Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 885
Filtrar
1.
Nat Commun ; 11(1): 4509, 2020 09 09.
Artigo em Inglês | MEDLINE | ID: mdl-32908151

RESUMO

Glycolysis is one of the primordial pathways of metabolism, playing a pivotal role in energy metabolism and biosynthesis. Glycolytic enzymes are known to form transient multi-enzyme assemblies. Here we examine the wider protein-protein interactions of plant glycolytic enzymes and reveal a moonlighting role for specific glycolytic enzymes in mediating the co-localization of mitochondria and chloroplasts. Knockout mutation of phosphoglycerate mutase or enolase resulted in a significantly reduced association of the two organelles. We provide evidence that phosphoglycerate mutase and enolase form a substrate-channelling metabolon which is part of a larger complex of proteins including pyruvate kinase. These results alongside a range of genetic complementation experiments are discussed in the context of our current understanding of chloroplast-mitochondrial interactions within photosynthetic eukaryotes.


Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/fisiologia , Cloroplastos/enzimologia , Glicólise/fisiologia , Mitocôndrias/enzimologia , Arabidopsis/citologia , Proteínas de Arabidopsis/genética , Metabolismo Energético/fisiologia , Mutação , Fosfoglicerato Mutase/genética , Fosfoglicerato Mutase/metabolismo , Fosfopiruvato Hidratase/genética , Fosfopiruvato Hidratase/metabolismo , Fotossíntese/fisiologia , Plantas Geneticamente Modificadas , Mapeamento de Interação de Proteínas , Mapas de Interação de Proteínas/fisiologia , Piruvato Quinase/genética , Piruvato Quinase/metabolismo
2.
Zhongguo Ying Yong Sheng Li Xue Za Zhi ; 36(2): 184-188, 2020 Mar.
Artigo em Chinês | MEDLINE | ID: mdl-32744017

RESUMO

Objective: To study the effects of α-enolase (ENO1) gene interference expression on proliferation, and cell cycle of follicular granulosa cells from Zi geese. Methods: F1 follicular granulosa cells were primary cultured (mixed culture), which were divided into four groups: ENO1 interference expression group (RNAi), unrelated sequence group (NC), culture group (Control), transfection reagent group (Lip). The apoptosis rate and cell cycle phase of the interference group and the control group were detected by the flow cytometry. Results: ENO1 gene interference expression slowed the proliferation of granulosa cells, increased the apoptosis, and increased the proportion of granulosa cells in G2/M phase. Conclusion: ENO1 gene interference expression could cause G2/M phase arrest in primary cultured goose follicular granulosa cells, induce cell apoptosis and inhibit cell proliferation.


Assuntos
Apoptose , Proliferação de Células , Gansos , Células da Granulosa/citologia , Fosfopiruvato Hidratase , Animais , Pontos de Checagem do Ciclo Celular , Feminino , Fosfopiruvato Hidratase/genética , Interferência de RNA
3.
Gene ; 763: 145069, 2020 Dec 30.
Artigo em Inglês | MEDLINE | ID: mdl-32827683

RESUMO

Neuropathic pain is caused by damage or disease happened to somatosensory nerve system. Due to the high prevalence and inefficient clinic intervention, neuropathic pain has brought considerable burden for world health care system. It is urgent to find novel targets for neuropathic pain basic research and clinical management. In this study, we found that miR-22-3p was decreased in Chronic Constriction Injury (CCI)rats and involved in neuropathic pain progression. Furthermore, it was found that ENO1 was a downstream target of miR-22-3p using bioinformatics analysis and luciferase reporter assays. MiR-22-3p downregulation promoted neuropathic pain via regulating inflammation factors expression by targeting ENO1. Then, we explored the upstream regulator of miR-22-3p using Miranda database. It was found that circular RNA ZNF609 sponged miR-22-3p by biotinylated RNA pull-down, AGO2-RIP, and luciferase reporter assays. Collectively, our study revealed that circZNF609 promoted inflammation factors expression to aggravate neuropathic pain progression via miR-22-3p/ENO1 axis in CCI rat models. Our study might provide a new direction for neuropathic pain basic research.


Assuntos
MicroRNAs/metabolismo , Neuralgia/metabolismo , Fosfopiruvato Hidratase/genética , RNA Circular/metabolismo , Animais , Células Cultivadas , Células HEK293 , Humanos , Masculino , MicroRNAs/genética , Neuralgia/genética , Fosfopiruvato Hidratase/metabolismo , RNA Circular/genética , Ratos , Ratos Sprague-Dawley
4.
Gene ; 756: 144911, 2020 Sep 25.
Artigo em Inglês | MEDLINE | ID: mdl-32574756

RESUMO

Enolase, a multifunctional glycolytic enzyme, is known to act as a plasminogen receptor in many species, involved in the pivotal processes such as motility, adhesion, invasion, growth, and differentiation of the parasites. Knowledge on the function of enolase from Dermanyssus gallinae is very limited. Here we report on the molecular cloning, enzymatic activity, tissue distribution and plasminogen binding activity of enolase from D. gallinae (DgENO). The full-length of cDNA was 1305 bp, specifying a peptide of 434 amino acids. Bioinformatics analysis showed that DgENO was highly conserved compared with a range of organisms, indicating the potentially similar functions in D. gallinae. A recombinant DgENO (rDgENO) protein was produced and characterized, it catalyzed the dehydration of 2-phospho-D-glycerate to phosphoenolpyruvate, the optimal pH was 7.5. Polyclonal antibodies were generated in mice and western blotting indicated that antiserum specifically recognized the native enolase in the somatic extracts from D. gallinae. Immunohistochemical staining of mite sections revealed that the distribution of DgENO was ubiquitous with high level in salivary gland, mite digestive tissues and fat bodies in D. gallinae. Expression level of DgENO was observed mostly in engorged adult mites. Moreover, ELISA binding assay showed that rDgENO could bind plasminogen, and lysine analog ε-aminocaproic acid significantly inhibited this binding activity, indicating that D. gallinae enolase is a receptor of plasminogen. The present study provided foundation for understanding of the biological functions of DgENO and its application in development of vaccines against D. gallinae.


