Identification of a family of species-selective complex I inhibitors as potential anthelmintics.

Soil-transmitted helminths (STHs) are major pathogens infecting over a billion people. There are few classes of anthelmintics and there is an urgent need for new drugs. Many STHs use an unusual form of anaerobic metabolism to survive the hypoxic conditions of the host gut. This requires rhodoquinone (RQ), a quinone electron carrier. RQ is not made or used by vertebrate hosts making it an excellent therapeutic target. Here we screen 480 structural families of natural products to find compounds that kill Caenorhabditis elegans specifically when they require RQ-dependent metabolism. We identify several classes of compounds including a family of species-selective inhibitors of mitochondrial respiratory complex I. These identified complex I inhibitors have a benzimidazole core and we determine key structural requirements for activity by screening 1,280 related compounds. Finally, we show several of these compounds kill adult STHs. We suggest these species-selective complex I inhibitors are potential anthelmintics.

Heat stress-induced NO enhanced perylenequinone biosynthesis of Shiraia sp. via calcium signaling pathway.

Perylenequinones (PQs) are natural photosensitizing compounds used as photodynamic therapy, and heat stress (HS) is the main limiting factor of mycelial growth and secondary metabolism of fungi. This study aimed to unravel the impact of HS-induced Ca2+ and the calcium signaling pathway on PQ biosynthesis of Shiraia sp. Slf14(w). Meanwhile, the intricate interplay between HS-induced NO and Ca2+ and the calcium signaling pathway was investigated. The outcomes disclosed that Ca2+ and the calcium signaling pathway activated by HS could effectively enhance the production of PQs in Shiraia sp. Slf14(w). Further investigations elucidated the specific mechanism through which NO signaling molecules induced by HS act upon the Ca2+/CaM (calmodulin) signaling pathway, thus propelling PQ biosynthesis in Shiraia sp. Slf14(w). This was substantiated by decoding the downstream positioning of the CaM/CaN (calcineurin) pathway in relation to NO through comprehensive analyses encompassing transcript levels, enzyme assays, and the introduction of chemical agents. Concurrently, the engagement of Ca2+ and the calcium signaling pathway in heat shock signaling was also evidenced. The implications of our study underscore the pivotal role of HS-induced Ca2+ and the calcium signaling pathway, which not only participate in heat shock signal transduction but also play an instrumental role in promoting PQ biosynthesis. Consequently, our study not only enriches our comprehension of the mechanisms driving HS signaling transduction in fungi but also offers novel insights into the PQ synthesis paradigm within Shiraia sp. Slf14(w). KEY POINTS: • The calcium signaling pathway was proposed to participate in PQ biosynthesis under HS. • HS-induced NO was revealed to act upon the calcium signaling pathway for the first time.

Quinones-enhanced humification in food waste composting: A novel strategy for hazard mitigation and nitrogen retention.

The harmless and high-value conversion of organic waste are the core problems to be solved by composting technology. This study introduced an innovative method of promoting targeted humification and nitrogen retention in composting by adding p-benzoquinone (PBQ), the composting without any additives was set as control group (CK). The results indicated that the addition of exogenous quinones led to a 30.1% increase in humic acid (HA) content during the heating and thermophilic phases of composting. Spectroscopic analyses confirmed that exogenous quinones form the core skeleton structure of amino-quinones in HA through composting biochemical reactions. This accelerated the transformation of quinones into recalcitrant HA in the early stages of composting, and reduced CO2 and NH3 by 8% and 78%, respectively. Redundancy analysis (RDA) revealed that the decrease in carbon and nitrogen losses primarily correlated with quinones enhancing HA formation and greater nitrogen incorporation into HA (P < 0.05). Furthermore, the compost treated with quinones demonstrated a decrease in phytotoxicity and earthworm mortality, alongside a significant increase in the relative abundance of actinobacteria, which are associated with the humification process. This research establishes and proposes that co-composting with quinones-containing waste is an effective approach for the sustainable recycling of hazardous solid waste.

Mitochondrial quinone redox states as a marker of mitochondrial metabolism.

