The tricarboxylic acid (TCA) cycle: a malleable metabolic network to counter cellular stress.

The tricarboxylic acid (TCA) cycle is a primordial metabolic pathway that is conserved from bacteria to humans. Although this network is often viewed primarily as an energy producing engine fueling ATP synthesis via oxidative phosphorylation, mounting evidence reveals that this metabolic hub orchestrates a wide variety of pivotal biological processes. It plays an important part in combatting cellular stress by modulating NADH/NADPH homeostasis, scavenging ROS (reactive oxygen species), producing ATP by substrate-level phosphorylation, signaling and supplying metabolites to quell a range of cellular disruptions. This review elaborates on how the reprogramming of this network prompted by such abiotic stress as metal toxicity, oxidative tension, nutrient challenge and antibiotic insult is critical for countering these conditions in mostly microbial systems. The cross-talk between the stressors and the participants of TCA cycle that results in changes in metabolite and nucleotide concentrations aimed at combatting the abiotic challenge is presented. The fine-tuning of metabolites mediated by disparate enzymes associated with this metabolic hub is discussed. The modulation of enzymatic activities aimed at generating metabolic moieties dedicated to respond to the cellular perturbation is explained. This ancient metabolic network has to be recognized for its ability to execute a plethora of physiological functions beyond its well-established traditional roles.

Acute Impacts of Ionizing Radiation Exposure on the Gastrointestinal Tract and Gut Microbiome in Mice.

Radiation therapy for abdominopelvic malignancies often results in damage to the gastrointestinal tract (GIT) and permanent changes in bowel function. An overlooked component of the pathophysiology of radiation-induced bowel injury is the role of the gut microbiome. The goal of this research was to identify the impacts of acute radiation exposure on the GIT and gut microbiome. C57BL/6 mice exposed to whole-body X-rays (0.1-3 Gy) were assessed for histological and microbiome changes 48 h post-radiation exposure. Within the ileum, a dose of 3 Gy significantly decreased crypt depth as well as the number of goblet cells, but increased overall goblet cell size. Overall, radiation altered the microbial distribution within each of the main phyla in a dose- and tissue-dependent manner. Within the Firmicutes phylum, high dose irradiation resulted in significant alterations in bacteria from the class Bacilli within the small bowels, and from the class Clostridia in the large bowels. The 3 Gy radiation also significantly increased the abundance of bacterial families from the Bacteroidetes phylum in the colon and feces. Overall, we identified various alterations in microbiome composition following acute radiation exposure, which could potentially lead to novel biomarkers for tracking patient toxicities or could be used as targets for mitigation strategies against radiation damage.

Metabolic adaptation and ATP homeostasis in Pseudomonas fluorescens exposed to phosphate stress.

Phosphate (Pi) is essential for life as it is an integral part of the universal chemical energy adenosine triphosphate (ATP), and macromolecules such as, DNA, RNA proteins and lipids. Despite the core roles and the need of this nutrient in living cells, some bacteria can grow in environments that are poor in Pi. The metabolic mechanisms that enable bacteria to proliferate in a low phosphate environment are not fully understood. In this study, the soil microbe Pseudomonas (P.) fluorescens was cultured in a control and a low Pi (stress) medium in order to delineate how energy homeostasis is maintained. Although there was no significant variation in biomass yield in these cultures, metabolites like isocitrate, oxaloacetate, pyruvate and phosphoenolpyruvate (PEP) were markedly increased in the phosphate-starved condition. Components of the glycolytic, glyoxylate and tricarboxylic acid cycles operated in tandem to generate ATP by substrate level phosphorylation (SLP) as NADH-producing enzymes were impeded. The α-ketoglutarate (KG) produced when glutamine, the sole carbon nutrient was transformed into phosphoenol pyruvate (PEP) and succinyl-CoA (SC), two high energy moieties. The metabolic reprogramming orchestrated by isocitrate lyase (ICL), phosphoenolpyruvate synthase (PEPS), pyruvate phosphate dikinase (PPDK), and succinyl-CoA synthetase fulfilled the ATP budget. Cell free extract experiments confirmed ATP synthesis in the presence of such substrates as PEP, oxaloacetate and isocitrate respectively. Gene expression profiling revealed elevated transcripts associated with numerous enzymes including ICL, PEPS, and succinyl-CoA synthetase (SCS). This microbial adaptation will be critical in promoting biological activity in Pi-poor ecosystems.