Assuntos
Antígenos/imunologia , Ácaros/imunologia , Fosfopiruvato Hidratase/química , Vacinas/imunologia , Sequência de Aminoácidos , Animais , Anticorpos/imunologia , Anticorpos/isolamento & purificação , Antígenos/química , Antígenos/genética , Clonagem Molecular , Regulação da Expressão Gênica no Desenvolvimento , Ácaros/enzimologia , Ácaros/genética , Ácaros/crescimento & desenvolvimento , Fosfopiruvato Hidratase/análise , Fosfopiruvato Hidratase/genética , Plasminogênio/metabolismo , Alinhamento de Sequência
5.
Appl Environ Microbiol ; 86(13)2020 06 17.
Artigo em Inglês | MEDLINE | ID: mdl-32303553

RESUMO

At present, little is known about the RNA metabolism driven by the RNA degradosome in cyanobacteria. RNA helicase and enolase are the common components of the RNA degradosome in many bacteria. Here, we provide evidence that both enolase and the DEAD-box RNA helicase CrhB can interact with RNase E in Anabaena (Nostoc) sp. strain PCC 7120 (referred to here as PCC 7120). Furthermore, we found that the C-terminal domains of CrhB and AnaEno (enolase of PCC 7120) are required for the interaction, respectively. Moreover, their recognition motifs for AnaRne (RNase E of PCC 7120) turned out to be located in the N-terminal catalytic domain, which is obviously different from those identified previously in Proteobacteria We also demonstrated in enzyme activity assays that CrhB can induce AnaRne to degrade double-stranded RNA with a 5' tail. Furthermore, we investigated the localization of CrhB and AnaRne by green fluorescent protein (GFP) translation fusion in situ and found that they both localized in the center of the PCC 7120 cytoplasm. This localization pattern is also different from the membrane binding of RNase E and RhlB in Escherichia coli Together with the previous identification of polynucleotide phosphorylase (PNPase) in PCC 7120, our results show that there is an RNA degradosome-like complex with a different assembly mechanism in cyanobacteria.IMPORTANCE In all domains of life, RNA turnover is important for gene regulation and quality control. The process of RNA metabolism is regulated by many RNA-processing enzymes and assistant proteins, where these proteins usually exist as complexes. However, there is little known about the RNA metabolism, as well as about the RNA degradation complex. In the present study, we described an RNA degradosome-like complex in cyanobacteria and revealed an assembly mechanism different from that of E. coli Moreover, CrhB could help RNase E in Anabaena sp. strain PCC 7120 degrade double-stranded RNA with a 5' tail. In addition, CrhB and AnaRne have similar cytoplasm localizations, in contrast to the membrane localization in E. coli.


Assuntos
Anabaena/genética , Proteínas de Bactérias/genética , RNA Helicases DEAD-box/genética , Endorribonucleases/genética , Fosfopiruvato Hidratase/genética , Anabaena/enzimologia , Proteínas de Bactérias/metabolismo , RNA Helicases DEAD-box/metabolismo , Endorribonucleases/metabolismo , Complexos Multienzimáticos/genética , Complexos Multienzimáticos/metabolismo , Fosfopiruvato Hidratase/metabolismo , Polirribonucleotídeo Nucleotidiltransferase/genética , Polirribonucleotídeo Nucleotidiltransferase/metabolismo , RNA Helicases/genética , RNA Helicases/metabolismo
6.
Hum Cell ; 33(3): 790-800, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32304027

RESUMO

Anterior gradient 2 (AGR2) was proved to modulate cancer progression. However, the role of AGR2 on endometrial cancer was not established. Here, we investigated the effects of AGR2 expression on endometrial cancer and explored the regulation mechanism. In the study, we found that AGR2 was overexpressed in tumor tissues of 30 endometrial cancer patients. A high level of AGR2 promoted endometrial cancer cells proliferation, migration and invasion. AGR2 induced the expression of lactate dehydrogenase A (LDHA), phosphoglycerate kinase 1 (PGK1), kallikrein 2 (HK2), and enolase 1-α (ENO1), glucose uptake and lactate production. AGR2 could bind to MUC1 and induce MUC1 and hypoxia-inducible factor 1α (HIF-1α). The inhibition effects of AGR2 knockdown on cells proliferation, migration and invasion ability were abolished by the overexpression of MUC1. Besides, the overexpression of MUC1 also reversed the inhibition effects of AGR2 knockdown on the expression of LDHA, HK2, PGK1 and ENO1, glucose uptake and lactate production. AGR2 knockdown inhibited tumor growth, the levels of Ki-67, MUC1, HIF-1α and glycolysis. In conclusion, AGR2 was overexpressed in endometrial cancer and AGR2-induced glucose metabolism facilitated the progression of endometrial carcinoma via the MUC1/HIF-1α pathway. AGR2 may be an effective therapeutic target for endometrial carcinoma.