Mitochondrial and thus cellular energetics are highly regulated both thermodynamically and kinetically. Cellular energetics is of prime importance in the regulation of cellular functions since it provides ATP for their accomplishment. However, cellular energetics is not only about ATP production but also about the ability to re-oxidize reduced coenzymes at a proper rate, such that the cellular redox potential remains at a level compatible with enzymatic reactions. However, this parameter is not only difficult to assess due to its dual compartmentation (mitochondrial and cytosolic) but also because it is well known that most NADH in the cells is bound to the enzymes. In this paper, we investigated the potential relevance of mitochondrial quinones redox state as a marker of mitochondrial metabolism and more particularly mitochondrial redox state. We were able to show that Q2 is an appropriate redox mediator to assess the mitochondrial quinone redox states. On isolated mitochondria, the mitochondrial quinone redox states depend on the mitochondrial substrate and the mitochondrial energetic state (phosphorylating or not phosphorylating). Last but not least, we show that the quinones redox state response allows to better understand the Krebs cycle functioning and respiratory substrates oxidation. Taken together, our results suggest that the quinones redox state is an excellent marker of mitochondrial metabolism.

Ongoing evolution of the Mycobacterium tuberculosis lactate dehydrogenase reveals the pleiotropic effects of bacterial adaption to host pressure.

The bacterial determinants that facilitate Mycobacterium tuberculosis (Mtb) adaptation to the human host environment are poorly characterized. We have sought to decipher the pressures facing the bacterium in vivo by assessing Mtb genes that are under positive selection in clinical isolates. One of the strongest targets of selection in the Mtb genome is lldD2, which encodes a quinone-dependent L-lactate dehydrogenase (LldD2) that catalyzes the oxidation of lactate to pyruvate. Lactate accumulation is a salient feature of the intracellular environment during infection and lldD2 is essential for Mtb growth in macrophages. We determined the extent of lldD2 variation across a set of global clinical isolates and defined how prevalent mutations modulate Mtb fitness. We show the stepwise nature of lldD2 evolution that occurs as a result of ongoing lldD2 selection in the background of ancestral lineage-defining mutations and demonstrate that the genetic evolution of lldD2 additively augments Mtb growth in lactate. Using quinone-dependent antibiotic susceptibility as a functional reporter, we also find that the evolved lldD2 mutations functionally increase the quinone-dependent activity of LldD2. Using 13C-lactate metabolic flux tracing, we find that lldD2 is necessary for robust incorporation of lactate into central carbon metabolism. In the absence of lldD2, label preferentially accumulates in dihydroxyacetone phosphate (DHAP) and glyceraldehyde-3-phosphate (G3P) and is associated with a discernible growth defect, providing experimental evidence for accrued lactate toxicity via the deleterious buildup of sugar phosphates. The evolved lldD2 variants increase lactate incorporation to pyruvate while altering triose phosphate flux, suggesting both an anaplerotic and detoxification benefit to lldD2 evolution. We further show that the mycobacterial cell is transcriptionally sensitive to the changes associated with altered lldD2 activity which affect the expression of genes involved in cell wall lipid metabolism and the ESX- 1 virulence system. Together, these data illustrate a multifunctional role of LldD2 that provides context for the selective advantage of lldD2 mutations in adapting to host stress.

Evidence for cytochrome P450 3A4-mediated metabolic activation of SCO-267.

SCO-267 is a potent G-protein-coupled receptor 40 agonist that is undergoing clinical development for the treatment of type 2 diabetes mellitus. The current work was undertaken to investigate the bioactivation potential of SCO-267 in vitro and in vivo. Three SCO-267-derived glutathione (GSH) conjugates (M1-M3) were found both in rat and human liver microsomal incubations supplemented with GSH and nicotinamide adenine dinucleotide phosphate. Two GSH conjugates (M1-M2) together with two N-acetyl-cysteine conjugates (M4-M5) were detected in the bile of rats receiving SCO-267 at 10 mg/kg. The identified conjugates suggested the generation of quinone-imine and ortho-quinone intermediates. CYP3A4 was demonstrated to primarily catalyze the bioactivation of SCO-267. In addition, SCO-267 concentration-, time-, and NADPH-dependently inactivated CYP3A in human liver microsomes using testosterone as a probe substrate, along with KI and kinact values of 4.91 µM and 0.036 min-1 , respectively. Ketoconazole (a competitive inhibitor of CYP3A) displayed no significant protective effect on SCO-267-induced CYP3A inactivation. However, inclusion of GSH showed significant protection. These findings revealed that SCO-267 undergoes a facile CYP3A4-catalyzed bioactivation with the generation of quinone-imine and ortho-quinone intermediates, which were assumed to be involved in SCO-267 induced CYP3A inactivation. These findings provide further insight into the bioactivation pathways involved in the generation of reactive, potentially toxic metabolites of SCO-267. Further studies are needed to evaluate the influence of SCO-267 metabolism on the safety of this drug in vivo.

