Glutamine Modulates Expression and Function of Glucose 6-Phosphate Dehydrogenase via NRF2 in Colon Cancer Cells.
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ABSTRACT:
SUBMITTER: Polat IH
PROVIDER: S-EPMC8472416 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Rationale: NRF2, a redox sensitive transcription factor, is up-regulated in head and neck squamous cell carcinoma (HNSCC), however, the associated impact and regulatory mechanisms remain unclear. Methods: The protein expression of NRF2 in HNSCC specimens was examined by IHC. The regulatory effect of c-MYC on NRF2 was validated by ChIP-qPCR, RT-qPCR and western blot. The impacts of NRF2 on malignant progression of HNSCC were determined through genetic manipulation and pharmacological inhibition in vitro and in vivo. The gene-set enrichment analysis (GSEA) on expression data of cDNA microarray combined with ChIP-qPCR, RT-qPCR, western blot, transwell migration/ invasion, cell proliferation and soft agar colony formation assays were used to investigate the regulatory mechanisms of NRF2. Results: NRF2 expression is positively correlated with malignant features of HNSCC. In addition, carcinogens, such as nicotine and arecoline, trigger c-MYC-directed NRF2 activation in HNSCC cells. NRF2 reprograms a wide range of cancer metabolic pathways and the most notable is the pentose phosphate pathway (PPP). Furthermore, glucose-6-phosphate dehydrogenase (G6PD) and transketolase (TKT) are critical downstream effectors of NRF2 that drive malignant progression of HNSCC; the coherently expressed signature NRF2/G6PD/TKT gene set is a potential prognostic biomarker for prediction of patient overall survival. Notably, G6PD- and TKT-regulated nucleotide biosynthesis is more important than redox regulation in determining malignant progression of HNSCC. Conclusions: Carcinogens trigger c-MYC-directed NRF2 activation. Over-activation of NRF2 promotes malignant progression of HNSCC through reprogramming G6PD- and TKT-mediated nucleotide biosynthesis. Targeting NRF2-directed cellular metabolism is an effective strategy for development of novel treatments for head and neck cancer.
| S-EPMC8039948 | biostudies-literature
Project description:Why do the activities of some enzymes greatly exceed the flux capacity of the embedding pathways? This is a puzzling open problem in quantitative evolutionary design. In this work we investigate reasons for high expression of a thoroughly characterized enzyme: glucose 6-phosphate dehydrogenase (G6PD) in human erythrocytes. G6PD catalyses the first step of the pathway that supplies NADPH for antioxidant defense mechanisms. Normal G6PD activity far exceeds the capacity of human erythrocytes for a steady NADPH supply, which is limited upstream of G6PD. However, the distribution of erythrocyte G6PD activity in human populations reveals a selective pressure for maintaining high activity. To clarify the nature of this selective pressure, we studied how G6PD activity and other parameters in a model of the NADPH redox cycle affect metabolic performance. Our analysis indicates that normal G6PD activity is sufficient but not superfluous to avoid NADPH depletion and ensure timely adaptation of the NADPH supply during pulses of oxidative load such as those that occur during adherence of erythrocytes to phagocytes. These results suggest that large excess capacities found in some biochemical and physiological systems, rather than representing large safety factors, may reflect a close match of system design to unscrutinized performance requirements. Understanding quantitative evolutionary design thus calls for careful consideration of the various performance specifications that biological components/processes must meet in order for the organism to be fit. The biochemical systems framework used in this paper is generally applicable for such a detailed examination of the quantitative evolutionary design of gene expression levels in other systems.
| S-EPMC283614 | biostudies-literature
Project description:G6PDH is the rate-limiting enzyme of the pentose phosphate pathway and one of the principal source of NADPH, a major cellular reductant. Importantly, in ruminant's metabolism the aforementioned NADPH provided, is utilized for de novo fatty acid synthesis. Previous work of cloning the ovine (Ovis aries) og6pdh gene has revealed the presence of two cDNA transcripts (og6pda and og6pdb), og6pdb being a product of alternative splicing not similar to any other previously reported.(1) In the current study the effect of energy balance in the ovine G6PDH protein expression was investigated, shedding light on the biochemical features and potential physiological role of the oG6PDB isoform. Changes in energy balance leads to protein expression changes in both transcripts, to the opposite direction and not in a proportional way. Negative energy balance was not in favor of the presence of any particular isoform, while both protein expression levels were not significantly different (P > 0.05). In contrast, at the transition point from negative to positive and on the positive energy balance, there is a significant increase of oG6PDA compared with oG6PDB protein expression (P < 0.001). Both oG6PDH protein isoforms changed significantly toward the positive energy balance. oG6PDA is escalating, while oG6PDB is falling, under the same stimulus (positive energy balance alteration). This change is also positively associated with increasing levels in enzyme activity, 4 weeks post-weaning in ewes' adipose tissue. Furthermore, regression analysis clearly demonstrated the linear correlation of both proteins in response to the WPW, while energy balance, enzyme activity, and oG6PDA relative protein expression follow the same escalating trend; in contrast, oG6PDB relative protein expression falls in time, similar to both transcripts accumulation pattern, as reported previously.(2.)