Assuntos
Carcinoma/genética , Carcinoma/patologia , Neoplasias do Endométrio/genética , Neoplasias do Endométrio/patologia , Glucose/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Mucina-1/genética , Mucina-1/metabolismo , Mucoproteínas/fisiologia , Proteínas Oncogênicas/fisiologia , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Carcinoma/metabolismo , Carcinoma/terapia , Movimento Celular/genética , Proliferação de Células/genética , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Progressão da Doença , Neoplasias do Endométrio/metabolismo , Neoplasias do Endométrio/terapia , Feminino , Expressão Gênica , Hexoquinase/genética , Hexoquinase/metabolismo , Humanos , L-Lactato Desidrogenase/genética , L-Lactato Desidrogenase/metabolismo , Terapia de Alvo Molecular , Mucoproteínas/genética , Mucoproteínas/metabolismo , Invasividade Neoplásica/genética , Proteínas Oncogênicas/genética , Proteínas Oncogênicas/metabolismo , Fosfoglicerato Quinase/genética , Fosfoglicerato Quinase/metabolismo , Fosfopiruvato Hidratase/genética , Fosfopiruvato Hidratase/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Proteínas Supressoras de Tumor/genética , Proteínas Supressoras de Tumor/metabolismo
7.
J Virol ; 94(8)2020 03 31.
Artigo em Inglês | MEDLINE | ID: mdl-31969437

RESUMO

Kaposi's sarcoma-associated herpesvirus (KSHV) is etiologically associated with endothelial Kaposi's sarcoma (KS) in immunocompromised individuals. KS lesion cells exhibit many similarities to neuroendocrine (NE) cancers, such as highly vascular and red/purple tumor lesions, spindle-shaped cells, an insignificant role for classic oncogenes in tumor development, the release of bioactive amines, and indolent growth of the tumors. However, the mechanistic basis for the similarity of KS lesion endothelial cells to neuroendocrine tumors remains unknown. Next-generation sequencing and bioinformatics analysis in the present study demonstrate that endothelial cells latently infected with KSHV express several neuronal and NE genes. De novo infection of primary dermal endothelial cells with live and UV-inactivated KSHV demonstrated that viral gene expression is responsible for the upregulation of five selected NE genes (adrenomedullin 2 [ADM2], histamine receptor H1 [HRH1], neuron-specific enolase [NSE] [ENO2], neuronal protein gene product 9.5 [PGP9.5], and somatostatin receptor 1 [SSTR1]). Immunofluorescence and immunohistochemistry examinations demonstrated the robust expression of the NE genes HRH1 and NSE/ENO2 in KSHV-infected KS tissue samples and KS visceral tissue microarrays. Further analysis demonstrated that KSHV latent open reading frame K12 (ORFK12) gene (kaposin A)-mediated decreased host REST/NRSF (RE1-silencing transcription factor/neuron-restrictive silencer factor) protein, a neuronal gene transcription repressor protein, is responsible for NE gene expression in infected endothelial cells. The NE gene expression observed in KSHV-infected cells was recapitulated in uninfected endothelial cells by the exogenous expression of ORFK12 and by the treatment of cells with the REST inhibitor X5050. When the neuroactive ligand-activating receptor HRH1 and inhibitory SSTR1 were knocked out by CRISPR, HRH1 knockout (KO) significantly inhibited cell proliferation, while SSTR1 KO induced cell proliferation, thus suggesting that HRH1 and SSTR1 probably counteract each other in regulating KSHV-infected endothelial cell proliferation. These results demonstrate that the similarity of KS lesion cells to neuroendocrine tumors is probably a result of KSHV infection-induced transformation of nonneuronal endothelial cells into cells with neuroendocrine features. These studies suggest a potential role of neuroendocrine pathway genes in the pathobiological characteristics of KSHV-infected endothelial cells, including a potential mechanism of escape from the host immune system by the expression of immunologically privileged neuronal-site NE genes, and NE genes could potentially serve as markers for KSHV-infected KS lesion endothelial cells as well as novel therapeutic targets to control KS lesions.IMPORTANCE Kaposi's sarcoma-associated herpesvirus (KSHV) manipulates several cellular pathways for its survival advantage during its latency in the infected human host. Here, we demonstrate that KSHV infection upregulates the expression of genes related to neuronal and neuroendocrine (NE) functions that are characteristic of NE tumors, both in vitro and in KS patient tissues and the heterogeneity of neuroendocrine receptors having opposing roles in KSHV-infected cell proliferation. Induction of NE genes by KSHV could also provide a potential survival advantage, as the expression of proteins at immunologically privileged sites such as neurons on endothelial cells may be an avenue to escape host immune surveillance functions. The NE gene products identified here could serve as markers for KSHV-infected cells and could potentially serve as therapeutic targets to combat KSHV-associated KS.