Metabolic adaptation and NADPH homeostasis evoked by a sulfur-deficient environment in Pseudomonas fluorescens.

Sulfur is essential for all living organisms due to its ability to mediate a variety of enzymatic reactions, signalling networks, and redox processes. The interplay between sulfhydryl group (SH) and disulfide bond (S-S) is central to the maintenance of intracellular oxidative balance. Although most aerobic organisms succumb to sulfur starvation, the nutritionally versatile soil microbe Pseudomonas fluorescens elaborates an intricate metabolic reprogramming in order to adapt to this challenge. When cultured in a sulfur-deficient medium with glutamine as the sole carbon and nitrogen source, the microbe reconfigures its metabolism aimed at the enhanced synthesis of NADPH, an antioxidant and the limited production of NADH, a pro-oxidant. While oxidative phosphorylation (OXPHOS) and tricarboxylic acid (TCA) cycle, metabolic modules known to generate reactive oxygen species are impeded, the activities NADPH-producing enzymes such as malic enzyme, and glutamate dehydrogenase (GDH) NADP-dependent are increased. The α-ketoglutarate (KG) generated from glutamine rapidly enters the TCA cycle via α-ketoglutarate dehydrogenase (KGDH), an enzyme that was prominent in the control cultures. In the S-deficient media, the severely impeded KGDH coupled with the increased activity of the reversible isocitrate dehydrogenase (ICDH) that fixes KG into isocitrate in the presence of NADH and HCO3- ensures a constant supply of this critical tricarboxylic acid. The up-regulation of ICDH-NADP dependent in the soluble fraction of the cells obtained from the S-deficient media results in enhanced NADPH synthesis, a reaction aided by the concomitant increase in NAD kinase activity. The latter converts NAD into NADP in the presence of ATP. Taken together, the data point to a metabolic network involving isocitrate, α-KG, and ICDH that converts NADH into NADPH in P. fluorescens subjected to a S-deprived environment.

Reactive nitrogen species (RNS)-resistant microbes: adaptation and medical implications.

Nitrosative stress results from an increase in reactive nitrogen species (RNS) within the cell. Though the RNS - nitric oxide (·NO) and peroxynitrite (ONOO-) - play pivotal physiological roles, at elevated concentrations, these moieties can be poisonous to both prokaryotic and eukaryotic cells alike due to their capacity to disrupt a variety of essential biological processes. Numerous microbes are known to adapt to nitrosative stress by elaborating intricate strategies aimed at neutralizing RNS. In this review, we will discuss both the enzymatic systems dedicated to the elimination of RNS as well as the metabolic networks that are tailored to generate RNS-detoxifying metabolites - α-keto-acids. The latter has been demonstrated to nullify RNS via non-enzymatic decarboxylation resulting in the production of a carboxylic acid, many of which are potent signaling molecules. Furthermore, as aerobic energy production is severely impeded during nitrosative stress, alternative ATP-generating modules will be explored. To that end, a holistic understanding of the molecular adaptation to nitrosative stress, reinforces the notion that neutralization of toxicants necessitates significant metabolic reconfiguration to facilitate cell survival. As the alarming rise in antimicrobial resistant pathogens continues unabated, this review will also discuss the potential for developing therapies that target the alternative ATP-generating machinery of bacteria.

The role of glutamine synthetase in energy production and glutamine metabolism during oxidative stress.

Oxidative stress is known to severely impede aerobic adenosine triphosphate (ATP) synthesis. However, the metabolically-versatile Pseudomonas fluorescens survives this challenge by invoking alternative ATP-generating networks. When grown in a medium with glutamine as the sole organic nutrient in the presence of H2O2, the microbe utilizes glutamine synthetase (GS) to modulate its energy budget. The activity of this enzyme that mediates the release of energy stored in glutamine was sharply increased in the stressed cells compared to the controls. The enhanced activities of such enzymes as acetate kinase, adenylate kinase and nucleotide diphosphate kinase ensured the efficacy of this ATP producing-machine by transferring the high energy phosphate. The elevated amounts of phosphoenol pyruvate carboxylase and pyruvate orthophosphate dikinase recorded in the H2O2 exposed cells provided another route to ATP independent of the reduction of O2. This is the first demonstration of a metabolic pathway involving GS dedicated to ATP synthesis. The phospho-transfer network that is pivotal to the survival of the microorganism under oxidative stress may reveal therapeutic targets against infectious microbes reliant on glutamine for their proliferation.

