scholarly journals ETMM-05. LACTIC ACID FACILITATES GLIOBLASTOMA GROWTH THROUGH MODULATION OF THE EPIGENOME

2021 ◽  
Vol 3 (Supplement_1) ◽  
pp. i15-i15
Author(s):  
Consuelo Torrini ◽  
Trang Nguyen ◽  
Chang Shu ◽  
Angeliki Mela ◽  
Nelson Humala ◽  
...  
Keyword(s):  
Cell Death ◽  
Lactic Acid ◽  
Critical Role ◽  
Tca Cycle ◽  
Regulatory Elements ◽  
Acid Uptake ◽  
Gene Promoters ◽  
Acetyl Coa ◽  
The Tca Cycle

Abstract Glioblastoma (GBM) is the most common primary malignant brain tumor with an unfavorable prognosis. While GBMs utilize glucose, there are other carbon sources at their disposal. Lactate accumulates to a significant amount in the infiltrative margin of GBMs. In the current study, we demonstrated that lactate rescued patient-derived xenograft (PDX) GBM cells from nutrient deprivation mediated cell death and inhibition of growth. Transcriptome analysis, ATAC-seq and CHIP-seq. showed that lactic acid exposure entertained a signature of cell cycle progression and oxidative phosphorylation (OXPHOS) /tricarboxylic acid (TCA)-cycle. LC/MS analysis demonstrated that U-13C-Lactate elicited substantial labeling of TCA-cycle metabolites, acetyl-CoA and histone protein acetyl-residues in PDX derived GBM cells. Given that acetyl-CoA is pivotal for histone acetylation we observed a dose-dependent elevation of histone marks (e.g. H3K27ac), which was rescued by genetic and pharmacological inhibition of lactic acid-uptake, ATP-citrate lyase, p300 histone-acetyl-transferase and OXPHOS, resulting in reversal of lactate mediated protection from cell death. CHIP-seq. analysis demonstrated that lactic acid facilitated enhanced binding of H3K27ac to gene promoters and cis-regulatory elements. Consistently, ATAC-seq. analysis highlighted enhanced accessibility of the chromatin by lactic acid. In a combined tracer experiment (U-13C-glucose and 3-C13-lactate), we made the fundamental observation that lactic acid carbons were predominantly labeling the TCA cycle metabolites over glucose, implying a critical role of lactic acid in GBMs. Finally, pharmacological blockage of the TCA-cycle, using a clinically validated drug, extended overall survival in an orthotopic PDX model in mice without induction of toxicity, implying a critical role of lactic acid in GBMs and establishing lactic acid metabolism as a novel drug target for GBM.

scholarly journals EPCO-16. LACTIC ACID IS AN EPIGENETIC METABOLITE THAT DRIVES GLIOBLASTOMA SURVIVAL AND GROWTH

2020 ◽  
Vol 22 (Supplement_2) ◽  
pp. ii72-ii72
Author(s):  
Consuelo Torrini ◽  
Trang Nguyen ◽  
Chang Shu ◽  
Angeliki Mela ◽  
Nelson Humala ◽  
...  
Keyword(s):  
Cell Death ◽  
Lactic Acid ◽  
Critical Role ◽  
Waste Product ◽  
Tca Cycle ◽  
Regulatory Elements ◽  
Acid Uptake ◽  
Gene Promoters ◽  
Histone Acetyl Transferase ◽  
Acetyl Coa

