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GFER Represents a Target for Dual Disruption of Redox Homeostasis and Reactivation of the Immune Response in Pancreatic Adenocarcinoma.

UNLABELLED: Both metabolic dysregulation and the immunosuppressive tumor microenvironment of pancreatic ductal adenocarcinoma (PDAC) contribute to the recalcitrance of this lethal disease to treatment. Accordingly, we aimed to identify and characterize a target that elicits an anticancer response through both disrupting cancer cell redox homeostasis and increasing the immunogenicity of PDAC. First, mitochondrial metabolic dependencies in PDAC were identified by using a CRISPR-Cas9 screening system with a custom single-guide RNA library. Functional validation analyses revealed GFER, a mitochondrial FAD-dependent sulfhydryl oxidase, as an essential regulator of tumor growth. In vitro and in vivo methodologies demonstrated that GFER depletion perturbed redox homeostasis and stimulated tumor immunogenicity, including sensitization to immune checkpoint blockade. In patient-derived xenograft models of PDAC, the growth-inhibitory response induced by GFER depletion was mediated by an altered oxidative balance that released damaged mitochondrial DNA into the cytoplasm of tumor cells, leading to the activation of the cGAS-STING pathway and expression of type I IFNs. This effect was recapitulated in a mouse immunocompetent syngeneic PDAC model, in which GFER depletion suppressed tumor growth and promoted T-cell infiltration to enhance tumor-killing effects. Consequently, GFER depletion significantly increased the antitumor efficacy of immune checkpoint blockade. Overall, these findings identify GFER as a critical node for both mitochondrial redox homeostasis and immunomodulation in PDAC and reveal a therapeutic opportunity for sensitizing PDAC to immune checkpoint blockade. SIGNIFICANCE: GFER is essential for mitochondrial redox balance and suppressing tumor immunogenicity in pancreatic tumors, with the combination of GFER inhibition with immune checkpoint blockade resulting in a strong antitumor response.

Animals↗

Degradation of myxovirus virion RNA by periodate.

Extensively degraded RNA was isolated from virions of influenza virus which had been oxidized with sodium m-periodate. Similarly, although to a lesser extent, RNA isolated from periodate-treated ribonucleoprotein of influenza virus was also degraded. In contrast, influenza virus RNA, if first freed from other virion components, was not degraded by periodate oxidation.

Hydrogen-Ion Concentration↗

Perm1 enhances Nrf2-driven antioxidant defense through Keap1 oxidation during myocardial ischemia/reperfusion injury.

Ischemia/reperfusion (IR) enhances oxidative stress, leading to myocardial injury. Although Perm1 promotes cytoprotective mechanisms, the underlying mechanisms are poorly understood. Cysteine oxidation of Keap1 alleviates Cul3-mediated ubiquitination/degradation of Nrf2 and promotes antioxidant transcription. Here we show that Perm1 activates Nrf2 through cysteine oxidation of Keap1 and stabilization of Nrf2. Endogenous Perm1 was downregulated during IR, whereas the rescue of Perm1 reduced IR injury. Downregulation of Perm1 exacerbated oxidative stress, whereas upregulation of Perm1 alleviated it, accompanied by downregulation and upregulation of Nrf2-regulated antioxidant genes, respectively. Perm1 promoted oxidation of cysteine residues in Keap1, possibly through thiol-disulfide exchange reactions, which decreases Keap1-Nrf2 interaction and inhibits Cul3-mediated degradation of Nrf2. We identified Cys121 and Cys746 in Perm1 as critical for Keap1 oxidation and cardioprotection. Thus, Perm1 induces cysteine oxidation of Keap1, thereby conferring myocardial resistance to IR injury by inducing Nrf2 stabilization and transcriptional activation of antioxidant genes.

Kelch-Like ECH-Associated Protein 1↗

Dependence on dose of the acute effects of ethanol on liver metabolism in vivo.