A novel ATP-generating machinery to counter nitrosative stress is mediated by substrate-level phosphorylation.

BACKGROUND: It is well-known that elevated amounts of nitric oxide and other reactive nitrogen species (RNS) impact negatively on the tricarboxylic acid (TCA) cycle and oxidative phosphorylation. These perturbations severely compromise O2-dependent energy production. While bacteria are known to adapt to RNS, a key tool employed by macrophages to combat infections, the exact mechanisms are unknown. METHODS: The bacterium was cultured in a defined mineral medium and cell-free extracts obtained at the same growth phase were utilized for various biochemical studies Blue native polyacrylamide gel electrophoresis followed by in-gel activity assays, high performance liquid chromatography and co-immunoprecipitaton are applied to investigate the effects of RNS on the model microbe Pseudomonas fluorescens. RESULTS: Citrate is channeled away from the tricarboxylic acid cycle using a novel metabolon consisting of citrate lyase (CL), phosphoenolpyruvate carboxylase (PEPC) and pyruvate phosphate dikinase (PPDK). This metabolic engine comprising three disparate enzymes appears to transiently assemble as a supercomplex aimed at ATP synthesis. The up-regulation in the activities of adenylate kinase (AK) and nucleoside diphosphate kinase (NDPK) ensured the efficacy of this ATP-making machine. CONCLUSION: Microbes may escape the effects of nitrosative stress by re-engineering metabolic networks in order to generate and store ATP anaerobically when the electron transport chain is defective. GENERAL SIGNIFICANCE: The molecular configuration described herein provides further understanding of how metabolism plays a key role in the adaptation to nitrosative stress and reveals novel targets that will inform the development of antimicrobial agents to counter RNS-resistant pathogens.

The role of formate in combatting oxidative stress.

The interaction of keto-acids with reactive oxygen species (ROS) is known to produce the corresponding carboxylic acid with the concomitant formation of CO2. Formate is liberated when the keto-acid glyoxylate neutralizes ROS. Here we report on how formate is involved in combating oxidative stress in the nutritionally-versatile Pseudomonas fluorescens. When the microbe was subjected to hydrogen peroxide (H2O2), the levels of formate were 8 and two-fold higher in the spent fluid and the soluble cell-free extracts obtained in the stressed cultures compared to the controls respectively. Formate was subsequently utilized as a reducing force to generate NADPH and succinate. The former is mediated by formate dehydrogenase (FDH-NADP), whose activity was enhanced in the stressed cells. Fumarate reductase that catalyzes the conversion of fumarate into succinate was also markedly increased in the stressed cells. These enzymes were modulated by H2O2. While the stressed whole cells produced copious amounts of formate in the presence of glycine, the cell-free extracts synthesized ATP and succinate from formate. Although the exact role of formate in anti-oxidative defence has to await further investigation, the data in this report suggest that this carboxylic acid may be a potent reductive force against oxidative stress.

Nuclear lactate dehydrogenase modulates histone modification in human hepatocytes.

It is becoming increasingly apparent that the nucleus harbors metabolic enzymes that affect genetic transforming events. Here, we describe a nuclear isoform of lactate dehydrogenase (nLDH) and its ability to orchestrate histone deacetylation by controlling the availability of nicotinamide adenine dinucleotide (NAD(+)), a key ingredient of the sirtuin-1 (SIRT1) deacetylase system. There was an increase in the expression of nLDH concomitant with the presence of hydrogen peroxide (H2O2) in the culture medium. Under oxidative stress, the NAD(+) generated by nLDH resulted in the enhanced deacetylation of histones compared to the control hepatocytes despite no discernable change in the levels of SIRT1. There appeared to be an intimate association between nLDH and SIRT1 as these two enzymes co-immunoprecipitated. The ability of nLDH to regulate epigenetic modifications by manipulating NAD(+) reveals an intricate link between metabolism and the processing of genetic information.

Mitochondrial lactate metabolism is involved in antioxidative defense in human astrocytoma cells.