Assuntos
Carcinoma Neuroendócrino/genética , Células Endoteliais/virologia , Regulação Neoplásica da Expressão Gênica , Infecções por Herpesviridae/virologia , Herpesvirus Humano 8/fisiologia , Sarcoma de Kaposi/genética , Sarcoma de Kaposi/virologia , Carcinoma Neuroendócrino/patologia , Linhagem Celular , Proliferação de Células , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Regulação Viral da Expressão Gênica , Técnicas de Inativação de Genes , Infecções por Herpesviridae/patologia , Humanos , Fases de Leitura Aberta/genética , Hormônios Peptídicos/genética , Fosfopiruvato Hidratase/genética , Receptores Histamínicos/genética , Receptores de Somatostatina/genética , Proteínas Repressoras/genética , Ubiquitina Tiolesterase/genética , Regulação para Cima , Proteínas Virais/genética , Latência Viral/genética , Latência Viral/fisiologia
8.
PLoS One ; 15(1): e0228134, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31990932

RESUMO

Chronic lameness affects bovine welfare and has a negative economic impact in dairy industry. Moreover, due to the translational gap between traditional pain models and new drugs development for treating chronic pain states, naturally occurring painful diseases could be a potential translational tool for chronic pain research. We therefore employed liquid chromatography tandem mass spectrometry (LC-MS/MS) to stablish the proteomic profile of the spinal cord samples from lumbar segments (L2-L4) of chronic lame dairy cows. Data were validated and quantified through software tool (Scaffold® v 4.0) using output data from two search engines (SEQUEST® and X-Tandem®). Search Tool for the Retrieval of Interacting Genes/Proteins (STRING) analysis was performed to detect proteins interactions. LC-MS/MS identified a total amount of 177 proteins; of which 129 proteins were able to be quantified. Lame cows showed a strong upregulation of interacting proteins with chaperone and stress functions such as Hsp70 (p < 0.006), Hsc70 (p < 0.0079), Hsp90 (p < 0.015), STIP (p > 0.0018) and Grp78 (p <0.0068), and interacting proteins associated to glycolytic pathway such as; γ-enolase (p < 0.0095), α-enolase (p < 0.013) and hexokinase-1 (p < 0.028). It was not possible to establish a clear network of interaction in several upregulated proteins in lame cows. Non-interacting proteins were mainly associated to redox process and cytoskeletal organization. The most relevant down regulated protein in lame cows was myelin basic protein (MBP) (p < 0.02). Chronic inflammatory lameness in cows is associated to increased expression of stress proteins with chaperone, metabolism, redox and structural functions. A state of endoplasmic reticulum stress and unfolded protein response (UPR) might explain the changes in protein expression in lame cows; however, further studies need to be performed in order to confirm these findings.


Assuntos
Doenças dos Bovinos/genética , Dor Crônica/veterinária , Regulação da Expressão Gênica , Coxeadura Animal/genética , Proteína Básica da Mielina/genética , Proteínas do Tecido Nervoso/genética , Animais , Bovinos , Doenças dos Bovinos/metabolismo , Doenças dos Bovinos/fisiopatologia , Dor Crônica/genética , Dor Crônica/metabolismo , Dor Crônica/fisiopatologia , Indústria de Laticínios , Feminino , Perfilação da Expressão Gênica , Ontologia Genética , Redes Reguladoras de Genes , Proteínas de Choque Térmico/genética , Proteínas de Choque Térmico/metabolismo , Isoenzimas/genética , Isoenzimas/metabolismo , Lactação/fisiologia , Coxeadura Animal/metabolismo , Coxeadura Animal/fisiopatologia , Anotação de Sequência Molecular , Proteína Básica da Mielina/metabolismo , Proteínas do Tecido Nervoso/classificação , Proteínas do Tecido Nervoso/metabolismo , Fosfopiruvato Hidratase/genética , Fosfopiruvato Hidratase/metabolismo , Proteômica/métodos , Corno Dorsal da Medula Espinal/metabolismo , Corno Dorsal da Medula Espinal/fisiopatologia
9.
Exp Cell Res ; 386(1): 111713, 2020 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-31705846

RESUMO

Reprogrammed glucose metabolism is essential for tumor initiation and development, especially for pancreatic ductal adenocarcinoma (PDAC). Most cancer cells rely on aerobic glycolysis, a phenomenon termed "the Warburg effect", to support uncontrolled proliferation and evade apoptosis. However, the direct regulators of the Warburg effect remain areas of active investigation. In this study, we found that the highly conserved transcription factor, TWIST1, is a crucial regulator of aerobic glycolysis in PDAC. Genetic silencing of TWIST1 significantly inhibited the glycolytic phenotypes of PDAC cells as revealed by reduced glucose uptake, lactate production, and extracellular acidification rate, which can be restored by re-expression of siRNA-resistant TWIST1. Moreover, tamoxifen-inducible expression of TWIST1 promoted the Warburg metabolism of PDAC cells. Mechanistically, by luciferase reporter assay and chromatin immunoprecipitation experiment, we showed that TWIST1 can directly increase the expression of several glycolytic genes, including SLC2A1, HK2, ENO1, and PKM2. Of note, the transcriptional regulation by TWIST1 was not dependent on HIF1α or c-Myc. In The Cancer Genome Atlas and Gene Expression Omnibus accession GSE15471, we confirmed that TWIST1 was closely associated with the glycolysis pathway. Collectively, our findings indicate that TWIST1 is likely to act as important regulator of the Warburg effect in PDAC.


