RESUMO
Some cyanobacteria can grow photoautotrophically or photomixotrophically by using simultaneously CO2 and glucose. The switch between these trophic modes and the role of glycogen, their main carbon storage macromolecule, was investigated. We analysed the effect of glucose addition on the physiology, metabolic and photosynthetic state of Synechocystis sp. PCC 6803 and mutants lacking phosphoglucomutase and ADP-glucose pyrophosphorylase, with limitations in glycogen synthesis. Glycogen acted as a metabolic buffer: glucose addition increased growth and glycogen reserves in the wild-type (WT), but arrested growth in the glycogen synthesis mutants. Already 30 min after glucose addition, metabolites from the Calvin-Benson-Bassham cycle and the oxidative pentose phosphate shunt increased threefold more in the glycogen synthesis mutants than the WT. These alterations substantially affected the photosynthetic performance of the glycogen synthesis mutants, as O2 evolution and CO2 uptake were both impaired. We conclude that glycogen synthesis is essential during transitions to photomixotrophy to avoid metabolic imbalance that induces inhibition of electron transfer from PSII and subsequently accumulation of reactive oxygen species, loss of PSII core proteins, and cell death. Our study lays foundations for optimising photomixotrophy-based biotechnologies through understanding the coordination of the crosstalk between photosynthetic electron transport and metabolism.
Assuntos
Glicogênio , Fotossíntese , Complexo de Proteína do Fotossistema II , Synechocystis , Synechocystis/metabolismo , Synechocystis/efeitos dos fármacos , Synechocystis/crescimento & desenvolvimento , Synechocystis/genética , Glicogênio/metabolismo , Transporte de Elétrons , Complexo de Proteína do Fotossistema II/metabolismo , Mutação/genética , Glucose/metabolismo , Dióxido de Carbono/metabolismo , Oxigênio/metabolismo , Glucose-1-Fosfato Adenililtransferase/metabolismo , Glucose-1-Fosfato Adenililtransferase/genética , Fosfoglucomutase/metabolismo , Fosfoglucomutase/genéticaRESUMO
Pneumocystis cyst life forms contain abundant ß-glucan carbohydrates, synthesized using ß-1,3 and ß-1,6 glucan synthase enzymes and the donor uridine diphosphate (UDP)-glucose. In yeast, phosphoglucomutase (PGM) plays a crucial role in carbohydrate metabolism by interconverting glucose 1-phosphate and glucose 6-phosphate, a vital step in UDP pools for ß-glucan cell wall formation. This pathway has not yet been defined in Pneumocystis. Herein, we surveyed the Pneumocystis jirovecii and Pneumocystis murina genomes, which predicted a homolog of the Saccharomyces cerevisiae major PGM enzyme. Furthermore, we show that PjPgm2p and PmPgm2p function similarly to the yeast counterpart. When both Pneumocystis pgm2 homologs are heterologously expressed in S. cerevisiae pgm2Δ cells, both genes can restore growth and sedimentation rates to wild-type levels. Additionally, we demonstrate that yeast pgm2Δ cell lysates expressing the two Pneumocystis pgm2 transcripts individually can restore PGM activities significantly altered in the yeast pgm2Δ strain. The addition of lithium, a competitive inhibitor of yeast PGM activity, significantly reduces PGM activity. Next, we tested the effects of lithium on P. murina viability ex vivo and found the compound displays significant anti-Pneumocystis activity. Finally, we demonstrate that a para-aryl derivative (ISFP10) with known inhibitory activity against the Aspergillus fumigatus PGM protein and exhibiting 50-fold selectivity over the human PGM enzyme homolog can also significantly reduce Pmpgm2 activity in vitro. Collectively, our data genetically and functionally validate phosphoglucomutases in both P. jirovecii and P. murina and suggest the potential of this protein as a selective therapeutic target for individuals with Pneumocystis pneumonia.