Although lactate has traditionally been known to be an end product of anaerobic metabolism, recent studies have revealed its disparate biological functions. Oxidative energy production and cell signaling are two important roles assigned to this monocarboxylic acid. Here we demonstrate that mitochondrial lactate metabolism to pyruvate mediated by lactate dehydrogenase (LDH) in a human astrocytic cell line is involved in antioxidative defense. The pooling of this α-ketoacid helps to detoxify reactive oxygen species, with the concomitant formation of acetate. In-gel activity assays following blue native PAGE electrophoresis were utilized to demonstrate the increase in mitochondrial LDH activity coupled to the decrease in pyruvate dehydrogenase activity in the cells challenged by oxidative stress. The enhanced production of pyruvate with the concomitant formation of acetate in astrocytoma cells was monitored by high-performance liquid chromatography. The ability of pyruvate to fend off oxidative stress was visualized by fluorescence microscopy with the aid of the dye 2',7'-dichlorodihydrofluorescein diacetate. Immunoblotting helped confirm the presence of elevated levels of LDH in cells exposed to oxidative stress, and recovery experiments were performed with pyruvate to diminish the oxidative burden on the astrocytoma. The acetate, generated as a consequence of the antioxidative attribute of pyruvate, was subsequently channeled toward the production of lipids, a process facilitated by the upregulation in activity of acetyl-CoA synthetase and acetyl-CoA carboxylase, as demonstrated by in-gel activity assays. The mitochondrial lactate metabolism mediated by LDH appears to play an important role in antioxidative defence in this astrocytic system.

How aluminum, an intracellular ROS generator promotes hepatic and neurological diseases: the metabolic tale.

Metal pollutants are a global health risk due to their ability to contribute to a variety of diseases. Aluminum (Al), a ubiquitous environmental contaminant is implicated in anemia, osteomalacia, hepatic disorder, and neurological disorder. In this review, we outline how this intracellular generator of reactive oxygen species (ROS) triggers a metabolic shift towards lipogenesis in astrocytes and hepatocytes. This Al-evoked phenomenon is coupled to diminished mitochondrial activity, anerobiosis, and the channeling of α-ketoacids towards anti-oxidant defense. The resulting metabolic reconfiguration leads to fat accumulation and a reduction in ATP synthesis, characteristics that are common to numerous medical disorders. Hence, the ability of Al toxicity to create an oxidative environment promotes dysfunctional metabolic processes in astrocytes and hepatocytes. These molecular events triggered by Al-induced ROS production are the potential mediators of brain and liver disorders.

The unravelling of metabolic dysfunctions linked to metal-associated diseases by blue native polyacrylamide gel electrophoresis.

Gel electrophoresis is routinely used to separate and analyse macromolecules in biological systems. Although many of these electrophoretic techniques necessitate the denaturing of the analytes prior to their analysis, blue native polyacrylamide gel electrophoresis (BN-PAGE) permits the investigation of proteins/enzymes and their supramolecular structures such as the metabolon in native form. This attribute renders this analytical tool conducive to deciphering the metabolic perturbations invoked by metal toxicity. In this review, we elaborate on how BN-PAGE has led to the discovery of the dysfunctional metabolic pathways associated with disorders such as Alzheimer's disease, Parkinson's disease, and obesity that have been observed as a consequence of exposure to various metal toxicants.

The Effects of Oral Probiotics on Type 2 Diabetes Mellitus (T2DM): A Clinical Trial Systematic Literature Review.

Type 2 diabetes mellitus (T2DM) remains a global health concern. Emerging clinical trial (CT) evidence suggests that probiotic intervention may promote a healthy gut microbiome in individuals with T2DM, thereby improving management of the disease. This systematic literature review summarizes thirty-three CTs investigating the use of oral probiotics for the management of T2DM. Here, twenty-one studies (64%) demonstrated an improvement in at least one glycemic parameter, while fifteen studies (45%) showed an improvement in at least one lipid parameter. However, no article in this review was able to establish a uniform decrease in glycemic, lipid, or blood pressure profiles. The lack of consistency across the studies may be attributed to differences in probiotic composition, duration of probiotic consumption, and probiotic dose. An interesting finding of this literature review was the beneficial trend of metformin and probiotic co-administration. Here, patients with T2DM taking metformin demonstrated enhanced glycemic control via the co-administration of probiotics. Taken together, the overall positive findings reported across the studies in combination with minimal adverse effects constitute ground for further quality CTs. This review provides recommendations for future CTs that may address the shortcomings of the current studies and help to extract useful data from future investigations of the use of probiotics in T2DM management.

Molecular Mechanisms Associated with Antifungal Resistance in Pathogenic Candida Species.