The dose dependence of the acute effects of ethanol upon liver intermediary metabolism in vivo has been demonstrated in rats. Ethanol was given i.p. in doses of 0.69, 1.7, and 3.0 g/kg in equal volumes (20 ml/kg). The liver was freeze-clamped 120 min after injection, and multiple metabolites were measured in the perchloric acid extract of the tissue. Each group showed a significantly different pattern of metabolites, redox states, and phosphorylation potentials although the rate of ethanol disappearance, at least between the two highest dose groups, was not significantly different. The mitochondrial free [NAD+]/[NADH] ratios and the cytoplasmic free [NADP+]/[NADPH] ratio were paradoxically most reduced with the lowest dose of ethanol and became progressively more oxidized with increasing dose. Once established, the differences in these ratios between the groups tended to persist with time, relatively independent of the concentration of ethanol. In a somewhat different pattern, the phosphorylation potential ([ATP]/[ADP][P1]) remained at the control level in the low-dose group but was significantly elevated in the two higher-dose groups. The results, therefore, show distinct and complicated dose-dependent patterns of intermediary metabolism that cannot be explained completely by any one hypothesis but that imply significant dose-dependent effects of ethanol upon intermediary metabolism not directly related to NADH production.

Adenosine Triphosphate↗

A probabilistic generative model for quantification of DNA modifications enables analysis of demethylation pathways.

We present a generative model, Lux, to quantify DNA methylation modifications from any combination of bisulfite sequencing approaches, including reduced, oxidative, TET-assisted, chemical-modification assisted, and methylase-assisted bisulfite sequencing data. Lux models all cytosine modifications (C, 5mC, 5hmC, 5fC, and 5caC) simultaneously together with experimental parameters, including bisulfite conversion and oxidation efficiencies, as well as various chemical labeling and protection steps. We show that Lux improves the quantification and comparison of cytosine modification levels and that Lux can process any oxidized methylcytosine sequencing data sets to quantify all cytosine modifications. Analysis of targeted data from Tet2-knockdown embryonic stem cells and T cells during development demonstrates DNA modification quantification at unprecedented detail, quantifies active demethylation pathways and reveals 5hmC localization in putative regulatory regions.

5-Methylcytosine↗

Two worlds beneath: Distinct microbial strategies of the rock-attached and planktonic subsurface biosphere.

BACKGROUND: Microorganisms in groundwater ecosystems exist either as planktonic cells or as attached communities on aquifer rock surfaces. Attached cells outnumber planktonic ones by at least three orders of magnitude, suggesting a critical role in aquifer ecosystem function. However, particularly in consolidated carbonate aquifers, where research has predominantly focused on planktonic microbes, the metabolic potential and ecological roles of attached communities remain poorly understood. RESULTS: To investigate the differences between attached and planktonic communities, we sampled the attached microbiome from passive samplers filled with crushed carbonate rock exposed to oxic and anoxic groundwater in the Hainich Critical Zone Exploratory and compared it to a previously published, extensive dataset of planktonic communities from the same aquifer ecosystem. Microbial lifestyle (attached vs. planktonic) explained more variance in community composition than redox conditions, prompting us to further investigate its role in shaping functional and activity profiles. Metagenomic analysis revealed a striking taxonomic and functional segregation: the 605 metagenome-assembled genomes (MAGs) from attached communities were dominated by Proteobacteria (358 MAGs) and were enriched in genes for biofilm formation, chemolithoautotrophy, and redox cycling (e.g., iron and sulfur metabolism). In contrast, the 891 MAGs from planktonic communities were dominated by Cand. Patescibacteria (464 MAGs) and Nitrospirota (60 MAGs) and showed lower functional versatility. Only a few genera were shared, and even closely related MAGs (> 90% average nucleotide identity) differed in assembly size and metabolic traits, demonstrating lifestyle-specific functional adaptation. Analysis of active replication indicated that the active fraction of the attached community was primarily represented by the most abundant MAGs. Planktonic communities featured a higher fraction of active MAGs compared to attached communities, but overall with lower relative abundances. CONCLUSIONS: The high abundance, metabolic specialization, and carbon fixation potential of attached microbes suggest that they are key drivers of subsurface biogeochemical processes. Carbonate aquifers may act as much larger inorganic carbon sinks than previously estimated based on CO2 fixation rates of the planktonic communities alone. Our findings underscore the need to incorporate attached microbial communities into models of subsurface ecosystem function. Video Abstract.

Plankton↗

Potential survival strategies of novel comammox and nitrite-oxidizing Nitrospira synthesizing osmoprotectants in a wastewater microbiome treating high-ammonia brackish landfill leachate.