Assuntos
Pneumocystis carinii , Pneumocystis , Pneumonia por Pneumocystis , beta-Glucanas , Humanos , Pneumocystis carinii/genética , Pneumonia por Pneumocystis/tratamento farmacológico , Fosfoglucomutase/genética , Fosfoglucomutase/metabolismo , Fosfoglucomutase/farmacologia , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Lítio/metabolismo , Lítio/farmacologia , Pneumocystis/genética , beta-Glucanas/metabolismo , Fosfatos/farmacologia , Glucose/metabolismo , Difosfato de Uridina/metabolismo , Difosfato de Uridina/farmacologiaRESUMO
Hyper-IgE Syndrome (HIES) is a heterogeneous group of primary immune-deficiency disorders characterized by elevated levels of IgE, eczema, and recurrent skin and lung infections. HIES that is autosomally dominant in the signal transducer and activator of transcription 3 (STAT3), and autosomal recessive mutations in phosphoglucomutase 3 (PGM3) have been reported in humans. An early diagnosis, based on clinical suspicion and immunological assessments, is challenging. Patients' metabolomics, proteomics, and cytokine profiles were compared to DOCK 8-deficient and atopic dermatitis patients. The PGM3 metabolomics profile identified significant dysregulation in hypotaurine, hypoxanthine, uridine, and ribothymidine. The eight proteins involved include bifunctional arginine demethylase and lysyl hydroxylase (JMJD1B), type 1 protein phosphatase inhibitor 4 (PPI 4), and platelet factor 4 which aligned with an increased level of the cytokine GCSF. Patients with STAT3 deficiency, on the other hand, showed significant dysregulation in eight metabolites, including an increase in protocatechuic acid, seven proteins including ceruloplasmin, and a plasma protease C1 inhibitor, in addition to cytokine VEGF being dysregulated. Using multi-omics profiling, we identified the dysregulation of endothelial growth factor (EGFR) and tumor necrosis factor (TNF) signaling pathways in PGM3 and STAT3 patients, respectively. Our findings may serve as a stepping stone for larger prospective HIES clinical cohorts to validate their future use as biomarkers.
Assuntos
Imunoglobulina E , Síndrome de Job , Humanos , Fosfoglucomutase/metabolismo , Fator de Transcrição STAT3/metabolismo , Multiômica , Estudos Prospectivos , Síndrome de Job/diagnóstico , Mutação , Citocinas/metabolismoRESUMO
The hexosamine biosynthetic pathway (HBP) is connected to abnormal N- and O-linked protein glycosylation in cancer, which performs critical roles in tumorigenesis. However, the regulation mechanisms of HBP and its role in colorectal cancer (CRC) progression remain unexplained. This study analyzed the expression level of phosphoglucomutase 3 (PGM3), a key enzyme in HBP, and identified its function in CRC cell lines. Analysis of publicly available CRC microarray data determined that PGM3 is upregulated in CRC tumor tissues. Furthermore, functional experiments emphasized the significant roles of PGM3 in facilitating CRC cell proliferation and migration. Mechanistically, we demonstrated that the activity of ß-catenin in CRC was maintained by PGM3-mediated O-GlcNAcylation. PGM3 knockdown or inhibition of O-GlcNAc transferase decreased ß-catenin activity and the expression levels of its downstream targets. Collectively, our findings indicate that PGM3 exhibits tumor-promoting roles by elevating O-GlcNAcylation level and maintaining ß-catenin activity, and might serve as a prognostic biomarker and treatment target in CRC.
Assuntos
Neoplasias Colorretais , Fosfoglucomutase , beta Catenina , Biomarcadores , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Transformação Celular Neoplásica , Neoplasias Colorretais/patologia , Regulação Neoplásica da Expressão Gênica , Hexosaminas/metabolismo , Humanos , Fosfoglucomutase/metabolismo , Via de Sinalização Wnt , beta Catenina/metabolismoRESUMO
OBJECTIVE: Toxoplasma gondii is a ubiquitous parasite of medical and veterinary importance; however, there exists no cure for chronic toxoplasmosis. Metabolic enzymes required for the production and maintenance of tissue cysts represent promising targets for novel therapies. Here, we use reverse genetics to investigate the role of Toxoplasma phosphoglucomutase 1, PGM1, in Toxoplasma growth and cystogenesis. RESULTS: We found that disruption of pgm1 did not significantly affect Toxoplasma intracellular growth and the lytic cycle. pgm1-defective parasites could differentiate into bradyzoites and produced cysts containing amylopectin in vitro. However, cysts produced in the absence of pgm1 were significantly smaller than wildtype. Together, our findings suggest that PGM1 is dispensable for in vitro growth but contributes to optimal Toxoplasma cyst development in vitro, thereby necessitating further investigation into the function of this enzyme in Toxoplasma persistence in its host.