Residual Complex I activity and amphidirectional Complex II operation support glutamate catabolism through mtSLP in anoxia.

Anoxia halts oxidative phosphorylation (OXPHOS) causing an accumulation of reduced compounds in the mitochondrial matrix which impedes dehydrogenases. By simultaneously measuring oxygen concentration, NADH autofluorescence, mitochondrial membrane potential and ubiquinone reduction extent in isolated mitochondria in real-time, we demonstrate that Complex I utilized endogenous quinones to oxidize NADH under acute anoxia. 13C metabolic tracing or untargeted analysis of metabolites extracted during anoxia in the presence or absence of site-specific inhibitors of the electron transfer system showed that NAD+ regenerated by Complex I is reduced by the 2-oxoglutarate dehydrogenase Complex yielding succinyl-CoA supporting mitochondrial substrate-level phosphorylation (mtSLP), releasing succinate. Complex II operated amphidirectionally during the anoxic event, providing quinones to Complex I and reducing fumarate to succinate. Our results highlight the importance of quinone provision to Complex I oxidizing NADH maintaining glutamate catabolism and mtSLP in the absence of OXPHOS.

Inverted region in the reaction of the quinone reduction in the A1-site of photosystem I from cyanobacteria.

Photosystem I from the menB strain of Synechocystis sp. PCC 6803 containing foreign quinones in the A1 sites was used for studying the primary steps of electron transfer by pump-probe femtosecond laser spectroscopy. The free energy gap (- ΔG) of electron transfer between the reduced primary acceptor A0 and the quinones bound in the A1 site varied from 0.12 eV for the low-potential 1,2-diamino-anthraquinone to 0.88 eV for the high-potential 2,3-dichloro-1,4-naphthoquinone, compared to 0.5 eV for the native phylloquinone. It was shown that the kinetics of charge separation between the special pair chlorophyll P700 and the primary acceptor A0 was not affected by quinone substitutions, whereas the rate of A0 → A1 electron transfer was sensitive to the redox-potential of quinones: the decrease of - ΔG by 400 meV compared to the native phylloquinone resulted in a ~ fivefold slowing of the reaction The presence of the asymmetric inverted region in the ΔG dependence of the reaction rate indicates that the electron transfer in photosystem I is controlled by nuclear tunneling and should be treated in terms of quantum electron-phonon interactions. A three-mode implementation of the multiphonon model, which includes modes around 240 cm-1 (large-scale protein vibrations), 930 cm-1 (out-of-plane bending of macrocycles and protein backbone vibrations), and 1600 cm-1 (double bonds vibrations) was applied to rationalize the observed dependence. The modes with a frequency of at least 1600 cm-1 make the predominant contribution to the reorganization energy, while the contribution of the "classical" low-frequency modes is only 4%.

6PPD-quinone exposure induces neuronal mitochondrial dysfunction to exacerbate Lewy neurites formation induced by α-synuclein preformed fibrils seeding.

The emerging toxicant N-(1,3-dimethylbutyl)-N'-phenyl-p-phenylenediamine quinone (6PPD-Q) is of wide concern due to its ubiquitous occurrence and high toxicity. Despite regular human exposure, limited evidence exists about its presence in the body and potential health risks. Herein, we analyzed cerebrospinal fluid (CSF) samples from Parkinson's disease (PD) patients and controls. The CSF levels of 6PPD-Q were twice as high in PD patients compared to controls. Immunostaining assays performed with primary dopaminergic neurons confirm that 6PPD-Q at environmentally relevant concentrations can exacerbate the formation of Lewy neurites induced by α-synuclein preformed fibrils (α-syn PFF). Assessment of cellular respiration reveals a considerable decrease in neuronal spare respiratory and ATP-linked respiration, potentially due to changes in mitochondrial membrane potential. Moreover, 6PPD-Q-induced mitochondrial impairment correlates with an upsurge in mitochondrial reactive oxygen species (mROS), and Mito-TEMPO-driven scavenging of mROS can lessen the amount of pathologic phospho-serine 129 α-synuclein. Untargeted metabolomics provides supporting evidence for the connection between 6PPD-Q exposure and changes in neuronal metabolite profiles. In-depth targeted metabolomics further unveils an overall reduction in glycolysis metabolite pool and fluctuations in the quantity of TCA cycle intermediates. Given its potentially harmful attributes, the presence of 6PPD-Q in human brain could potentially be a risk factor for PD.