Assuntos
Adenocarcinoma/metabolismo , Regulação Neoplásica da Expressão Gênica , Glicólise , Proteínas Nucleares/genética , Neoplasias Pancreáticas/metabolismo , Proteína 1 Relacionada a Twist/genética , Adenocarcinoma/genética , Adenocarcinoma/patologia , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Linhagem Celular Tumoral , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Transportador de Glucose Tipo 1/genética , Transportador de Glucose Tipo 1/metabolismo , Hexoquinase/genética , Hexoquinase/metabolismo , Humanos , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patologia , Fosfopiruvato Hidratase/genética , Fosfopiruvato Hidratase/metabolismo , Hormônios Tireóideos/genética , Hormônios Tireóideos/metabolismo , Proteínas Supressoras de Tumor/genética , Proteínas Supressoras de Tumor/metabolismo
10.
J Biotechnol ; 308: 40-55, 2020 Jan 20.
Artigo em Inglês | MEDLINE | ID: mdl-31706887

RESUMO

Pyrimethamine is an effective drug in the cancer cell treatment and is a dihydrofolate reductase inhibitor. In this work, the amount of drug loading up on CNT and its cytotoxicity effect upon MCF-7 cell lines was surveyed. The novel applications of some drugs and nanocarriers can induce the differentiation of adipose mesenchymal cells into nerve cells. Hence carbon nanotube-pyrimethamine was used to differentiate mesenchymal stem cells into the neural category, for the first time. The results of NSE and NFM gene expression level were evaluated using the real-time PCR. A detailed study on the interaction between pyrimethamine anticancer drug and (6, 0) zigzag single-walled carbon nanotube was performed by DFT/B3LYP and DFT/M06-2X with 6-31G* basis set calculations in gas phase and in solvent using the PCM. Different configurations of the adsorbed pyrimethamine onto the CNT surface were studied. Based on the results, the process of pyrimethamine adsorption on diff ;erent sites on the outer wall of the nanotube was exothermic and configurations were stable. The adsorption energy values indicated that the pyrimethamine molecule could be physically adsorbed on the external surface of the SWCNT. The QTAIM was used for characterizing the nature of the interactions between the pyrimethamine and the selected nanocarrier.


Assuntos
Tecido Adiposo/citologia , Antineoplásicos/farmacologia , Neurônios/citologia , Pirimetamina/farmacologia , Tecido Adiposo/efeitos dos fármacos , Antineoplásicos/química , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Células MCF-7 , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Conformação Molecular , Nanotubos de Carbono , Proteínas de Neurofilamentos/genética , Neurônios/efeitos dos fármacos , Fosfopiruvato Hidratase/genética , Pirimetamina/química
11.
Vet Res ; 50(1): 106, 2019 Dec 05.
Artigo em Inglês | MEDLINE | ID: mdl-31806006

RESUMO

The binding and activation of host plasminogen (PLG) by worm surface enolases has been verified to participate in parasite invasion, but the role of this processes during Trichinella spiralis infection has not been clarified. Therefore, the expression and immunolocalization of a T. spiralis enolase (TsENO) and its binding activity with PLG were evaluated in this study. Based on the three-dimensional (3D) molecular model of TsENO, the protein interaction between TsENO and human PLG was analysed by the ZDOCK server. The interacting residues were identified after analysis of the protein-protein interface by bioinformatics techniques. The key interacting residues were confirmed by a series of experiments. The qPCR analysis results demonstrated that Ts-eno was transcribed throughout the whole life cycle of T. spiralis. The immunofluorescence assay (IFA) results confirmed that TsENO was distributed on the T. spiralis surface. The binding assays showed that recombinant TsENO (rTsENO) and native TsENO were able to bind PLG. Four lysine residues (90, 289, 291 and 300) of TsENO were considered to be active residues for PLG interaction. The quadruple mutant (Lys90Ala + Lys289Ala + Lys291Ala + Lys300Ala) TsENO, in which the key lysine residues were substituted with alanine (Ala) residues, exhibited a reduction in PLG binding of nearly 50% (45.37%). These results revealed that TsENO has strong binding activity with human PLG. The four lysine residues (90, 289, 291 and 300) of TsENO play an important role in PLG binding and could accelerate PLG activation and invasion of the host's intestinal wall by T. spiralis.


Assuntos
Proteínas de Helminto/genética , Fosfopiruvato Hidratase/genética , Plasminogênio/fisiologia , Trichinella spiralis/fisiologia , Triquinelose/imunologia , Animais , Feminino , Proteínas de Helminto/metabolismo , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Fosfopiruvato Hidratase/metabolismo , Trichinella spiralis/genética , Triquinelose/parasitologia
12.
Mol Cells ; 42(11): 804-809, 2019 Nov 30.
Artigo em Inglês | MEDLINE | ID: mdl-31697874

RESUMO

Oncogenic gain-of-function mutations are clinical biomarkers for most targeted therapies, as well as represent direct targets for drug treatment. Although loss-of-function mutations involving the tumor suppressor gene, STK11 (LKB1) are important in lung cancer progression, STK11 is not the direct target for anticancer agents. We attempted to identify cancer transcriptome signatures associated with STK11 loss-offunction mutations. Several new sensitive and specific gene expression markers (ENO3, TTC39C, LGALS3, and MAML2) were identified using two orthogonal measures, i.e., fold change and odds ratio analyses of transcriptome data from cell lines and tissue samples. Among the markers identified, the ENO3 gene over-expression was found to be the direct consequence of STK11 loss-of-function. Furthermore, the knockdown of ENO3 expression exhibited selective anticancer effect in STK11 mutant cells compared with STK11 wild type (or recovered) cells. These findings suggest that ENO3 -based targeted therapy might be promising for patients with lung cancer harboring STK11 mutations.