Assuntos
Fosfoglucomutase , Toxoplasma , Toxoplasmose , Humanos , Fosfoglucomutase/genética , Fosfoglucomutase/metabolismo , Toxoplasma/enzimologia , Toxoplasma/genética , Toxoplasmose/parasitologiaRESUMO
Aspergillus fumigatus is the causative agent of invasive aspergillosis, an infection with mortality rates of up to 50%. The glucan-rich cell wall of A. fumigatus is a protective structure that is absent from human cells and is a potential target for antifungal treatments. Glucan is synthesized from the donor uridine diphosphate glucose, with the conversion of glucose-6-phosphate to glucose-1-phosphate by the enzyme phosphoglucomutase (PGM) representing a key step in its biosynthesis. Here, we explore the possibility of selectively targeting A. fumigatus PGM (AfPGM) as an antifungal treatment strategy. Using a promoter replacement strategy, we constructed a conditional pgm mutant and revealed that pgm is required for A. fumigatus growth and cell wall integrity. In addition, using a fragment screen, we identified the thiol-reactive compound isothiazolone fragment of PGM as targeting a cysteine residue not conserved in the human ortholog. Furthermore, through scaffold exploration, we synthesized a para-aryl derivative (ISFP10) and demonstrated that it inhibits AfPGM with an IC50 of 2 µM and exhibits 50-fold selectivity over the human enzyme. Taken together, our data provide genetic validation of PGM as a therapeutic target and suggest new avenues for inhibiting AfPGM using covalent inhibitors that could serve as tools for chemical validation.
Assuntos
Aspergilose , Aspergillus fumigatus , Antifúngicos/farmacologia , Aspergilose/tratamento farmacológico , Aspergilose/microbiologia , Aspergillus fumigatus/enzimologia , Aspergillus fumigatus/genética , Glucanos/metabolismo , Humanos , Fosfoglucomutase/genética , Fosfoglucomutase/metabolismoRESUMO
In non-small-cell lung cancer (NSCLC), concurrent mutations in the oncogene KRAS and tumor suppressor STK11 (also known as LKB1) confer an aggressive malignant phenotype, an unfavourability towards immunotherapy, and overall poor prognoses in patients. In a previous study, we showed that murine KRAS/LKB1 co-mutant tumors and human co-mutant cancer cells have an enhanced dependence on glutamine-fructose-6-phosphate transaminase 2 (GFPT2), a rate-limiting enzyme in the hexosamine biosynthesis pathway (HBP), which could be targeted to reduce survival of KRAS/LKB1 co-mutants. Here, we found that KRAS/LKB1 co-mutant cells also exhibit an increased dependence on N-acetylglucosamine-phosphate mutase 3 (PGM3), an enzyme downstream of GFPT2. Genetic or pharmacologic suppression of PGM3 reduced KRAS/LKB1 co-mutant tumor growth in both in vitro and in vivo settings. Our results define an additional metabolic vulnerability in KRAS/LKB1 co-mutant tumors to the HBP and provide a rationale for targeting PGM3 in this aggressive subtype of NSCLC.
Assuntos
Quinases Proteína-Quinases Ativadas por AMP/genética , Neoplasias Pulmonares/genética , Terapia de Alvo Molecular , Fosfoglucomutase/metabolismo , Proteínas Proto-Oncogênicas p21(ras)/genética , Animais , Vias Biossintéticas/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Glutamina-Frutose-6-Fosfato Transaminase (Isomerizante)/antagonistas & inibidores , Glutamina-Frutose-6-Fosfato Transaminase (Isomerizante)/metabolismo , Glicosilação/efeitos dos fármacos , Hexosaminas/biossíntese , Humanos , Neoplasias Pulmonares/patologia , Camundongos , Fosfoglucomutase/antagonistas & inibidores , Fosfoglucomutase/genéticaRESUMO
Background: Bioinformatics showed that long non-coding RNA (lncRNA) pgm5-as1 was regulated in patients with colorectal cancer (CRC), and miR-484 was also regulated in CRC. We aimed at determining the modulatory pathway of lncRNA pgm5-as1 in CRC cells, and whether miR-484 was involved in the pathway. Materials and Methods: The target gene of pgm5-as1 was predicted by bioinformatics and verified by dual luciferase assay. Transcription levels of pgm5-as1 and miR-484 were determined by quantitative real-time polymerase chain reaction. Viability, migration rate, invasion, and growth of SW480 and HCT116 cells were determined by Cell Counting Kit-8 (CCK-8), wound healing assay, transwell, and colony formation assay, respectively. Results: pgm5-as1 was upregulated in CRC tissues and cell lines; however, its downregulation contributed to the decreasing of cell viability, growth, migration, and invasion of SW480 and HCT116 cells. Moreover, miR-484 was predicted as the target of pgm5-as1, and the downregulation of pgm5-as1 partially restored the elevated cell viability, growth, migration, and invasion that were induced by the inhibition of miR-484 expression in SW480 and HCT116 cells. Conclusions: The loss of miR-484 expression in CRC might be involved in the promotion and metastasis of CRC, which may be caused by the overexpression of pgm5-as1. Hence, the downregulation of pgm5-as1 could be a therapeutic target in the prevention or intervention of CRC.