| S-EPMC3917929 | biostudies-literature
Project description:Glutamine (Gln) and glucose (Glc) represent two important nutrients for proliferating cells, consistent with the observations that oncogenic processes are associated with enhanced glycolysis and glutaminolysis. Gln depletion and Glc depletion have been shown to trigger growth arrest and eventually cell death. Solid tumors often outgrow the blood supply, resulting in ischemia, which is associated with hypoxia and nutrient insufficiency. Whereas oxygen-sensing and adaptive mechanisms to hypoxia have been well-studied, how cells directly sense and respond to Gln and Glc insufficiency remains unclear. Using mRNA profiling techniques, we compared the gene expression profiles of acute Gln-depleted cells, Glc-depleted cells and cells adapted to Gln depletion. Here we report the global changes of the gene expression in those cells cultured under the defined nutrient conditions. Analysis of mRNA profiling data revealed that Gln and Glc depletion triggered dramatic gene expression reprogramming. Either Gln or Glc deletion leads to changes of the expression of cell cycle genes, but these conditions have distinctive effects on transcription regulators and gene expression profiles. Moreover, Gln and Glc depletion triggered distinguishable ER-stress responses. The gene expression patterns support that Gln and Glc have distinctive metabolic roles in supporting cell survival and proliferation, and cells use different mechanisms to sense and respond to Gln and Glc insufficiency. Our mRNA profiling database provides a resource for further investigating the nutrient-sensing mechanisms and potential effects of Glc and Gln abundance on the biological behaviors of cells.
| S-EPMC3478318 | biostudies-literature
Project description:Among the determinants contributing to the pathogenesis of asthma, antioxidant genetic factors play a leading role. Glucose-6-phosphate dehydrogenase (G6PD) is an enzyme that is competent to detoxify free radicals. Although a relationship between G6PD deficiency and asthma has been previously reported, the literature is still scanty. In this study, we test this hypothesis in a large cohort of patients from Sardinia, Italy. A retrospective case-control study was performed using data from 11,829 clinical records of outpatients referred to a teaching hospital for a medical visit. In total, 455 cases (asthma-positive) and 11,374 controls (asthma-negative) were compared for G6PD status using multivariable analysis, adjusting for all covariates. Overall, G6PD deficiency was detected in 11.2% of study participants and was associated with an increased risk of asthma (odds ratio (OR) 1.63; 95% confidence interval (CI) 1.27-2.10). Additional variables significantly associated with asthma were female sex (OR 1.66; 95% CI 1.34-2.06), overweight/obesity (OR 1.56; 95% CI 1.27-1.92), smoking (OR 1.44; 95% CI 1.449-3.963), and high socioeconomic status (OR 1.40; 95% CI 1.16-1.70), whereas age was inversely related with asthma (OR 0.49; 95% CI 0.39-0.61). Our study shows that G6PD deficiency is an independent risk for asthma. These findings suggest that G6PD should be assessed in asthmatic patients for better risk stratification.
| S-EPMC8658649 | biostudies-literature
Project description:Five bromophenols isolated from three Rhodomelaceae algae (Laurencia nipponica, Polysiphonia morrowii, Odonthalia corymbifera) showed inhibitory effects against glucose 6-phosphate dehydrogenase (G6PD). Among them, the symmetric bromophenol dimer (5) showed the highest inhibitory activity against G6PD.