Assuntos
Adenocarcinoma/genética , Mutação com Ganho de Função , Perfilação da Expressão Gênica/métodos , Regulação Neoplásica da Expressão Gênica , Neoplasias Pulmonares/genética , Fosfopiruvato Hidratase/genética , Proteínas Serina-Treonina Quinases/genética , Células A549 , Adenocarcinoma/patologia , Biomarcadores Tumorais/genética , Linhagem Celular Tumoral , Proliferação de Células/genética , Sobrevivência Celular/genética , Humanos , Neoplasias Pulmonares/patologia , Interferência de RNA
13.
Sci Rep ; 9(1): 17257, 2019 11 21.
Artigo em Inglês | MEDLINE | ID: mdl-31754158

RESUMO

Rapid modulation of RNA function by endoribonucleases during physiological responses to environmental changes is known to be an effective bacterial biochemical adaptation. We report a molecular mechanism underlying the regulation of enolase (eno) expression by two endoribonucleases, RNase G and RNase III, the expression levels of which are modulated by oxygen availability in Escherichia coli. Analyses of transcriptional eno-cat fusion constructs strongly suggested the existence of cis-acting elements in the eno 5' untranslated region that respond to RNase III and RNase G cellular concentrations. Primer extension and S1 nuclease mapping analyses of eno mRNA in vivo identified three eno mRNA transcripts that are generated in a manner dependent on RNase III expression, one of which was found to accumulate in rng-deleted cells. Moreover, our data suggested that RNase III-mediated cleavage of primary eno mRNA transcripts enhanced Eno protein production, a process that involved putative cis-antisense RNA. We found that decreased RNase G protein abundance coincided with enhanced RNase III expression in E. coli grown anaerobically, leading to enhanced eno expression. Thereby, this posttranscriptional up-regulation of eno expression helps E. coli cells adjust their physiological reactions to oxygen-deficient metabolic modes. Our results revealed a molecular network of coordinated endoribonuclease activity that post-transcriptionally modulates the expression of Eno, a key enzyme in glycolysis.


Assuntos
Endorribonucleases/metabolismo , Proteínas de Escherichia coli/metabolismo , Fosfopiruvato Hidratase/genética , Ribonuclease III/metabolismo , Endorribonucleases/genética , Escherichia coli/genética , Proteínas de Escherichia coli/genética , Expressão Gênica/genética , Regulação Bacteriana da Expressão Gênica/genética , Oxigênio/metabolismo , Fosfopiruvato Hidratase/metabolismo , Processamento Pós-Transcricional do RNA/genética , RNA Bacteriano/genética , RNA Mensageiro/genética , Ribonuclease III/genética
14.
Sichuan Da Xue Xue Bao Yi Xue Ban ; 50(3): 373-378, 2019 May.
Artigo em Chinês | MEDLINE | ID: mdl-31631606

RESUMO

Objective: To establish a radiomic model for predicting lymph node (LN) metastasis in patients with non-small cell lung cancer (NSCLC). Methods: The prediction model was developed using a training cohort comprising 100 patients with clinicopathologically confirmed NSCLC. Data were gathered from January 2014 to December 2015. Radiomic features of NSCLC were obtained from non-contrast and enhancement computed tomography (CT). Lasso-logistic regression models were established for data dimension reduction, feature selection, and radiomics signature building. Consistency coefficient ( ICCs) was used to evaluate the consistency between observer interior and interobserver.The consistency index (C-index)is used to evalutate the prediction of lymph node metastasis by using the radiomics signature, shown with the area under the receiver operating characteristic curve ( AUC).Multivariate logistic regression analyses were performed to develop the prediction model, considering radiomics signature and clinicopathologic risk factors. The radiomics model was validated in a validation cohort comprising 100 consecutive NSCLC patients from January 2016 to December 2017 in terms of its calibration and discrimination. AUC was used to evaluate the predictive effectiveness of the model, and Delong test was used to compare models. Hosmer-Lemeshow good of fit test was used to evaluate the calibration of prediction models.The results were represented by correction curves to compare the consistency between the predicted results of the model and the actual probability of LN metastasis. Results: The consistency between observer interior and interobserver was good, with ICCs higher than 0.75.The radiomics signature, including 22 selected features, was associated with LN metastasis. AUC was 0.781 in training cohort and 0.776 in validation cohort. The individualized prediction model identified radiomics signature, neuron specific enolase (CEA), cytokeratin 19 fragment antigen 21-1 (CYFRA21-1), and carbohydrate antigen 125 (CA125) as independent predictors. The model showed good discrimination, with 0.836 AUC in the training cohort, and 0.821 AUC in the validation cohort. The model in both the training and validation cohorts had good calibration,which demonstrated high consistency with the actual LN metastasis. Conclusion: The radiomics model incorporating radiomics signature and clinical risk factors can be conveniently used to facilitate preoperative individualized prediction of LN metastasis in patients with NSCLC.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/patologia , Neoplasias Pulmonares/patologia , Metástase Linfática/diagnóstico , Antígenos de Neoplasias/genética , Antígeno Ca-125/genética , Humanos , Queratina-19/genética , Modelos Logísticos , Linfonodos , Proteínas de Membrana/genética , Fosfopiruvato Hidratase/genética , Valor Preditivo dos Testes , Estudos Retrospectivos
15.
Plant Sci ; 289: 110243, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31623796

RESUMO

Maize (Zea mays) seeds are the major source of starch all over the world and the excellent model for researching starch synthesis. Seed starch content is a typical quantitative phenotype and many reports revealed that the glycolytic enzymes are involved in regulating starch synthesis, however the regulatory mechanism is still unclear. Here, we present a comparative phosphoproteomic study of three maize inbred lines with different seed starch content. It reveals that abundances of 62 proteins and 63 phosphoproteins were regulated during maize seed development. Dynamics of 17 enzymes related to glycolysis and starch synthesis were used to construct a phosphorylation regulatory network of starch synthesis. It shows that starch synthesis and glycolysis in maize seeds utilize the same hexose phosphates pool coming from sorbitol and sucrose as carbon source, and phosphorylation of ZmENO1 are suggested to contribute to increase starch content, because it is positively related to seed starch content in different developmental stages and different lines, and the phosphor-mimic mutant (ZmENO1S43D) damaged its enzyme activity which is vital in glycolysis. Our results provide a new sight into regulatory process of seed starch synthesis and can be used in maize breeding for high starch content.