Assuntos
Neoplasias Colorretais/metabolismo , Proteínas do Citoesqueleto/genética , MicroRNAs/metabolismo , Fosfoglucomutase/genética , RNA Longo não Codificante/metabolismo , Proliferação de Células/fisiologia , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Proteínas do Citoesqueleto/metabolismo , Regulação para Baixo , Humanos , MicroRNAs/biossíntese , MicroRNAs/genética , Metástase Neoplásica , Fosfoglucomutase/metabolismo , RNA Longo não Codificante/genéticaRESUMO
Cells lining the uterus are responsible for storage and secretion of carbohydrates to support early embryonic development. Histotrophic secretions contain glycogen and glycolytic products such as lactate and pyruvate. Insufficient carbohydrate storage as glycogen has been correlated with infertility in women. While it is clear that changes in estrogen (17-ß-estradiol (E2)) and progesterone (P4) in vivo affect the distribution of glucose in the uterine cells and secretions, the biochemical mechanism(s) by which they affect this crucial allocation is not well understood. Furthermore, in cultured uterine cells, neither E2 nor P4 affect glycogen storage without insulin present. We hypothesized that P4 and E2 alone affect the activity of glycolytic enzymes, glucose and glycolytic flux to increase glycogen storage (E2) and catabolism (P4) and increase pyruvate and lactate levels in culture. We measured the rate of glucose uptake and glycolysis in a mink immortalized epithelial cell line (GMMe) after 24-h exposure to 10 µM P4 and 10 nM E2 (pharmacologic levels) at 5 mM glucose and determined the kinetic parameters (Vmax, Km) of all enzymes. While the activities of many glycolytic enzymes in GMMe cells were shown to be decreased by E2 treatment, in contrast, glucose uptake, glycolytic flux and metabolites levels were not affected by the treatments. The cellular rationale for P4- and E2-induced decreases in the activity of enzymes may be to prime the system for other regulators such as insulin. In vivo, E2 and P4 may be necessary but not sufficient signals for uterine cycle carbohydrate allocation.
Assuntos
Estradiol/metabolismo , Ciclo Estral/metabolismo , Glucose/metabolismo , Progesterona/metabolismo , Útero/metabolismo , Animais , Linhagem Celular , Ensaios Enzimáticos , Células Epiteliais , Feminino , Glucosefosfato Desidrogenase/metabolismo , Glicogênio/metabolismo , Glicólise/fisiologia , Cinética , Vison , Modelos Animais , Fosfoglucomutase/metabolismoRESUMO
The second messenger cyclic di-AMP (c-di-AMP) is essential for growth of many bacteria because it controls osmolyte homeostasis. c-di-AMP can regulate the synthesis of potassium uptake systems in some bacteria and it also directly inhibits and activates potassium import and export systems, respectively. Therefore, c-di-AMP production and degradation have to be tightly regulated depending on the environmental osmolarity. The Gram-positive pathogen Listeria monocytogenes relies on the membrane-bound diadenylate cyclase CdaA for c-di-AMP production and degrades the nucleotide with two phosphodiesterases. While the enzymes producing and degrading the dinucleotide have been reasonably well examined, the regulation of c-di-AMP production is not well understood yet. Here we demonstrate that the extracytoplasmic regulator CdaR interacts with CdaA via its transmembrane helix to modulate c-di-AMP production. Moreover, we show that the phosphoglucosamine mutase GlmM forms a complex with CdaA and inhibits the diadenylate cyclase activity in vitro. We also found that GlmM inhibits c-di-AMP production in L. monocytogenes when the bacteria encounter osmotic stress. Thus, GlmM is the major factor controlling the activity of CdaA in vivo. GlmM can be assigned to the class of moonlighting proteins because it is active in metabolism and adjusts the cellular turgor depending on environmental osmolarity.
Assuntos
Proteínas de Bactérias/metabolismo , AMP Cíclico/biossíntese , Listeria monocytogenes/fisiologia , Fosfoglucomutase/metabolismo , Fosfatos de Dinucleosídeos/metabolismo , Homeostase , Listeria monocytogenes/enzimologia , Pressão Osmótica/fisiologiaRESUMO
Cancer cells undergo metabolic reprogramming to sustain their own survival under an environment of increased energy demand; however, the mechanism by which cancer cells ensure survival under glucose deprivation stressed conditions remains elusive. Here, we show that deprivation of glucose, dramatically activated the glycogen pathway, accompanied by elevated phosphoglucomutase 1 (PGM1) expression. We further identified that AMP-activated protein kinase (AMPK) stimulated PGM1 expression by inducing histone deacetylase 8 (HDAC8) phosphorylation. Moreover, we demonstrated that glucose deprivation-induced AMPK activation stimulated the translocation of HDAC8 from the nucleus to the cytoplasm, consequently disrupting the binding between HDAC8 and histone 3. PGM1 expression was also found to be critical for lung cancer glycolysis, the oxidative pentose phosphate pathway, and oxidative phosphorylation under glucose deprivation conditions, and further led to the aberrant expression of metabolic enzymes involved in glucose metabolism mediated by ERK1/2. Finally, PGM1 was found to be highly expressed in lung cancer tissues from patients, which correlated with a poor prognosis. Taken together, these results revealed that AMPK activation by glucose deprivation leads to enhanced PGM1 expression, an essential component of the metabolic switch, to facilitate cancer progression, suggesting PGM1 as promising anti-cancer treatment targets.
Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Glucose/metabolismo , Histona Desacetilases/metabolismo , Neoplasias Pulmonares/metabolismo , Fosfoglucomutase/genética , Fosfoglucomutase/metabolismo , Proteínas Repressoras/metabolismo , Células A549 , Linhagem Celular Tumoral , Núcleo Celular/metabolismo , Sobrevivência Celular , Citoplasma/metabolismo , Regulação Neoplásica da Expressão Gênica , Glicólise , Células HEK293 , Humanos , Neoplasias Pulmonares/genética , Fosforilação , Transdução de Sinais , Regulação para CimaRESUMO
OBJECTIVE: Enzymatic fingerprinting of key enzymes of glucose metabolism is a valuable analysis tool in cell physiological phenotyping of plant samples. Yet, a similar approach for mammalian cell line samples is missing. In this study, we applied semi-high throughput enzyme activity assays that were originally designed for plant samples and tested their feasibility in extracts of six frequently used mammalian cell lines (Caco2, HaCaT, C2C12, HEK293, HepG2 and INS-1E). RESULTS: Enzyme activities for aldolase, hexokinase, glucose-6-phosphate dehydrogenase, phosphoglucoisomerase, phosphoglucomutase, phosphofructokinase could be detected in samples of one or more mammalian cell lines. We characterized effects of sample dilution, assay temperature and repeated freeze-thaw cycles causing potential biases. After careful selection of experimental parameters, the presented semi-high throughput methods could be established as useful tool for physiological phenotyping of cultured mammalian cells.
Assuntos
Metabolismo dos Carboidratos , Ensaios Enzimáticos/métodos , Glucose/metabolismo , Glicólise , Animais , Células CACO-2 , Linhagem Celular , Linhagem Celular Tumoral , Estudos de Viabilidade , Frutose-Bifosfato Aldolase/metabolismo , Glucosefosfato Desidrogenase/metabolismo , Células HEK293 , Células Hep G2 , Hexoquinase/metabolismo , Humanos , Camundongos , Fenótipo , Fosfofrutoquinases/metabolismo , Fosfoglucomutase/metabolismo , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo , Projetos PilotoRESUMO
c-di-AMP is an important second messenger molecule that plays a pivotal role in regulating fundamental cellular processes, including osmotic and cell wall homeostasis in many Gram-positive organisms. In the opportunistic human pathogen Staphylococcus aureus, c-di-AMP is produced by the membrane-anchored DacA enzyme. Inactivation of this enzyme leads to a growth arrest under standard laboratory growth conditions and a re-sensitization of methicillin-resistant S. aureus (MRSA) strains to ß-lactam antibiotics. The gene coding for DacA is part of the conserved three-gene dacA/ybbR/glmM operon that also encodes the proposed DacA regulator YbbR and the essential phosphoglucosamine mutase GlmM, which is required for the production of glucosamine-1-phosphate, an early intermediate of peptidoglycan synthesis. These three proteins are thought to form a complex in vivo and, in this manner, help to fine-tune the cellular c-di-AMP levels. To further characterize this important regulatory complex, we conducted a comprehensive structural and functional analysis of the S. aureus DacA and GlmM enzymes by determining the structures of the S. aureus GlmM enzyme and the catalytic domain of DacA. Both proteins were found to be dimers in solution as well as in the crystal structures. Further site-directed mutagenesis, structural and enzymatic studies showed that multiple DacA dimers need to interact for enzymatic activity. We also show that DacA and GlmM form a stable complex in vitro and that S. aureus GlmM, but not Escherichia coli or Pseudomonas aeruginosa GlmM, acts as a strong inhibitor of DacA function without the requirement of any additional cellular factor. Based on Small Angle X-ray Scattering (SAXS) data, a model of the complex revealed that GlmM likely inhibits DacA by masking the active site of the cyclase and preventing higher oligomer formation. Together these results provide an important mechanistic insight into how c-di-AMP production can be regulated in the cell.