Functional characterization of CYP81C16 involved in the tanshinone biosynthetic pathway in Salvia miltiorrhiza.

Danshen, the dried roots and rhizomes of Salvia miltiorrhiza Bunge (S. miltiorrhiza), is widely used in the treatment of cardiovascular and cerebrovascular diseases. Tanshinones, the bioactive compounds from Danshen, exhibit a wide spectrum of pharmacological properties, suggesting their potential for future therapeutic applications. Tanshinone biosynthesis is a complex process involving at least six P450 enzymes that have been identified and characterized, most of which belong to the CYP76 and CYP71 families. In this study, CYP81C16, a member of the CYP71 clan, was identified in S. miltiorrhiza. An in vitro assay revealed that it could catalyze the hydroxylation of four para-quinone-type tanshinones, namely neocryptotanshinone, deoxyneocryptotanshinone, and danshenxinkuns A and B. SmCYP81C16 emerged as a potential broad-spectrum oxidase targeting the C-18 position of para-quinone-type tanshinones with an impressive relative conversion rate exceeding 90%. Kinetic evaluations andin vivo assays underscored its highest affinity towards neocryptotanshinone among the tested substrates. The overexpression of SmCYP81C16 promoted the accumulation of (iso)tanshinone in hairy root lines. The characterization of SmCYP81C16 in this study accentuates its potential as a pivotal tool in the biotechnological production of tanshinones, either through microbial or plant metabolic engineering.

How a natural antibiotic uses oxidative stress to kill oxidant-resistant bacteria.

Natural products that possess antibiotic and antitumor qualities are often suspected of working through oxidative mechanisms. In this study, two quinone-based small molecules were compared. Menadione, a classic redox-cycling compound, was confirmed to generate high levels of reactive oxygen species inside Escherichia coli. It inactivated iron-cofactored enzymes and blocked growth. However, despite the substantial levels of oxidants that it produced, it was unable to generate significant DNA damage and was not lethal. Streptonigrin, in contrast, was poorer at redox cycling and did not inactivate enzymes or block growth; however, even in low doses, it damaged DNA and killed cells. Its activity required iron and oxygen, and in vitro experiments indicated that its quinone moiety transferred electrons through the adjacent iron atom to oxygen. Additionally, in vitro experiments revealed that streptonigrin was able to damage DNA without inhibition by catalase, indicating that hydrogen peroxide was not involved. We infer that streptonigrin can reduce bound oxygen directly to a ferryl species, which then oxidizes the adjacent DNA, without release of superoxide or hydrogen peroxide intermediates. This scheme allows streptonigrin to kill a bacterial cell without interference by scavenging enzymes. Moreover, its minimal redox-cycling behavior avoids alerting either the OxyR or the SoxRS systems, which otherwise would block killing. This example highlights qualities that may be important in the design of oxidative drugs. These results also cast doubt on proposals that bacteria can be killed by stressors that merely stimulate intracellular O2- and H2O2 formation.

Characterization of the inner membrane cytochrome ImcH from Geobacter reveals its importance for extracellular electron transfer and energy conservation.

Electroactive bacteria combine the oxidation of carbon substrates with an extracellular electron transfer (EET) process that discharges electrons to an electron acceptor outside the cell. This process involves electron transfer through consecutive redox proteins that efficiently connect the inner membrane to the cell exterior. In this study, we isolated and characterized the quinone-interacting membrane cytochrome c ImcH from Geobacter sulfurreducens, which is involved in the EET process to high redox potential acceptors. Spectroscopic and electrochemical studies show that ImcH hemes have low midpoint redox potentials, ranging from -150 to -358 mV, and connect the oxidation of the quinol-pool to EET, transferring electrons to the highly abundant periplasmic cytochrome PpcA with higher affinity than to its homologues. Despite the larger number of hemes and transmembrane helices, the ImcH structural model has similarities with the NapC/NirT/NrfH superfamily, namely the presence of a quinone-binding site on the P-side of the membrane. In addition, the first heme, likely involved on the quinol oxidation, has apparently an unusual His/Gln coordination. Our work suggests that ImcH is electroneutral and transfers electrons and protons to the same side of the membrane, contributing to the maintenance of a proton motive force and playing a central role in recycling the menaquinone pool.