Abstract Glioblastoma (GBM) is the most common primary malignant brain tumor with an unfavorable prognosis and a reprogrammed metabolism. While tumors utilize glucose, there are other carbon sources at their disposal. Originally considered as a waste product of glucose catabolism, lactate accumulates to a significant amount in tumor tissue. We launched our studies with the central hypothesis that lactate is metabolized by GBM cells to promote their survival via modulation of the epigenome. We showed that lactate rescued patient-derived xenograft (PDX) GBM cells from nutrient deprivation mediated cell death and inhibition of growth. Transcriptome analysis, Assay for Transposase-Accessible Chromatin using sequencing (ATAC-seq), and CHIP-seq. showed that lactic acid exposure entertained a signature of cell cycle progression, oxidative phosphorylation (OXPHOS) and MYC target expression. LC/MS analysis demonstrated that U-13C-Lactate elicited substantial labeling of TCA-cycle metabolites, acetyl-CoA and histone protein acetyl-residues in PDX derived GBM cells. Given that acetyl-CoA is pivotal for histone acetylation we observed a dose-dependent elevation of histone marks (e.g. H3K27ac), which was rescued by genetic and pharmacological inhibition of lactic acid-uptake, ATP-citrate lyase, p300 histone-acetyl-transferase and OXPHOS, resulting in reversal of lactate mediated protection from cell death or facilitation of GBM growth. CHIP-seq. analysis demonstrated that lactic acid facilitated enhanced binding of H3K27ac to gene promoters and cis-regulatory elements (e.g. super-enhancers). Consistently, ATAC-seq. analysis highlighted enhanced accessibility of the chromatin by lactic acid. Finally, we assessed whether lactic acid is actively metabolized in vivo, utilizing an orthotopic PDX model of GBM. In a combined tracer experiment (U-13C-glucose and 3-C13-lactate), we made the fundamental observation that lactic acid carbons were predominantly labeling the TCA cycle metabolites over glucose, implying a critical role of lactic acid in GBMs and establishing lactic acid metabolism as a novel drug target for GBM that may be targeted with epigenetic drugs.

Liquid-liquid extraction of succinate using a dicopper cryptate

2020 ◽  
Author(s):  
Riccardo Mobili ◽  
Sonia La Cognata ◽  
Francesca Merlo ◽  
Andrea Speltini ◽  
Massimo Boiocchi ◽  
...  
Keyword(s):  
Aqueous Phase ◽  
Visible Spectrum ◽  
Tca Cycle ◽  
Residual Concentration ◽  
Waste Products ◽  
Liquid Liquid Extraction ◽  
The Tca Cycle ◽  
Quantitative Extraction ◽  
Uv Visible

<div> <p>The extraction of the succinate dianion from a neutral aqueous solution into dichloromethane is obtained using a lipophilic cage-like dicopper(II) complex as the extractant. The quantitative extraction exploits the high affinity of the succinate anion for the cavity of the azacryptate. The anion is effectively transferred from the aqueous phase, buffered at pH 7 with HEPES, into dichloromethane. A 1:1 extractant:anion adduct is obtained. Extraction can be easily monitored by following changes in the UV-visible spectrum of the dicopper complex in dichloromethane, and by measuring the residual concentration of succinate in the aqueous phase by HPLC−UV. Considering i) the relevance of polycarboxylates in biochemistry, as e.g. normal intermediates of the TCA cycle, ii) the relevance of dicarboxylates in the environmental field, as e.g. waste products of industrial processes, and iii) the recently discovered role of succinate and other dicarboxylates in pathophysiological processes including cancer, our results open new perspectives for research in all contexts where selective recognition, trapping and extraction of polycarboxylates is required. </p> </div>

scholarly journals A Dual Role of Heme Oxygenase-1 in Cancer Cells

2018 ◽  
Vol 20 (1) ◽  
pp. 39 ◽  
Author(s):  
Shih-Kai Chiang ◽  
Shuen-Ei Chen ◽  
Ling-Chu Chang
Keyword(s):  
Cell Death ◽  
Heme Oxygenase ◽  
Cancer Progression ◽  
Critical Role ◽  
Causative Factor ◽  
Protective Role ◽  
Dark Side ◽  
Heme Oxygenase 1 ◽  
Cellular Iron