Assuntos
Inibidores de Adenilil Ciclases/metabolismo , Adenilil Ciclases/metabolismo , Adenilil Ciclases/ultraestrutura , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Parede Celular/metabolismo , Fosfatos de Dinucleosídeos/antagonistas & inibidores , Fosfatos de Dinucleosídeos/metabolismo , Staphylococcus aureus Resistente à Meticilina/genética , Staphylococcus aureus Resistente à Meticilina/metabolismo , Óperon/genética , Fosfoglucomutase/metabolismo , Fósforo-Oxigênio Liases/metabolismo , Domínios Proteicos , Espalhamento a Baixo Ângulo , Sistemas do Segundo Mensageiro/genética , Infecções Estafilocócicas/genética , Staphylococcus aureus/metabolismo , Staphylococcus aureus/fisiologia , Difração de Raios X/métodosRESUMO
Helicobacter pylori growth requirements is a prerequisite to invade gastric epithelium and the process of injury to gastric cells will eventually lead to gastric cancer. The aim of this study is to investigate the effect of iron challenge on the expression of genes involved in iron homeostasis. The presence of Phosphoglucosamine mutase (glmM), cytotoxin-associated gene A (cagA) and vacuolating cytotoxin A (vacA) genes and mRNA expression of Iron Regulatory Protein 2 (IRP2), Transferrin Receptor (TFRC) and Ferritin Light Chain (FTL) genes in samples of 28 normal gastric mucosa, 33 chronic gastritis, 29 gastritis with intestinal metaplasia, 29 intestinal type adenocarcinoma patients were examined by real-time PCR. Immunohistochemistry was used to analyze cellular localization and protein levels. In the all H. pylori positive tissues, particularly in the basal regions of foveolar cells, TFRC was overexpressed (Pâ¯<â¯0.05), and regardless of the H. pylori infection, FTL was overexpressed in all patient, exclusively in metaplastic glandular cells (Pâ¯<â¯0.05). Furthermore, overexpression of IRP2 was associated with H. pylori positive chronic gastritis and intestinal metaplasia (Pâ¯<â¯0.05). Our findings confirm the role of transferrin receptor in H. pylori attachment into the gastric mucosa to capture iron. Overexpression of FTL gene in metaplastic cells could be considered as a research background to investigate the role of this gene in the differentiation of gastric cells into intestinal metaplasia. In addition, this gene could be suggested as a diagnostic marker to be included among the other markers routinely performed by clinical diagnostic laboratories.
Assuntos
Apoferritinas/metabolismo , Biomarcadores , Infecções por Helicobacter/metabolismo , Helicobacter pylori/metabolismo , Helicobacter pylori/patogenicidade , Metaplasia/metabolismo , Receptores da Transferrina/metabolismo , Adenocarcinoma , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Antígenos de Bactérias/genética , Antígenos de Bactérias/metabolismo , Apoferritinas/genética , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Feminino , Mucosa Gástrica/metabolismo , Mucosa Gástrica/patologia , Gastrite/patologia , Gastrite Atrófica/patologia , Expressão Gênica , Infecções por Helicobacter/patologia , Humanos , Ferro/metabolismo , Proteína 2 Reguladora do Ferro/genética , Proteína 2 Reguladora do Ferro/metabolismo , Masculino , Metaplasia/diagnóstico , Pessoa de Meia-Idade , Fosfoglucomutase/genética , Fosfoglucomutase/metabolismo , RNA Mensageiro/biossíntese , Neoplasias Gástricas/patologia , Adulto JovemRESUMO
BACKGROUND: Heart injury is one of the most important concerns after exposure to a high dose of radiation in chest cancer radiotherapy or whole body exposure to a radiation disaster. Studies have proposed that increased level of inflammatory and pro-fibrotic cytokines following radiotherapy or radiation events play a key role in the development of several side effects such as cardiovascular disorders. In the current study, we aimed to evaluate the expression of IL-4 and IL-13 cytokines as well as signaling pathways such as IL4Ra1, IL13Ra2, Duox1 and Duox2. In addition, we detected the possible protective effect of curcumin on the expression of these factors and infiltration of inflammatory cells. MATERIALS AND METHODS: Twenty rats were divided into 4 groups including control; curcumin treated; radiation; and radiation plus curcumin. After 10 weeks, rats were sacrificed for evaluation of mentioned parameters. RESULTS: Results showed an increase in the level of IL-4 and all evaluated genes, as well as increased infiltration of lymphocytes and macrophages. Treatment with curcumin could attenuate these changes. CONCLUSION: Curcumin could reduce radiation-induced heart injury markers in rats.