Machine Learning Enables Accurate Prediction of Quinone Formation during Drug Metabolism.

Metabolism helps in the elimination of drugs from the human body by making them more hydrophilic. Sometimes, drugs can be bioactivated to highly reactive metabolites or intermediates during metabolism. These reactive metabolites are often responsible for the toxicities associated with the drugs. Identification of reactive metabolites of drug candidates can be very helpful in the initial stages of drug discovery. Quinones are soft electrophiles that are generated as reactive intermediates during metabolism. Quinones make up more than 40% of the reactive metabolites. In this work, a reliable data set of 510 molecules was used to develop machine learning and deep learning-based predictive models to predict the formation of quinone-type metabolites. For representing molecules, two-dimensional (2D) descriptors, PubChem fingerprints, electro-topological state (E-state) fingerprints, and metabolic reactivity-based descriptors were used. Developed models were compared to the existing Xenosite web server using the untouched test set of 102 molecules. The best model achieved an accuracy of 86.27%, while the Xenosite server could achieve an accuracy of only 52.94% on the test set. Descriptor analysis revealed that the presence of greater numbers of polar moieties in a molecule can prevent the formation of quinone-type metabolites. In addition, the presence of a nitrogen atom in an aromatic ring and the presence of metabolophores V51, V52, and V53 (SMARTCyp descriptors) decrease the probability of quinone formation. Finally, a tool based on the best machine learning models was developed, which is accessible at http://14.139.57.41/quinonepred/.

Bamboo polysaccharides elicit hypocrellin A biosynthesis of a bambusicolous fungus Shiraia sp. S9.

Hypocrellin A (HA), a fungal perylenequinone from bambusicolous Shiraia species, is a newly developed photosensitizer for photodynamic therapy in cancer and other infectious diseases. The lower yield of HA is an important bottleneck for its biomedical application. This study is the first report of the enhancement of HA production in mycelium culture of Shiraia sp. S9 by the polysaccharides from its host bamboo which serve as a strong elicitor. A purified bamboo polysaccharide (BPSE) with an average molecular weight of 34.2 kDa was found to be the most effective elicitor to enhance fungal HA production and characterized as a polysaccharide fraction mainly composed of arabinose and galactose (53.7: 36.9). When BPSE was added to the culture at 10 mg/L on day 3, the highest HA production of 422.8 mg/L was achieved on day 8, which was about 4.0-fold of the control. BPSE changed the gene expressions mainly responsible for central carbon metabolism and the cellular oxidative stress. The induced generation of H2O2 and nitric oxide was found to be involved in both the permeabilization of cell membrane and HA biosynthesis, leading to enhancements in both intra- and extracellular HA production. Our results indicated the roles of plant polysaccharides in host-fungal interactions and provided a new elicitation technique to improve fungal perylenequinone production in mycelium cultures.

Biosynthesis and function of microbial methylmenaquinones.

The membranous quinone/quinol pool is essential for the majority of life forms and its composition has been widely used as a biomarker in microbial taxonomy. The most abundant quinone is menaquinone (MK), which serves as an essential redox mediator in various electron transport chains of aerobic and anaerobic respiration. Several methylated derivatives of MK, designated methylmenaquinones (MMKs), have been reported to be present in members of various microbial phyla possessing either the classical MK biosynthesis pathway (Men) or the futalosine pathway (Mqn). Due to their low redox midpoint potentials, MMKs have been proposed to be specifically involved in appropriate electron transport chains of anaerobic respiration. The class C radical SAM methyltransferases MqnK, MenK and MenK2 have recently been shown to catalyse specific MK methylation reactions at position C-8 (MqnK/MenK) or C-7 (MenK2) to synthesise 8-MMK, 7-MMK and 7,8-dimethylmenaquinone (DMMK). MqnK, MenK and MenK2 from organisms such as Wolinella succinogenes, Adlercreutzia equolifaciens, Collinsella tanakaei, Ferrimonas marina and Syntrophus aciditrophicus have been functionally produced in Escherichia coli, enabling extensive quinone/quinol pool engineering of the native MK and 2-demethylmenaquinone (DMK). Cluster and phylogenetic analyses of available MK and MMK methyltransferase sequences revealed signature motifs that allowed the discrimination of MenK/MqnK/MenK2 family enzymes from other radical SAM enzymes and the identification of C-7-specific menaquinone methyltransferases of the MenK2 subfamily. It is envisaged that this knowledge will help to predict the methylation status of the menaquinone/menaquinol pool of any microbial species (or even a microbial community) from its (meta)genome.