BACKGROUND: In the late stages of landfill operation, leachate becomes brackish and contains high concentrations of ammonia with limited organic carbon. At leachate treatment facilities, it is typically subjected to nitrification followed by denitrification, with methanol supplied as an external electron donor. This unique environment may harbor novel microorganisms, including nitrifiers. Although a variety of microorganisms are involved in nitrification, their substrate specificity and salinity tolerance remain insufficiently understood. In this study, a genome-centric metagenome analysis was conducted on the microbiome from a leachate treatment facility at a closed landfill. RESULTS: A total of 68 metagenome-assembled genomes (MAGs) were reconstructed, including 64 putative novel species. Among these, two Nitrospira MAGs were recovered: a novel complete ammonia-oxidizing bacterium (comammox), Nitrospira LAS72 (88.72% completeness, 2.10% contamination), and canonical nitrite-oxidizing Nitrospira LAS18 (99.98% completeness, 2.29% contamination). Comparative genomic analysis with 260 publicly available Nitrospira genomes revealed that LAS18 represents a new sub-lineage within lineage VII of the Nitrospira genus. Two ammonia-oxidizing archaea (AOA), Candidatus Nitrosocosmicus LAS21 and Nitrosarchaeum LAS73, were also identified, while canonical ammonia-oxidizing bacteria were not detected. Given the brackish conditions (1.23% salinity) and the methanol-fed operation of the treatment facility, the genomic potential for osmotic stress adaptation and methanol metabolism was investigated. Comammox Nitrospira LAS72 harbors biosynthetic pathways for several compatible solutes (osmoprotectants), including glycine betaine, proline, trehalose, and L-glutamate. Moreover, comammox Nitrospira LAS72 possesses genetic potential for oxidizing formaldehyde, suggesting that it may exploit these methanol-derived intermediates as energy sources. These features indicate that LAS72 may withstand osmotic fluctuations through the production of various osmoprotectants and thrive under the unique conditions of a methanol-fed environment. CONCLUSIONS: The discovery of novel comammox Nitrospira and canonical Nitrospira forming a new sub-lineage within lineage VII of the Nitrospira genus in an ammonia-rich brackish environment provides the first genomic evidence for evolutionary adaptation among nitrifiers to saline, methanol-fed environments. These findings enhance our understanding of the ecological and evolutionary dynamics shaping nitrifier communities in complex treatment ecosystems. Video Abstract.

Ammonia↗

Exercise Therapy in Down Syndrome: A Systematic Review and Meta-Analysis Focused on Muscle Strength, Redox Balance, and Inflammatory Profile.

OBJECTIVE: This study systematically reviewed and meta-analyzed randomized and quasi-randomized controlled trials investigating the impact of exercise therapy on muscle strength, redox balance, and inflammatory profile in individuals with Down syndrome. DESIGN: Systematic review and meta-analysis. DATA SOURCES: Cochrane Central Register of Controlled Trials, MEDLINE, CINAHL, SPORTDiscus, EMBASE, and PEDro. ELIGIBILITY CRITERIA FOR SELECTING STUDIES: Randomized and quasi-randomized controlled trials exploring exercise therapy effects on muscle strength and redox balance in individuals with Down syndrome. Although no initial restrictions on age, gender, or health condition were applied during the search process, all included studies focused on adult participants (>18 yr old). No language restrictions were applied, and the search covered the period from 1970 to 2021. RESULTS: We assessed the abstract of 1964 studies. Of the 46 studies meeting the inclusion criteria for the period 2004-2021, 32 focused on muscle strength, and 14 examined redox balance and inflammation. A total of 1611 participants with a mean age of 27 yr were included. This review confirmed that different exercise modalities are prone to improve muscle strength (random effect (95% confidence interval): 0.66, 0.54 to 0.78), redox balance and inflammatory profile (random effect (95% confidence interval): -1.04, -1.31 to -0.76) in this population. The multimodel inference suggested that the frequency of training (times per week) might play a significant role in the main effect. Unsupervised machine learning algorithms displayed a pattern-based graphic representation to assess heterogeneity. CONCLUSIONS: Exercise training demonstrated a positive impact on muscle strength in adults with Down syndrome. The review provides valuable insights into the effects of exercise therapy on individuals with Down syndrome, emphasizing the need for tailored training prescriptions.