Assuntos
Curcumina/farmacologia , Coração/efeitos dos fármacos , Coração/efeitos da radiação , Protetores contra Radiação/farmacologia , Animais , Oxidases Duais/metabolismo , Ensaio de Imunoadsorção Enzimática , Interleucina-13/metabolismo , Interleucina-4/metabolismo , Fosfoglucomutase/metabolismo , Radiação Ionizante , Ratos , Reação em Cadeia da Polimerase em Tempo Real , Transdução de SinaisRESUMO
Glycogen metabolism commonly altered in cancer is just beginning to be understood. Phosphoglucomutase 1 (PGM1), the first enzyme in glycogenesis that catalyzes the reversible conversion between glucose 1-phosphate (G-1-P) and glucose 6-phosphate (G-6-P), participates in both the breakdown and synthesis of glycogen. Here, we show that PGM1 is down-regulated in hepatocellular carcinoma (HCC), which is associated with the malignancy and poor prognosis of HCC. Decreased PGM1 expression obstructed glycogenesis pathway, which leads to the increased flow of glucose into glycolysis, thereby promoting tumor cell proliferation and HCC development. The loss of forkhead box protein J2 (FOXJ2), at least partly due to low genomic copy number in HCC, releases cellular nucleic acid-binding protein (CNBP), a nucleic acid chaperon, to bind to and promote G-quadruplex formation in PGM1 promoter and therefore decreases PGM1 expression. In addition, integrated analyses of PGM1 and FOXJ2 expression provide a better prediction for the malignance and prognosis of HCC. This study establishes a tumor-suppressive role of PGM1 by regulating glucose trafficking and uncovers a novel regulatory mechanism of PGM1 expression.
Assuntos
Carcinoma Hepatocelular/metabolismo , Glucose/metabolismo , Neoplasias Hepáticas/metabolismo , Fosfoglucomutase/metabolismo , Animais , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Proliferação de Células , Progressão da Doença , Regulação para Baixo , Feminino , Fatores de Transcrição Forkhead/genética , Fatores de Transcrição Forkhead/metabolismo , Regulação Enzimológica da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Glicólise , Células Hep G2 , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patologia , Masculino , Camundongos , Camundongos Nus , Pessoa de Meia-Idade , Fosfoglucomutase/deficiência , Fosfoglucomutase/genética , Prognóstico , Regiões Promotoras Genéticas , Proteínas de Ligação a RNA/antagonistas & inibidores , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismoRESUMO
BACKGROUND: A novel immunodeficiency, frequently accompanied by high serum-IgE, and caused by mutations in the PGM3 gene was described in 2014. To date there are no unique phenotype characteristics for PGM3 deficiency. PGM3 encodes a carbohydrate-modifying enzyme, phosphoglucomutase 3. Null-mutations are quite likely lethal, and to date only missense mutations or small deletions have been reported. Such mutations frequently cause a combination of reduced enzyme activity and protein instability, complicating determination of the enzyme level needed for survival. Here we present the first patient with a homozygous splice-modifying mutation in the PGM3 gene. An A > G substitution at position c.871 + 3 (transcript NM_001199917) is causing a deletion of exon 7 in the majority of PGM3 transcripts. In addition, this case further increases the clinical phenotypes of immunodeficiency caused by PGM3 mutations. CASE PRESENTATION: We describe the symptoms of a 3-year-old girl who was severely growth retarded, had vascular malformations, extensive eczema, multiple food-allergies, and was prone to infections. Unlike the majority of reported PGM3 deficient patients she lacked skeletal dysplasia and had normal neurocognitive development. In addition to the high serum-IgE, she displayed altered T cell numbers with reduced naïve CD4+ and CD8+ T-cells, increased number of activated effector memory CD8+ T cells and aberrant T-cell functions. The patient was homozygous for a new hypomorphic, splice-modifying mutation in the PGM3 gene, causing severely reduced mRNA levels. In the patient's cells, we observed 5% intact mRNA and approximately 11% of the protein levels seen in healthy controls. Treatment with allogeneic hematopoietic stem cell therapy was planned, but unfortunately the clinical condition deteriorated with multi-organ failure, which led to her death at 3 years of age. CONCLUSIONS: There is still no specific phenotype identified that distinguishes immunodeficiency caused by PGM3 mutations from other forms of immunodeficiency. The patient described here yields new information on the phenotypic variability among these patients. In addition, since all the synthesized protein is wild-type, it is possible for the first time to estimate the enzyme activity in vivo. The results suggest that1/10 of the normal PGM3 level is sufficient for survival but that it is insufficient for accurate carbohydrate processing.