Differential Pattern of Cell Death and ROS Production in Human Airway Epithelial Cells Exposed to Quinones Combined with Heated-PM2.5 and/or Asian Sand Dust.

The combined toxicological effects of airborne particulate matter (PM), such as PM2.5, and Asian sand dust (ASD), with surrounding chemicals, particularly quinones, on human airway epithelial cells remain underexplored. In this study, we established an in vitro combination exposure model using 1,2-naphthoquinones (NQ) and 9,10-phenanthroquinones (PQ) along with heated PM (h-PM2.5 and h-ASD) to investigate their potential synergistic effects. The impacts of quinones and heated PM on tetrazolium dye (WST-1) reduction, cell death, and cytokine and reactive oxygen species (ROS) production were examined. Results revealed that exposure to 9,10-PQ with h-PM2.5 and/or h-ASD dose-dependently increased WST-1 reduction at 1 µM compared to the corresponding control while markedly decreasing it at 10 µM. Higher early apoptotic, late apoptotic, or necrotic cell numbers were detected in 9,10-PQ + h-PM2.5 exposure than in 9,10-PQ + h-ASD or 9,10-PQ + h-PM2.5 + h-ASD. Additionally, 1,2-NQ + h-PM2.5 exposure also resulted in an increase in cell death compared to 1,2-NQ + h-ASD and 1,2-NQ + h-PM2.5 + h-ASD. Quinones with or without h-PM2.5, h-ASD, or h-PM2.5 + h-ASD significantly increased ROS production, especially with h-PM2.5. Our findings suggest that quinones, at relatively low concentrations, induce cell death synergistically in the presence of h-PM2.5 rather than h-ASD and h-PM2.5 + h-ASD, partially through the induction of apoptosis with increased ROS generation.

Role of the Escherichia coli ubiquinone-synthesizing UbiUVT pathway in adaptation to changing respiratory conditions.

Isoprenoid quinones are essential for cellular physiology. They act as electron and proton shuttles in respiratory chains and various biological processes. Escherichia coli and many α-, ß-, and γ-proteobacteria possess two types of isoprenoid quinones: ubiquinone (UQ) is mainly used under aerobiosis, while demethylmenaquinones (DMK) are mostly used under anaerobiosis. Yet, we recently established the existence of an anaerobic O2-independent UQ biosynthesis pathway controlled by ubiT, ubiU, and ubiV genes. Here, we characterize the regulation of ubiTUV genes in E. coli. We show that the three genes are transcribed as two divergent operons that are both under the control of the O2-sensing Fnr transcriptional regulator. Phenotypic analyses using a menA mutant devoid of DMK revealed that UbiUV-dependent UQ synthesis is essential for nitrate respiration and uracil biosynthesis under anaerobiosis, while it contributes, though modestly, to bacterial multiplication in the mouse gut. Moreover, we showed by genetic study and 18O2 labeling that UbiUV contributes to the hydroxylation of ubiquinone precursors through a unique O2-independent process. Last, we report the crucial role of ubiT in allowing E. coli to shift efficiently from anaerobic to aerobic conditions. Overall, this study uncovers a new facet of the strategy used by E. coli to adjust its metabolism on changing O2 levels and respiratory conditions. This work links respiratory mechanisms to phenotypic adaptation, a major driver in the capacity of E. coli to multiply in gut microbiota and of facultative anaerobic pathogens to multiply in their host. IMPORTANCE Enterobacteria multiplication in the gastrointestinal tract is linked to microaerobic respiration and associated with various inflammatory bowel diseases. Our study focuses on the biosynthesis of ubiquinone, a key player in respiratory chains, under anaerobiosis. The importance of this study stems from the fact that UQ usage was for long considered to be restricted to aerobic conditions. Here we investigated the molecular mechanism allowing UQ synthesis in the absence of O2 and searched for the anaerobic processes that UQ is fueling in such conditions. We found that UQ biosynthesis involves anaerobic hydroxylases, that is, enzymes able to insert an O atom in the absence of O2. We also found that anaerobically synthesized UQ can be used for respiration on nitrate and the synthesis of pyrimidine. Our findings are likely to be applicable to most facultative anaerobes, which count many pathogens (Salmonella, Shigella, and Vibrio) and will help in unraveling microbiota dynamics.