Heme oxygenase (HO)-1 is known to metabolize heme into biliverdin/bilirubin, carbon monoxide, and ferrous iron, and it has been suggested to demonstrate cytoprotective effects against various stress-related conditions. HO-1 is commonly regarded as a survival molecule, exerting an important role in cancer progression and its inhibition is considered beneficial in a number of cancers. However, increasing studies have shown a dark side of HO-1, in which HO-1 acts as a critical mediator in ferroptosis induction and plays a causative factor for the progression of several diseases. Ferroptosis is a newly identified iron- and lipid peroxidation-dependent cell death. The critical role of HO-1 in heme metabolism makes it an important candidate to mediate protective or detrimental effects via ferroptosis induction. This review summarizes the current understanding on the regulatory mechanisms of HO-1 in ferroptosis. The amount of cellular iron and reactive oxygen species (ROS) is the determinative momentum for the role of HO-1, in which excessive cellular iron and ROS tend to enforce HO-1 from a protective role to a perpetrator. Despite the dark side that is related to cell death, there is a prospective application of HO-1 to mediate ferroptosis for cancer therapy as a chemotherapeutic strategy against tumors.

scholarly journals A critical role of superoxide anion in selenite-induced mitophagic cell death

Autophagy ◽  
2008 ◽  
Vol 4 (1) ◽  
pp. 76-78 ◽  
Author(s):  
Eun Hee Kim ◽  
Kyeong Sook Choi
Keyword(s):  
Cell Death ◽  
Superoxide Anion ◽  
Critical Role

Gluconeogenesis from labeled carbon: estimating isotope dilution

1986 ◽  
Vol 250 (3) ◽  
pp. E296-E305 ◽  
Author(s):  
J. K. Kelleher
Keyword(s):  
Steady State ◽  
Isotope Dilution ◽  
Experimental Tests ◽  
Tca Cycle ◽  
Tricarboxylic Acid ◽  
Complex Model ◽  
Acetyl Coa ◽  
The Tca Cycle ◽  
Carbon Compounds ◽  
Mathematical Techniques

To estimate the rate of gluconeogenesis from steady-state incorporation of labeled 3-carbon precursors into glucose, isotope dilution must be considered so that the rate of labeling of glucose can be quantitatively converted to the rate of gluconeogenesis. An expression for the value of this isotope dilution can be derived using mathematical techniques and a model of the tricarboxylic acid (TCA) cycle. The present investigation employs a more complex model than that used in previous studies. This model includes the following pathways that may affect the correction for isotope dilution: 1) flux of 3-carbon precursor to the oxaloacetate pool via acetyl-CoA and the TCA cycle; 2) flux of 4- or 5-carbon compounds into the TCA cycle; 3) reversible flux between oxaloacetate (OAA) and pyruvate and between OAA and fumarate; 4) incomplete equilibrium between OAA pools; and 5) isotope dilution of 3-carbon tracers between the experimentally measured pool and the precursor for the TCA-cycle OAA pool. Experimental tests are outlined which investigators can use to determine whether these pathways are significant in a specific steady-state system. The study indicated that flux through these five pathways can significantly affect the correction for isotope dilution. To correct for the effects of these pathways an alternative method for calculating isotope dilution is proposed using citrate to relate the specific activities of acetyl-CoA and OAA.

Ammonia metabolism

1978 ◽  
Vol 235 (4) ◽  
pp. F265-F277 ◽  
Author(s):  
R. L. Tannen
Keyword(s):  
Critical Role ◽  
Tca Cycle ◽  
Gamma Glutamyl Transferase ◽  
Urine Ph ◽  
Ammonia Metabolism ◽  
Adaptive Process ◽  
Purine Nucleotide Cycle ◽  
Tca Cycle Intermediates ◽  
Glutamyl Transferase

