabolismo , Aciltransferases/genética , Camellia/classificação , Camellia/crescimento & desenvolvimento , Resposta ao Choque Frio , DNA de Plantas/genética , Ativação Enzimática , Resposta ao Choque Térmico , Isoenzimas/genética , Microssomos/enzimologia , Filogenia , Proteínas de Plantas/genética , Sementes/crescimento & desenvolvimento , Especificidade por Substrato
16.
Biomolecules ; 11(8)2021 08 04.
Artigo em Inglês | MEDLINE | ID: mdl-34439822

RESUMO

The genome of the halophilic archaea Haloferax mediterranei contains three ORFs that show homology with glutamine synthetase (GS) (glnA-1, glnA-2, and glnA-3). Previous studies have focused on the role of GlnA-1, suggesting that proteins GlnA-2 and GlnA-3 could play a different role to that of GS. Glutamine synthetase (EC 6.3.1.2) belongs to the class of ligases, including 20 subclasses of other different enzymes, such as aspartate-ammonia ligase (EC 6.3.1.1), glutamate-ethylamine ligase (EC 6.3.1.6), and glutamate-putrescine ligase (EC 6.3.1.11). The reaction catalyzed by glutamate-putrescine ligase is comparable to the reaction catalyzed by glutamine synthetase (GS). Both enzymes can bind a glutamate molecule to an amino group: ammonium (GS) or putrescine (glutamate-putrescine ligase). In addition, they present the characteristic catalytic domain of GS, showing significant similarities in their structure. Although these proteins are annotated as GS, the bioinformatics and experimental results obtained in this work indicate that the GlnA-2 protein (HFX_1688) is a glutamate-putrescine ligase, involved in polyamine catabolism. The most significant results are those related to glutamate-putrescine ligase's activity and the analysis of the transcriptional and translational expression of the glnA-2 gene in the presence of different nitrogen sources. This work confirms a new metabolic pathway in the Archaea domain which extends the knowledge regarding the utilization of alternative nitrogen sources in this domain.


Assuntos
Proteínas Arqueais/genética , Proteínas de Escherichia coli/genética , Regulação da Expressão Gênica em Archaea , Ácido Glutâmico/metabolismo , Haloferax mediterranei/enzimologia , Ligases/genética , Fixação de Nitrogênio/genética , Putrescina/metabolismo , Trifosfato de Adenosina/metabolismo , Sequência de Aminoácidos , Amônia/metabolismo , Proteínas Arqueais/metabolismo , Clonagem Molecular , Biologia Computacional/métodos , Escherichia coli/enzimologia , Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Expressão Gênica , Vetores Genéticos/química , Vetores Genéticos/metabolismo , Haloferax mediterranei/genética , Isoenzimas/genética , Isoenzimas/metabolismo , Ligases/metabolismo , Filogenia , Biossíntese de Proteínas , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Salmonella typhimurium/enzimologia , Salmonella typhimurium/genética , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Especificidade por Substrato , Transcrição Genética
17.
Int J Mol Sci ; 22(16)2021 Aug 10.
Artigo em Inglês | MEDLINE | ID: mdl-34445292

RESUMO

The genes involved in implantation and placentation are tightly regulated to ensure a healthy pregnancy. The endoplasmic reticulum aminopeptidase 2 (ERAP2) gene is associated with preeclampsia (PE). Our studies have determined that an isoform of ERAP2-arginine (N), expressed in trophoblast cells (TC), significantly activates immune cells, and ERAP2N-expressing TCs are preferentially killed by both cytotoxic T lymphocytes (CTLs) and Natural Killer cells (NKCs). To understand the cause of this phenomenon, we surveyed differentially expressed genes (DEGs) between ERAP2N expressing and non-expressing TCs. Our RNAseq data revealed 581 total DEGs between the two groups. 289 genes were up-regulated, and 292 genes were down-regulated. Interestingly, most of the down-regulated genes of significance were pro-survival genes that play a crucial role in cell survival (LDHA, EGLN1, HLA-C, ITGB5, WNT7A, FN1). However, the down-regulation of these genes in ERAP2N-expressing TCs translates into a propensity for cell death. The Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that 64 DEGs were significantly enriched in nine pathways, including "Protein processing in endoplasmic reticulum" and "Antigen processing and presentation", suggesting that the genes may be associated with peptide processes involved in immune recognition during the reproductive cycle.


Assuntos
Aminopeptidases/genética , Morte Celular/genética , Trofoblastos/metabolismo , Substituição de Aminoácidos/genética , Aminopeptidases/metabolismo , Arginina/genética , Células Cultivadas , Citotoxicidade Imunológica/genética , Feminino , Perfilação da Expressão Gênica , Regulação Enzimológica da Expressão Gênica , Humanos , Isoenzimas/genética , Isoenzimas/metabolismo , Pré-Eclâmpsia/genética , Pré-Eclâmpsia/metabolismo , Pré-Eclâmpsia/patologia , Gravidez , Trofoblastos/patologia , Trofoblastos/fisiologia , Regulação para Cima/genética
18.
Int J Mol Sci ; 22(16)2021 Aug 10.
Artigo em Inglês | MEDLINE | ID: mdl-34445316