Humans↗

Repair of oxidized methionine residues in the chaperone Spy maintains periplasmic proteostasis under chlorite stress in Escherichia coli.

The bacterial cell envelope is exposed to various stresses, including oxidative stress caused by different types of oxidants, such as reactive oxygen species (ROS) and reactive chlorine species (RCS). In Escherichia coli, the reduction of chlorate into chlorite, a toxic RCS compound, induces the expression of the MsrPQ system, which repairs periplasmic proteins oxidized at methionine residues (methionine sulfoxide, Met-O). In this study, using a proteomic-based approach, we show that chlorite stress also triggers the overproduction of the periplasmic molecular chaperone Spheroplast Protein Y (Spy). This response is mediated by the activation of the BaeSR two-component system. Furthermore, both in vivo and in vitro evidence reveal that Spy's susceptibility to oxidation is critical for its chaperone activity. We demonstrate that the MsrPQ repair system ensures Spy's functionality by reducing its Met-O, thereby safeguarding its role in periplasmic protein homeostasis. Overall, this work reveals Spy as a key target of chlorite-induced oxidative damage and underscores the essential role of MsrPQ in preserving periplasmic protein quality control.

Methionine↗

Ecdysone Oxidase, an enzyme from the blowfly Calliphora erythrocephala (Meigen).

In the blowfly, the formation of 3-dehydroecdysone from the insect molting hormone ecdysone is catalyzed by an enzyme which carries hydrogen from ecdysone and ecdysterone to oxygen. The enzyme is therefore called "ecdysone oxidase". Two methods are described for the detection of ecdysone oxidase activity, one using a radiolabelled substrate which is separated from the product by thin-layer chromatography after the reaction, and the other using dichloroindophenol, which is discoloured by the redox reaction. The ecdysone oxidase is purified by a factor of 2200 from prepupae of Calliphora erythrocephala using salt precipitation and ion exchange chromatography. The ecdysone oxidase has a Km value for ecdysone of 42muM. The pH optimum is 6.5. The temperature optimum lies at 45 degrees C. The ecdysone oxidase has a molecular weight of 240000.

2,6-Dichloroindophenol↗

Distribution and removal of added mercury in milk.

Distribution patterns of added mercury in raw whole milk after equilibration for 30 min and 2 h at 37 C showed a distribution among acid casein, whey proteins, fat globule membrane, and soluble fat globule membrane of 33, 28, 16, and 2%. On the basis of protein content, the fat globule membrane had the highest amount of mercury. Mercury added to milk as mercuric chloride was removed by treatment with thiolated aminoethyl celluloses and reduced human hair. In a 5 min treatment, 70, 43, and 41% of the mercury was removed by thiosuccinylated aminoethyl cellulose, thionitrocarboxyphenylated aminoethyl cellulose, and reduced human hair, respectively, from whole milk initially containing 1 ppm mercury and equilibrated for 2 h at 37 C prior to treatment. After treatment for 60 min, 82, 52, and 64% of the mercury was removed by thiosuccinilated aminoethyl cellulose, thionitrocarboxyphenylated aminoethyl cellulose, and reduced hair, respectively. However, increasing incubation temperature and time prior to treatment decreased the removal efficiencies. Thiosuccinilated aminoethyl cellulose and reduced human hair showed increasing efficiency directly with pH, while thionitrocarboxyphenylated aminoethyl cellulose showed the opposite effect and had higher affinity for mercury at pH 5.5 than at pH 7.5. Moreover, the rate of removal of mercury at 4 C compared to 37 C was much slower. The removal of mercury from soluble casein and soluble whey proteins was more efficient than from micellar casein. Protein, lactose content, and pH of milk were not changed by the polymer treatments.

Animals↗

Molecular and Physiological Insights into CAT- and SOD-Associated Redox Homeostasis Under Salt Stress in Artemisia argyi.