The pathways responsible for an the mechanisms underlying the adaptive increase in ammonia production in response to acidosis are considered. It seems unlikely that the cytosolic pathways (glutamine synthetase, glutaminase II, phosphate-independent glutaminase, and gamma-glutamyl transferase) are of primary importance in the adaptive process, but the role of the purine nucleotide cycle has not been resolved. The intramitochondrially located phosphate-dependent glutaminase pathway is generally believed to be of primary importance. Adaptation involved either enhanced glutamine entry into the mitrochondria and/or activation of phosphate-dependent glutaminase, but the relative importance of each has not been resolved definitively. The overall adaptive response is probably modulated by factors regulating alpha-ketoglutarate metabolism to phosphoenolpyruvate, and possibly also by metabolism of TCA cycle intermediates. It seems unlikely that a decrease in systemic pH is the direct effector for the acidosis-induced increase in ammonia formation; however, the resulting decrease in urine pH may play a critical role. Other potential messengers, including potassium, glucocorticoids, mineralocorticoids, cyclic AMP, and calcium probably do not serve a primary function, but the importance of other circulating factor(s) is unclear.

scholarly journals Comprehensive Genomic Discovery of Non-Coding Transcriptional Enhancers in the African Malaria Vector Anopheles coluzzii

2022 ◽  
Vol 12 ◽  
Author(s):  
Inge Holm ◽  
Luisa Nardini ◽  
Adrien Pain ◽  
Emmanuel Bischoff ◽  
Cameron E. Anderson ◽  
...  
Keyword(s):  
Malaria Vector ◽  
Regulatory Elements ◽  
Specific Cell ◽  
Insect Vector ◽  
Anopheles Coluzzii ◽  
Gene Promoters ◽  
Transcriptional Enhancers ◽  
Genome Wide ◽  
A Genome

Almost all regulation of gene expression in eukaryotic genomes is mediated by the action of distant non-coding transcriptional enhancers upon proximal gene promoters. Enhancer locations cannot be accurately predicted bioinformatically because of the absence of a defined sequence code, and thus functional assays are required for their direct detection. Here we used a massively parallel reporter assay, Self-Transcribing Active Regulatory Region sequencing (STARR-seq), to generate the first comprehensive genome-wide map of enhancers in Anopheles coluzzii, a major African malaria vector in the Gambiae species complex. The screen was carried out by transfecting reporter libraries created from the genomic DNA of 60 wild A. coluzzii from Burkina Faso into A. coluzzii 4a3A cells, in order to functionally query enhancer activity of the natural population within the homologous cellular context. We report a catalog of 3,288 active genomic enhancers that were significant across three biological replicates, 74% of them located in intergenic and intronic regions. The STARR-seq enhancer screen is chromatin-free and thus detects inherent activity of a comprehensive catalog of enhancers that may be restricted in vivo to specific cell types or developmental stages. Testing of a validation panel of enhancer candidates using manual luciferase assays confirmed enhancer function in 26 of 28 (93%) of the candidates over a wide dynamic range of activity from two to at least 16-fold activity above baseline. The enhancers occupy only 0.7% of the genome, and display distinct composition features. The enhancer compartment is significantly enriched for 15 transcription factor binding site signatures, and displays divergence for specific dinucleotide repeats, as compared to matched non-enhancer genomic controls. The genome-wide catalog of A. coluzzii enhancers is publicly available in a simple searchable graphic format. This enhancer catalogue will be valuable in linking genetic and phenotypic variation, in identifying regulatory elements that could be employed in vector manipulation, and in better targeting of chromosome editing to minimize extraneous regulation influences on the introduced sequences.Importance: Understanding the role of the non-coding regulatory genome in complex disease phenotypes is essential, but even in well-characterized model organisms, identification of regulatory regions within the vast non-coding genome remains a challenge. We used a large-scale assay to generate a genome wide map of transcriptional enhancers. Such a catalogue for the important malaria vector, Anopheles coluzzii, will be an important research tool as the role of non-coding regulatory variation in differential susceptibility to malaria infection is explored and as a public resource for research on this important insect vector of disease.