RESUMO

Inhibition of pyruvate dehydrogenase kinase (PDK) emerged as a potential strategy for treatment of cancer and metabolic disorders. Dichloroacetate (DCA), a prototypical PDK inhibitor, reduces the abundance of some PDK isoenzymes. However, the underlying mechanisms are not fully characterized and may differ across cell types. We determined that DCA reduced the abundance of PDK1 in breast (MDA-MB-231) and prostate (PC-3) cancer cells, while it suppressed both PDK1 and PDK2 in skeletal muscle cells (L6 myotubes). The DCA-induced PDK1 suppression was partially dependent on hypoxia-inducible factor-1α (HIF-1α), a transcriptional regulator of PDK1, in cancer cells but not in L6 myotubes. However, the DCA-induced alterations in the mRNA and the protein levels of PDK1 and/or PDK2 did not always occur in parallel, implicating a role for post-transcriptional mechanisms. DCA did not inhibit the mTOR signaling, while inhibitors of the proteasome or gene silencing of mitochondrial proteases CLPP and AFG3L2 did not prevent the DCA-induced reduction of the PDK1 protein levels. Collectively, our results suggest that DCA reduces the abundance of PDK in an isoform-dependent manner via transcriptional and post-transcriptional mechanisms. Differential response of PDK isoenzymes to DCA might be important for its pharmacological effects in different types of cells.


Assuntos
Ácido Dicloroacético/farmacologia , Inibidores Enzimáticos/farmacologia , Fibras Musculares Esqueléticas/metabolismo , Piruvato Desidrogenase Quinase de Transferência de Acetil/antagonistas & inibidores , Proteases Dependentes de ATP/antagonistas & inibidores , Proteases Dependentes de ATP/metabolismo , ATPases Associadas a Diversas Atividades Celulares/antagonistas & inibidores , ATPases Associadas a Diversas Atividades Celulares/metabolismo , Animais , Linhagem Celular Tumoral , Endopeptidase Clp/antagonistas & inibidores , Endopeptidase Clp/metabolismo , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Isoenzimas/antagonistas & inibidores , Isoenzimas/genética , Isoenzimas/metabolismo , Células PC-3 , Piruvato Desidrogenase Quinase de Transferência de Acetil/genética , Piruvato Desidrogenase Quinase de Transferência de Acetil/metabolismo , Ratos
19.
J Enzyme Inhib Med Chem ; 36(1): 1874-1883, 2021 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-34340614

RESUMO

A library of variously decorated N-phenyl secondary sulphonamides featuring the bicyclic tetrahydroquinazole scaffold was synthesised and biologically evaluated for their inhibitory activity against human carbonic anhydrase (hCA) I, II, IV, and IX. Of note, several compounds were identified showing submicromolar potency and excellent selectivity for the tumour-related hCA IX isoform. Structure-activity relationship data attained for various substitutions were rationalised by molecular modelling studies in terms of both inhibitory activity and selectivity.


Assuntos
Inibidores da Anidrase Carbônica/farmacologia , Biologia Computacional/métodos , Isoenzimas/antagonistas & inibidores , Quinazolinas/química , Sulfonamidas/farmacologia , Espectroscopia de Ressonância Magnética Nuclear de Carbono-13 , Inibidores da Anidrase Carbônica/síntese química , Inibidores da Anidrase Carbônica/química , Avaliação Pré-Clínica de Medicamentos , Simulação de Acoplamento Molecular , Espectroscopia de Prótons por Ressonância Magnética , Relação Estrutura-Atividade , Sulfonamidas/química
20.
J Enzyme Inhib Med Chem ; 36(1): 1783-1797, 2021 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-34340630

RESUMO

Carbonic Anhydrase Activators (CAAs) could represent a novel approach for the treatment of Alzheimer's disease, ageing, and other conditions that require remedial achievement of spatial learning and memory therapy. Within a research project aimed at developing novel CAAs selective for certain isoforms, three series of indole-based derivatives were investigated. Enzyme activation assay on human CA I, II, VA, and VII isoforms revealed several effective micromolar activators, with promising selectivity profiles towards the brain-associated cytosolic isoform hCA VII. Molecular modelling studies suggested a theoretical model of the complex between hCA VII and the new activators and provide a possible explanation for their modulating as well as selectivity properties. Preliminary biological evaluations demonstrated that one of the most potent CAA 7 is not cytotoxic and is able to increase the release of the brain-derived neurotrophic factor (BDNF) from human microglial cells, highlighting its possible application in the treatment of CNS-related disorders.


Assuntos
Anidrases Carbônicas/efeitos dos fármacos , Ativadores de Enzimas/farmacologia , Indóis/farmacologia , Isoenzimas/efeitos dos fármacos , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Espectroscopia de Ressonância Magnética Nuclear de Carbono-13 , Anidrases Carbônicas/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Ativação Enzimática , Ativadores de Enzimas/química , Ensaio de Imunoadsorção Enzimática/métodos , Humanos , Indóis/química , Isoenzimas/metabolismo , Microglia/citologia , Microglia/efeitos dos fármacos , Modelos Moleculares , Espectroscopia de Prótons por Ressonância Magnética , Especificidade por Substrato
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