Assuntos
Regulação da Expressão Gênica de Plantas , Fosfoproteínas/genética , Fosfopiruvato Hidratase/genética , Proteínas de Plantas/genética , Proteoma/genética , Amido/metabolismo , Zea mays/metabolismo , Fosfoproteínas/metabolismo , Fosfopiruvato Hidratase/metabolismo , Proteínas de Plantas/metabolismo , Proteoma/metabolismo , Sementes/crescimento & desenvolvimento , Sementes/metabolismo , Zea mays/enzimologia , Zea mays/genética , Zea mays/crescimento & desenvolvimento
16.
J Biosci ; 44(4)2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31502568

RESUMO

The glycolytic enzyme enolase of Staphylococcus aureus is a highly conserved enzyme which binds to human plasminogen thereby aiding the infection process. The cloning, over expression and purification of S. aureus enolase as well as the effect of various metals upon the catalytic activity and structural stability of the enzyme have been reported. The recombinant enzyme (rSaeno) has been purified to homogeneity in abundant amounts (60 mg/L of culture) and the kinetic parameters (Km = 0.23 +/- 0.013 x 10-3 M; Vmax = 90.98 +/- 0.00052 U/mg) and the optimum pH were calculated. This communication further reports that increasing concentrations of Na+ ions inhibit the enzyme while increasing concentrations of K+ ions were stimulatory. In case of divalent cations, it was found that Mg2+ stimulates the activity of rSaeno while the rest of the divalent cations (Zn2+, Mn2+, Fe2+, Cu2+, Ni2+ and Ca2+) lead to a dose-dependent loss in the activity with a total loss of activity in the presence of Hg2+ and Cr2+. The circular dichroism data indicate that other than Hg2+, Ni2+ and to a certain extent Cu2+, none of the other ions destabilized rSaeno. The inhibitory roles of fluorides, as well as neurotoxic compounds upon the catalytic activity of rSaeno, have also been studied. Conformational changes in rSaeno (induced by ions) were studied using partial trypsin digestion.


Assuntos
Estabilidade Enzimática/efeitos dos fármacos , Metais/farmacologia , Fosfopiruvato Hidratase/genética , Conformação Proteica/efeitos dos fármacos , Catálise/efeitos dos fármacos , Dicroísmo Circular , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Humanos , Íons/química , Íons/farmacologia , Metais/química , Fosfopiruvato Hidratase/química , Fosfopiruvato Hidratase/isolamento & purificação , Staphylococcus aureus/enzimologia , Staphylococcus aureus/patogenicidade
17.
Microb Pathog ; 135: 103651, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31398532

RESUMO

Bartonella infection is distributed worldwide with animal and public health. Recent studies have shown that host cells infection by Bartonella has a series of different infection stages, beginning with encounter and adherence to the cells. In this study, we expressed and purified recombinant Bartonella henselae (B. henselae) α-enolase. And we found that B. henselae α-enolase is highly conserved in Bartonella species. The interacting protein partners of B. henselae α-enolase were showed by String-11. The interactions between B. henselae α-enolase and human plasminogen were subsequently confirmed by ELISA, pull down, T7 phage display and molecular docking assays. And the plasminogen-binding sites of B. henselae α-enolase are predicted at 247FYKNGSYFY255. These findings will help elucidate and improve the understanding of the molecular mechanisms of Bartonella infection.


Assuntos
Bartonella/enzimologia , Bartonella/genética , Proteínas de Transporte/metabolismo , Fosfopiruvato Hidratase/genética , Fosfopiruvato Hidratase/isolamento & purificação , Plasminogênio/metabolismo , Sequência de Aminoácidos , Bartonella henselae/enzimologia , Bartonella henselae/genética , Sítios de Ligação , Proteínas de Transporte/química , Clonagem Molecular , Regulação Bacteriana da Expressão Gênica , Humanos , Modelos Moleculares , Simulação de Acoplamento Molecular , Fosfopiruvato Hidratase/química , Fosfopiruvato Hidratase/classificação , Filogenia , Plasminogênio/química , Proteínas Recombinantes
18.
Biosci Rep ; 39(9)2019 09 30.
Artigo em Inglês | MEDLINE | ID: mdl-31431517

RESUMO

Bladder cancer (BC) is the ninth most common malignancy throughout the world. The molecular mechanisms of this disease remain largely unclear. The glycolytic enzyme enolase 1 (ENO1) has been shown to regulate the development of various cancers. However, the significance of ENO1 in BC is underdetermined. In this study, we found that ENO1 was highly expressed in BC tissues and cells. High expression of ENO1 was associated with the poor survival of BC patients. Using lentivirus-mediated knockdown and over-expression, we revealed that ENO1 was critical for the growth and proliferation of BC cells. ENO1 over-expression also promoted the proliferation of SV-HUC-1 cells. At the molecular level, the cell cycle and apoptosis related genes were regulated by ENO1. ß-catenin expression was positively regulated by ENO1. Furthermore, ectopic expression of ß-catenin reversed the effect of ENO1 knockdown on T24 cell proliferation and growth. Opposite results were observed in ß-catenin knockdown T24 cells. Our findings suggested that ENO1 functioned as an oncogene in BC through regulating cell cycle, apoptosis and ß-catenin. Targeting ENO1/ß-catenin cascade may benefit for BC patients.