scholarly journals Liquid-liquid extraction of succinate using a dicopper cryptate

2020 ◽  
Author(s):  
Riccardo Mobili ◽  
Sonia La Cognata ◽  
Francesca Merlo ◽  
Andrea Speltini ◽  
Massimo Boiocchi ◽  
...  
Keyword(s):  
Aqueous Phase ◽  
Visible Spectrum ◽  
Tca Cycle ◽  
Residual Concentration ◽  
Waste Products ◽  
Liquid Liquid Extraction ◽  
The Tca Cycle ◽  
Quantitative Extraction ◽  
Uv Visible

<div> <p>The extraction of the succinate dianion from a neutral aqueous solution into dichloromethane is obtained using a lipophilic cage-like dicopper(II) complex as the extractant. The quantitative extraction exploits the high affinity of the succinate anion for the cavity of the azacryptate. The anion is effectively transferred from the aqueous phase, buffered at pH 7 with HEPES, into dichloromethane. A 1:1 extractant:anion adduct is obtained. Extraction can be easily monitored by following changes in the UV-visible spectrum of the dicopper complex in dichloromethane, and by measuring the residual concentration of succinate in the aqueous phase by HPLC−UV. Considering i) the relevance of polycarboxylates in biochemistry, as e.g. normal intermediates of the TCA cycle, ii) the relevance of dicarboxylates in the environmental field, as e.g. waste products of industrial processes, and iii) the recently discovered role of succinate and other dicarboxylates in pathophysiological processes including cancer, our results open new perspectives for research in all contexts where selective recognition, trapping and extraction of polycarboxylates is required. </p> </div>

scholarly journals Genome-Wide Analysis of PR-1 Genes From Piper Nigrum Reveals Critical Role in Defense Response During Phytophthora Capsici Infection

2020 ◽  
Author(s):  
Divya Kattupalli ◽  
Asha Sriniva ◽  
Soniya E V
Keyword(s):  
Function Analysis ◽  
Phytophthora Capsici ◽  
Critical Role ◽  
Defense Responses ◽  
Regulatory Elements ◽  
Black Pepper ◽  
Piper Nigrum ◽  
Binding Motif ◽  
Genome Wide

Abstract Background: Black pepper is a prominent spice which is an indispensable ingredient in culinary and traditional medicine. Phytophthora capsici, the causative agent of foot rot disease causes drastic constraint in black pepper cultivation and productivity. To counterattack various biotic and abiotic stresses plants employ a broad array of mechanisms one such includes the accumulation of pathogenesis-related (PR) proteins. Several studies have reported the role of PR-1 proteins in triggering the plant defenses during plant-oomycete interaction.Results: Through the genome-wide survey, eleven PR-1 genes that belongs to a CAP superfamily protein with Caveolin-Binding Motif (CBM) and CAP-derived peptide (CAPE) were identified from P. nigrum. Despite the critical functional domains, PnPR1 homologs differ in their signal peptide motifs, and core amino acid sequence composition in the functional protein domains. The GO, biological function analysis reveals their role in defense responses and response to biotic stimulus whereas the KEGG functional annotation predicted their function in the plant-pathogen interactions. Furthermore, transcriptome-assisted FPKM analysis revealed PnPR-1 genes mapped to P. nigrum - P. capsici interaction pathway. The differentially expressed pathogen-responsive PR-1 gene was validated through qRT-PCR. Subsequent analysis revealed the structural details, phylogenetic relationships, conserved sequence motifs and critical cis-regulatory elements of PnPR-1 genes.Conclusion: This is the first genome-wide study that identified the role of PR-1 genes during P. nigrum - P. capsici interactions. The detailed in silico experimental analysis revealed the vital role of PnPR-1 genes in regulating the first layer of defense towards P. capsici infection in Panniyur-1 plants.

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