| S-EPMC3826149 | biostudies-literature
Project description:BackgroundGlucose-regulated proteins (GRP) are induced in the cancer microenvironment to promote tumor survival, metastasis and drug resistance. AST was obtained from the medicinal plant Astragalus membranaceus, which possesses anti-tumor and pro-apoptotic properties in colon cancer cells and tumor xenograft. The present study aimed to investigate the involvement of GRP in endoplasmic reticulum (ER) stress-mediated apoptosis during colon cancer development, with focus on the correlation between AST-evoked regulation of GRP and calpain activation.MethodsThe effects of AST on GRP and apoptotic activity were assessed in HCT 116 human colon adenocarcinoma cells. Calpain activity was examined by using a fluorescence assay kit. Immunofluorescence staining and immunoprecipitation were employed to determine the localization and association between calpains and GRP. GRP78 gene silencing was performed to confirm the importance of GRP in anticancer drug activities. The modulation of GRP and calpains was also studied in nude mice xenograft.ResultsER stress-mediated apoptosis was induced by AST, as shown by elevation in both spliced XBP-1 and CHOP levels, with parallel up-regulation of GRP. The expression of XBP-1 and CHOP continued to increase after the peak level of GRP was attained at 24 h. Nevertheless, the initial increase in calpain activity as well as calpain I and II protein level was gradually declined at later stage of drug treatment. Besides, the induction of GRP was partly reversed by calpain inhibitors, with concurrent promotion of AST-mediated apoptosis. The knockdown of GRP78 by gene silencing resulted in higher sensitivity of colon cancer cells to AST-induced apoptosis and reduction of colony formation. The association between calpains and GRP78 had been confirmed by immunofluorescence staining and immunoprecipitation. Modulation of GRP and calpains by AST was similarly demonstrated in nude mice xenograft, leading to significant inhibition of tumor growth.ConclusionsOur findings exemplify that calpains, in particular calpain II, play a permissive role in the modulation of GRP78 and consequent regulation of ER stress-induced apoptosis. Combination of calpain inhibitors and AST could exhibit a more pronounced pro-apoptotic effect. These results help to envisage a new therapeutic approach in colon cancer by targeting calpain and GRP.
| S-EPMC4210535 | biostudies-literature
Project description:Glucose 6-phosphate dehydrogenase (G6PD) (EC 1.1.1.363) is a crucial regulatory enzyme in the oxidative pentose phosphate pathway that provides reductive potential in the form of NADPH, as well as carbon skeletons for the synthesis of macromolecules. In this study, we report the cloning, expression, and characterization of G6PD (SpG6PD1) from a lichen-associated psychrophilic bacterium Sphingomonas sp. PAMC 26621. SpG6PD1 was expressed in Escherichia coli as a soluble protein, having optimum activity at pH 7.5?8.5 and 30 °C for NADP? and 20 °C for NAD?. SpG6PD1 utilized both NADP? and NAD?, with the preferential utilization of NADP?. A high Km value for glucose 6-phosphate and low activation enthalpy (?H‡) compared with the values of mesophilic counterparts indicate the psychrophilic nature of SpG6PD1. Despite the secondary structure of SpG6PD1 being maintained between 4?40 °C, its activity and tertiary structure were better preserved between 4?20 °C. The results of this study indicate that the SpG6PD1 that has a flexible structure is most suited to a psychrophilic bacterium that is adapted to a permanently cold habitat.
| S-EPMC6471386 | biostudies-literature
Project description:Total glucose-6-phosphate dehydrogenase (G6PDH) activity, protein abundance, and transcript levels of G6PDH isoforms were measured in response to exogenous abscisic acid (ABA) supply to barley (Hordeum vulgare cv Nure) hydroponic culture. Total G6PDH activity increased by 50% in roots treated for 12?h with exogenous 0.1?mM ABA. In roots, a considerable increase (35%) in plastidial P2-G6PDH transcript levels was observed during the first 3?h of ABA treatment. Similar protein variations were observed in immunoblotting analyses. In leaves, a 2-fold increase in total G6PDH activity was observed after ABA treatment, probably related to an increase in the mRNA level (increased by 50%) and amount of protein (increased by 85%) of P2-G6PDH. Together these results suggest that the plastidial P2-isoform plays an important role in ABA-treated barley plants.
| S-EPMC3134356 | biostudies-literature
Project description:The relationship between glucose-6-phosphate dehydrogenase (G6PD) deficiency and clinical phenomena such as primaquine-sensitivity and protection from severe malaria remains poorly defined, with past association studies yielding inconsistent and conflicting results. One possibility is that examination of a single genetic variant might underestimate the presence of true effects in the presence of unrecognized functional allelic diversity.We systematically examined this possibility in Kenya, conducting a fine-mapping association study of erythrocyte G6PD activity in 1828 Kenyan children across 30 polymorphisms at or around the G6PD locus.We demonstrate a strong functional role for c.202G>A (rs1050828), which accounts for the majority of variance in enzyme activity observed (P=1.5×10?²??, additive model). Additionally, we identify other common variants that exert smaller, intercorrelated effects independent of c.202G>A, and haplotype analyses suggest that each variant tags one of two haplotype motifs that are opposite in sequence identity and effect direction. We posit that these effects are of biological and possible clinical significance, specifically noting that c.376A>G (rs1050829) augments 202AG heterozygote risk for deficiency trait by two-fold (OR = 2.11 [1.12 - 3.84], P=0.014).Our results suggest that c.202G>A is responsible for the majority of the observed prevalence of G6PD deficiency trait in Kenya, but also identify a novel role for c.376A>G as a genetic modifier which marks a common haplotype that augments the risk conferred to 202AG heterozygotes, suggesting that variation at both loci merits consideration in genetic association studies probing G6PD deficiency-associated clinical phenotypes.
| S-EPMC4236593 | biostudies-literature