Assuntos
Síndromes de Imunodeficiência/genética , Mutação , Fosfoglucomutase/genética , Sítios de Splice de RNA/genética , Pré-Escolar , Evolução Fatal , Feminino , Homozigoto , Humanos , Fosfoglucomutase/metabolismo , RNA Mensageiro/metabolismoRESUMO
Cancer aberrant N- and O-linked protein glycosylation, frequently resulting from an augmented flux through the Hexosamine Biosynthetic Pathway (HBP), play different roles in tumor progression. However, the low specificity and toxicity of the existing HBP inhibitors prevented their use for cancer treatment. Here we report the preclinical evaluation of FR054, a novel inhibitor of the HBP enzyme PGM3, with a remarkable anti-breast cancer effect. In fact, FR054 induces in different breast cancer cells a dramatic decrease in cell proliferation and survival. In particular, in a model of Triple Negative Breast Cancer (TNBC) cells, MDA-MB-231, we show that these effects are correlated to FR054-dependent reduction of both N- and O-glycosylation level that cause also a strong reduction of cancer cell adhesion and migration. Moreover we show that impaired survival of cancer cells upon FR054 treatment is associated with the activation of the Unfolded Protein Response (UPR) and accumulation of intracellular ROS. Finally, we show that FR054 suppresses cancer growth in MDA-MB-231 xenograft mice, supporting the advantage of targeting HBP for therapeutic purpose and encouraging further investigation about the use of this small molecule as a promising compound for breast cancer therapy.
Assuntos
Inibidores Enzimáticos/farmacologia , Inibidores Enzimáticos/uso terapêutico , Hexosaminas/biossíntese , Fosfoglucomutase/metabolismo , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Células MCF-7 , Camundongos , Fosfoglucomutase/antagonistas & inibidores , Transdução de Sinais/efeitos dos fármacos , Neoplasias de Mama Triplo Negativas/metabolismo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
To elucidate the starch synthesis pathway and the role of this reserve in rice pollen, we characterized mutations in the plastidic phosphoglucomutase, OspPGM, and the plastidic large subunit of ADP-glucose (ADP-Glc) pyrophosphorylase, OsAGPL4 Both genes were up-regulated in maturing pollen, a stage when starch begins to accumulate. Progeny analysis of self-pollinated heterozygous lines carrying the OspPGM mutant alleles, osppgm-1 and osppgm-2, or the OsAGPL4 mutant allele, osagpl4-1, as well as reciprocal crosses between the wild type (WT) and heterozygotes revealed that loss of OspPGM or OsAGPL4 caused male sterility, with the former condition rescued by the introduction of the WT OspPGM gene. While iodine staining and transmission electron microscopy analyses of pollen grains from homozygous osppgm-1 lines produced by anther culture confirmed the starch null phenotype, pollen from homozygous osagpl4 mutant lines, osagpl4-2 and osagpl4-3, generated by the CRISPR/Cas system, accumulated small amounts of starch which were sufficient to produce viable seed. Such osagpl4 mutant pollen, however, was unable to compete against WT pollen successfully, validating the important role of this reserve in fertilization. Our results demonstrate that starch is mainly polymerized from ADP-Glc synthesized from plastidic hexose phosphates in rice pollen and that starch is an essential requirement for successful fertilization in rice.
Assuntos
Glucose-1-Fosfato Adenililtransferase/metabolismo , Oryza/fisiologia , Fosfoglucomutase/metabolismo , Pólen/metabolismo , Amido/biossíntese , Fertilidade/fisiologia , Glucose-1-Fosfato Adenililtransferase/fisiologia , Microscopia , Mutação , Oryza/enzimologia , Oryza/metabolismo , Fosfoglucomutase/fisiologia , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
In recent years a number of primary immunodeficiencies (PIDs) characterized by elevated Immunoglobulin E (IgE) levels have been uncovered and termed as Hyper-IgE syndrome (HIES). In addition to the elevated levels of IgE, patients with these PIDs display a spectrum of infections by staphylococci and fungi, and in some cases viruses, particularly affecting skin and lungs. Most of these PIDs also have a non-infectious phenotype, comprising musculoskeletal, vascular, and neurological abnormalities. The genetic basis for the majority of conditions with elevated IgE has now been established and includes mutations in STAT3, DOCK8, TYK2, and most recently PGM3 molecules. However, in some patients with the relevant phenotype, mutations in these molecules are not identified, suggesting additional genetic etiologies of HIES not yet discovered. As the immunological and molecular basis of HIES is being unraveled, important insights are emerging that may have implications for our understanding of basic principles of immunology and protective immunity as well as for the pathogenesis and clinical management of patients with these complex and challenging PIDs. In this review, are presented the current knowledge on the clinical presentation, infectious phenotype, and the genetic and immunological pathogenesis of hyper-IgE syndromes as well as some other PIDs with elevated levels of IgE.