Soil salinity disrupts redox homeostasis and limits plant growth and development. Although catalase (CAT) and superoxide dismutase (SOD) are key enzymatic antioxidants, the CAT and SOD gene families have not been characterized in Artemisia argyi (A. argyi), a species of medicinal and ecological importance. While SOD and CAT serve as the primary enzymatic scavengers for reactive oxygen species (ROS) detoxification, their genomic architecture and stress-responsive regulatory networks in A. argyi have remained uncharacterized. In this study, we conducted the first comprehensive genome-wide analysis of these gene families in A. argyi, identifying 22 structurally conserved members (8 AarCATs and 14 AarSODs). Collinearity and synteny analyses revealed strict lineage-specific evolutionary conservation, while tertiary protein modeling and subcellular localization illustrated a highly organized multi-organelle defense compartmentalization. High salinity (up to 200 mM NaCl) reduced the stomatal conductance and net photosynthetic rate. Salt stress reduced growth and increased osmoprotectant and antioxidant accumulation in A. argyi. Furthermore, histochemical staining using nitroblue tetrazolium (NBT) and 3,3'-Diaminobenzidine (DAB) provided comprehensive evidence of significant accumulation of ROS in leaves, which indicates the intense oxidative stress triggered by ionic stress. Tissue-specific analysis revealed that AarCAT1, AarCSD1, and AarFSD2 were 3.9-, 7.9-, and 12.7-fold higher in leaves than in roots, respectively. Under stress, AarCAT6 and AarCSD1 were strongly repressed in leaves by ~50% and ~46-70%, respectively, whereas AarMSD2 and AarMSD3 were significantly induced in roots by ~2.2- and ~1.8-fold. These distinct expression patterns suggest their potential involvement in tissue-specific stress adaptation and ROS homeostasis. These findings uncover the evolutionary and physiological basis of salt tolerance in A. argyi, providing genetic targets for climate-resilient breeding.

Artemisia↗

Redox Regulation in Glioblastoma: Mechanisms, Biomarkers, and Therapeutic Implications.

Glioblastoma is the most aggressive primary tumor of the central nervous system, characterized by high invasiveness, rapid progression, and a poor prognosis despite the current treatment modalities. Molecular stratification, using biomarkers such as IDH1, TERT, and MGMT, is a crucial step in the 2021 WHO classification for improving diagnosis and prognosis. Oxidative stress, a feature of GB, has been identified as an important factor in the initiation, progression, and resistance to treatment. It occurs due to an imbalance between reactive oxygen species generated by mitochondrial metabolism, NADPH oxidases, and exogenous sources such as ionizing radiation and xenobiotics and antioxidant defense. This imbalance leads to DNA damage, genomic instability, and deregulation of signaling pathways involved in cell proliferation, apoptosis, and tumor progression. This review provides an overview of key oxidative stress biomarkers and their dual roles in tumor suppression and progression. It highlights how oxidative stress contributes to treatment responses and resistance to current GB treatments, including redox-adaptive mechanisms such as the Nrf2-Keap1 pathway, which promotes radioresistance. Finally, it discusses the potential of understanding these mechanisms to develop therapeutic strategies that target redox balance and homeostasis, aiming to overcome resistance and improve survival outcomes for glioblastoma patients.

Humans↗

[The effect of oxidazable substrates and ATP on the sensitivity of certain energy-dependent functions submitochondrial particles to phospholipases A, C and D].

The effect of NADH, succinate and ATP on the sensitivity of a number of energy-dependent functions of submitochondrial particles ot phospholipases A, C and D has been studied. It has been shown that in the conditions of oxidation of NADH and succinate by oxygen and also of ATP hydrolysis, the decrease in the phosphorylating activity of the particles under the action of phospholipases C and D accelerates. No such acceleration has been observed with phospholipase A. For other two functions, i. e. reverse electron transfer (ATP-dependent NAD+ reduction by succinate) and ATP-dependent transhydrogenase reaction the results proved to be different. Oxidizable substrates and ATP promoted the maintenance of these functions in the presence of phospholipase A, but did not retard their suppression by phospholipases C and D. The effects of NADH, succinate and ATP on the sensitivity of different energy-dependent functions of submitochondrial particles to phospholipases A, C and D could be removed by the uncoupling agent carbonyl cyanide-m-chlorophenyl hydrazone. The conclusion is made that the effects revealed are associated with an increase in the sensitivity of coupling sites II PAND/OR III to phospholipases C and D and with a decrease in the sensitivity of sites I and IV to phospholipase A on energization of submitochondrial particles.