NAD(P)H:Quinone Acceptor Oxidoreductase 1 (NQO1) Activatable Salicylamide H+ /Cl- Transporters.

NAD(P)H:quinone acceptor oxidoreductase 1 (NQO1), a detoxifying enzyme overexpressed in tumors, plays a key role in protecting cancer cells against oxidative stress and thus has been considered an attractive candidate for activating prodrug(s). Herein, we report the first use of NQO1 for the selective activation of 'protransporter' systems in cancer cells leading to the induction of apoptosis. Salicylamides, easily synthesizable small molecules, have been effectively used for efficient H+ /Cl- symport across lipid membranes. The ion transport activity of salicylamides was efficiently abated by caging the OH group with NQO1 activatable quinones via either ether or ester linkage. The release of active transporters, following the reduction of quinone caged 'protransporters' by NQO1, was verified. Both the transporters and protransporters exhibited significant toxicity towards the MCF-7 breast cancer line, mediated via the induction of oxidative stress, mitochondrial membrane depolarization, and lysosomal deacidification. Induction of cell death via intrinsic apoptotic pathway was verified by monitoring PARP1 cleavage.

Whole-genome sequence analysis reveals phenanthrene and pyrene degradation pathways in newly isolated bacteria Klebsiella michiganensis EF4 and Klebsiella oxytoca ETN19.

Polycyclic aromatic hydrocarbons (PAHs) are diverse pollutants of significant environmental concerns, requiring effective biodegradation. This study used different bioinformatics tools to conduct whole-genome sequencing of two novel bacterial strains, Klebsiella michiganensis EF4 and K. oxytoca ETN19, to improve our understanding of their many genomic functions and degradation pathways of phenanthrene and pyrene. After 28 days of cultivation, strain EF4 degraded approximately 80% and 60% of phenanthrene and pyrene, respectively. However, their combinations (EF4 +ETN19) showed tremendous phenanthrene degradation efficiency, supposed to be at the first-level kinetic model with a t1/2 value of approximately 6 days. In addition, the two bacterial genomes contained carbohydrate-active enzymes and secondary metabolites biosynthetic gene clusters associated with PAHs degradation. The two genomes contained the bZIP superfamily of transcription factors, primarily the cAMP-response element-binding protein (CREB), which could regulate the expression of several PAHs degradation genes and enzymes. Interestingly, the two genomes were found to uniquely degrade phenanthrene through a putative pathway that catabolizes 2-carboxybenzalpyruvate into the TCA cycle. An operon containing multicomponent proteins, including a novel gene (JYK05_14550) that could initiate the beginning step of phenanthrene and pyrene degradation, was found in the EF4 genome. However, the degradation pathway of ETN19 showed that the yhfP gene encoding putative quinone oxidoreductase was associated with phenanthrene and pyrene catabolic processes. Furthermore, the significant expression of catechol 1,2-dioxygenase and quinone oxidoreductase genes in EF4 +ETN19 and ETN19 following the quantitative reverse transcription polymerase chain reaction (qRT-PCR) analysis confirmed the ability of the bacteria combination to degrade pyrene and phenanthrene effectively. These findings present new insight into the possible co-metabolism of the two bacterial species in the rapid biodegradation of phenanthrene and pyrene in soil environments.

A sense-antisense RNA interaction promotes breast cancer metastasis via regulation of NQO1 expression.

Antisense RNAs are ubiquitous in human cells, yet their role is largely unexplored. Here we profiled antisense RNAs in the MDA-MB-231 breast cancer cell line and its highly lung metastatic derivative. We identified one antisense RNA that drives cancer progression by upregulating the redox enzyme NADPH quinone dehydrogenase 1 (NQO1), and named it NQO1-AS. Knockdown of either NQO1 or NQO1-AS reduced lung colonization in a mouse model, and investigation into the role of NQO1 indicated that it is broadly protective against oxidative damage and ferroptosis. Breast cancer cells in the lung are dependent on this pathway, and this dependence can be exploited therapeutically by inducing ferroptosis while inhibiting NQO1. Together, our findings establish a role for NQO1-AS in the progression of breast cancer by regulating its sense mRNA post-transcriptionally. Because breast cancer predominantly affects females, the disease models used in this study are of female origin and the results are primarily applicable to females.