Assuntos
Apoptose/genética , Biomarcadores Tumorais/metabolismo , Pontos de Checagem do Ciclo Celular/genética , Proteínas de Ligação a DNA/metabolismo , Fosfopiruvato Hidratase/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Neoplasias da Bexiga Urinária/patologia , beta Catenina/metabolismo , Biomarcadores Tumorais/genética , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Proteínas de Ligação a DNA/genética , Humanos , Fosfopiruvato Hidratase/genética , Interferência de RNA , RNA Interferente Pequeno/genética , Proteínas Supressoras de Tumor/genética , Regulação para Cima , Neoplasias da Bexiga Urinária/mortalidade
19.
Int J Mol Sci ; 20(16)2019 Aug 14.
Artigo em Inglês | MEDLINE | ID: mdl-31416219

RESUMO

Breast cancer is a complex and heterogeneous disease: Several molecular alterations cause cell proliferation and the acquisition of an invasive phenotype. Extracellular matrix (ECM) is considered essential for sustaining tumor growth and matrix metalloproteinases (MMPs) have been identified as drivers of many aspects of the tumor phenotype. Mounting evidence indicates that both α-enolase (ENO1) and Myc promoter-binding protein-1 (MBP-1) also played pivotal roles in tumorigenesis, although as antagonists. ENO1 is involved in cell growth, hypoxia tolerance and autoimmune activities besides its major role in the glycolysis pathway. On the contrary, MBP-1, an alternative product of ENO1, suppresses cell proliferation and the invasive ability of cancer cells. Since an important task in personalized medicine is to discriminate a different subtype of patients with different clinical outcomes including chances of recurrence and metastasis, we investigated the functional relationship between ENO1/MBP-1 expression and MMP-2 and MMP-9 activity levels in both tissues and sera of breast cancer patients. We focused on the clinical relevance of ENO1 and MMPs (MMP-2 and MMP-9) overexpression in breast cancer tissues: The association between the higher ENO1, MMP-2 and MMP-9 expression with a worse prognosis suggest that the elevated ENO1 and MMPs expression are promising biomarkers for breast cancer. A relationship seems to exist between MBP-1 expression and the decrease in the activity levels of MMP-9 in cancer tissues and MMP-2 in sera. Moreover, the sera of breast cancer patients grouped for MBP-1 expression differentially induced, in vitro, cell proliferation and migration. Our findings support the hypothesis of patient's stratification based on ENO1, MBP-1 and MMPs expression. Elucidating the molecular pathways through which MBP-1 influences MMPs expression and breast cancer regression can lead to the discovery of new management strategies.


Assuntos
Biomarcadores Tumorais/genética , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Proteínas de Ligação a DNA/genética , Regulação Neoplásica da Expressão Gênica , Metaloproteinase 2 da Matriz/genética , Metaloproteinase 9 da Matriz/genética , Fosfopiruvato Hidratase/genética , Proteínas Supressoras de Tumor/genética , Neoplasias da Mama/metabolismo , Neoplasias da Mama/mortalidade , Linhagem Celular Tumoral , Proliferação de Células , Progressão da Doença , Feminino , Humanos , Estimativa de Kaplan-Meier , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/metabolismo
20.
Artigo em Inglês | MEDLINE | ID: mdl-31263685

RESUMO

Enolase is an evolutionarily conserved enzyme involved in the processes of glycolysis and gluconeogenesis. Mycoplasma hyopneumoniae belongs to Mycoplasma, whose species are wall-less and among the smallest self-replicating bacteria, and is an important colonizing respiratory pathogen in the pig industry worldwide. Mycoplasma hyopneumoniae enolase (Mhp Eno) expression is significantly increased after infection and was previously found to be a virulence factor candidate. Our studies show that Mhp Eno is a cell surface-localized protein that can adhere to swine tracheal epithelial cells (STECs). Adhesion to STECs can be specifically inhibited by an Mhp Eno antibody. Mhp Eno can recognize and interact with plasminogen with high affinity. Here, the first crystal structure of the mycoplasmal enolase from Mycoplasma hyopneumoniae was determined. The structure showed unique features of Mhp Eno in the S3/H1, H6/S6, H7/H8, and H13 regions. All of these regions were longer than those of other enolases and were exposed on the Mhp Eno surface, making them accessible to host molecules. These results show that Mhp Eno has specific structural characteristics and acts as a multifunctional adhesin on the Mycoplasma hyopneumoniae cell surface.


Assuntos
Adesinas Bacterianas/química , Adesinas Bacterianas/metabolismo , Membrana Celular/metabolismo , Mycoplasma hyopneumoniae/enzimologia , Fosfopiruvato Hidratase/química , Fosfopiruvato Hidratase/metabolismo , Adesinas Bacterianas/genética , Adesinas Bacterianas/isolamento & purificação , Animais , Cristalografia por Raios X , Células Epiteliais/microbiologia , Modelos Moleculares , Mycoplasma hyopneumoniae/metabolismo , Mycoplasma hyopneumoniae/patogenicidade , Fosfopiruvato Hidratase/genética , Fosfopiruvato Hidratase/isolamento & purificação , Plasminogênio/metabolismo , Pneumonia Suína Micoplasmática/microbiologia , Conformação Proteica , Alinhamento de Sequência , Análise de Sequência de Proteína , Especificidade da Espécie , Ressonância de Plasmônio de Superfície , Suínos , Fatores de Virulência
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...