Adenosine Triphosphate↗

Oxidative biotransformation of 2-acetylaminofluorene in fetal and placental tissues of humans and monkeys. Correlations with aryl hydrocarbon hydroxylase activities.

The mixed-function oxidation of 14C-labled 2-acetylaminofluorene (AAF) was investigated in placental and fetal tissues of humans and monkeys (Macaca nemestrina) in vitro. The major metabolite formed in most tissues was 7-hydroxy-AAF. Rates of the hydroxylation reactions varied widely among the tissues investigated and were generally one to two orders of magnitude lower than those measured in rat hepatic tissues. High correlations among rates of 7-,5-, and 3- and between 1- and N-hydroxylations of AAF were observed. The latter two reactions were less responsive to inhibition by carbon monoxide. Rates of 3-hydroxylations of benzo[a]pyrene (BP) also were highly correlated with rates of 7-, 5-, and 3-hydroxylations of AAF but were not correlated with rates of 1- and N-hydroxylations in human placental microsomes. A lack of statistically significant correlations was observed among rates of many of these hydroxylation reactions studied in primate fetal tissues. Rates of 7-, 5-, and 3-hydroxylations of AAF were not statistically correlated with rates of 3-hydroxylation of BP in homogenates of primate fetal tissues in most instances, but statistically significant correlations among rates of 3-hydroxylation of BP and 1- and N-hydroxylations of AAF were observed in those preparations. The results suggested two separate mechanisms for the genetic control of rates of placental aromatic ring- and N-hydroxylation reactions as opposed to apparent multiple genetic controls for rates of these hydroxylation reactions in primate fetal tissues.

2-Acetylaminofluorene↗

[Analysis of inhibition in pathways of NADP.H2 and NAD.H2 oxidation in liver tissue microsomes].

Microsomal complexes of electron transfer were resistant to typical inhibitors of mitochondrial pathway of electron transport. In oxidation of NADP.H2 there were at least three point of molecular O2 reduction: NADP.H2-specific flavoprotein, Fe2+ participating in reactions of peroxidation of unsaturated fatty acids and cytochrome P-450. Efficiency of cytochrome P-450 inhibitors could not be evaluated by polarography as in the pathway several sites of molecular O2 activation were observed. In oxidation of NADP.H2 estimation of the rate of electron transfer reactions was carried out by monitoring of velocity of O2 absorption in presence of EDTA (inhibitor of the reaction of peroxidation) because about 50% of the total oxygen were utilized only in the process where NADP.H2 was oxidized. NAD.H2 oxidation, inhibited with EDTA, was activated by addition of Ca2+.

Animals↗

[Nitrates and nitrites in plants].

The formation of aminoacids and proteins from the nitrogen which enters the roots as nitra t involves a complex reaction requiring energy. The first step requires a metalloflavoprotein, the nitrate reductase and the successive intervention of NADPH, FAD and reduced molybdenum which transfers electrons to nitrate and reduces it to nitrite. The following steps involve NADPH, FAD, Copper, Iron and Manganese, the last steps of the successive reductions being ammonia, needed for the aminoacids synthesis. The activity of the different enzymes are under the dependence of the genetic equipment of the plant, of the nitrogen and oligo-element nutrition and of the different factors acting on the photosynthesis.

Amino Acids↗

[Content of nicotinamide coenzymes in rat liver under conditions of nicotinamide administration].

The content of NAD+, NADH, NADP+, NADPH in the liver of normal, fasting rats, those on the low-carbohydrate diet and suffering from alloxan diabetes was studied as affected by nictotinamide. Changes in the NAD+ content, sum of nicotinamide coenzymes, the [NAD+] + [NADP+]/[NADH] +/- [NADPH] and [NAD+] + [NADH] (sum of nicotinamide coenzymes) ratios are mainly due to nicotinamide administration. Changes in the content of reduced forms of both nucleotides depend equally on nicotinamide administration and the physiological state of animals. Response of the rat organism to nicotinamide administration consists in a sharp intensification of NAD+ synthesis and in a less pronounced intensification of NADH, NADP+ and NADPH synthesis.

Animals↗