Candidiasis is a highly pervasive infection posing major health risks, especially for immunocompromised populations. Pathogenic Candida species have evolved intrinsic and acquired resistance to a variety of antifungal medications. The primary goal of this literature review is to summarize the molecular mechanisms associated with antifungal resistance in Candida species. Resistance can be conferred via gain-of-function mutations in target pathway genes or their transcriptional regulators. Therefore, an overview of the known gene mutations is presented for the following antifungals: azoles (fluconazole, voriconazole, posaconazole and itraconazole), echinocandins (caspofungin, anidulafungin and micafungin), polyenes (amphotericin B and nystatin) and 5-fluorocytosine (5-FC). The following mutation hot spots were identified: (1) ergosterol biosynthesis pathway mutations (ERG11 and UPC2), resulting in azole resistance; (2) overexpression of the efflux pumps, promoting azole resistance (transcription factor genes: tac1 and mrr1; transporter genes: CDR1, CDR2, MDR1, PDR16 and SNQ2); (3) cell wall biosynthesis mutations (FKS1, FKS2 and PDR1), conferring resistance to echinocandins; (4) mutations of nucleic acid synthesis/repair genes (FCY1, FCY2 and FUR1), resulting in 5-FC resistance; and (5) biofilm production, promoting general antifungal resistance. This review also provides a summary of standardized inhibitory breakpoints obtained from international guidelines for prominent Candida species. Notably, N. glabrata, P. kudriavzevii and C. auris demonstrate fluconazole resistance.

Tellurite-exposed Escherichia coli exhibits increased intracellular α-ketoglutarate.

The tellurium oxyanion tellurite is toxic to most organisms because of its ability to generate oxidative stress. However, the detailed mechanism(s) how this toxicant interferes with cellular processes have yet to be fully understood. As part of our effort to decipher the molecular interactions of tellurite with living systems, we have evaluated the global metabolism of α-ketoglutarate a known antioxidant in Escherichia coli. Tellurite-exposed cells displayed reduced activity of the KG dehydrogenase complex (KGDHc), resulting in increased intracellular KG content. This complex's reduced activity seems to be due to decreased transcription in the stressed cells of sucA, a gene that encodes the E1 component of KGDHc. Furthermore, it was demonstrated that the increase in total reactive oxygen species and superoxide observed upon tellurite exposure was more evident in wild type cells than in E. coli with impaired KGDHc activity. These results indicate that KG may be playing a pivotal role in combating tellurite-mediated oxidative damage.

A facile electrophoretic technique to monitor phosphoenolpyruvate-dependent kinases.

Phosphoenolpyruvate (PEP)-dependent kinases are central to numerous metabolic processes and mediate the production of adenosine triphosphate (ATP) by substrate-level phosphorylation (SLP). While pyruvate kinase (PK, EC: 2.7.1.40), the final enzyme of the glycolytic pathway is critical in the anaerobic synthesis of ATP from ADP, pyruvate phosphate dikinase (PPDK, EC: 2.7.9.1), and phosphoenolpyruvate synthase (PEPS, EC: 2.7.9.2) help generate ATP from AMP coupled to PEP as a substrate. Here we demonstrate an inexpensive and effective electrophoretic technology to determine the activities of these enzymes by blue-native polyacrylamide gel electrophoresis (BN-PAGE). The generation of pyruvate is linked to exogenous lactate dehydrogenase (LDH), and the oxidation of reduced nicotinamide adenine dinucleotide (NADH) coupled to 2,6-dichloroindophenol (DCIP) and iodonitrotetrazolium chloride (INT) results in a formazan precipitate which is easily quantifiable. The selectivity of the enzymes is ensured by including either AMP or ADP and pyrophosphate (PP(i) ) or inorganic phosphate (P(i) ). Activity bands were readily obtained after incubation in the respective reaction mixtures for 20-30 min. Cell-free extract concentrations as low as 20 µg protein equivalent yielded activity bands and substrate levels were manipulated to optimize sensitivity of this analytical technique. High-pressure liquid chromatography (HPLC), two-dimensional (2-D) SDS-PAGE (where SDS is sodium dodecyl sulfate), and immunoblot studies of the excised activity band help further characterize these PEP-dependent kinases. Furthermore, these enzymes were readily identified on the same gel by incubating it sequentially in the respective reaction mixtures. This technique provides a facile method to elucidate these kinases in biological systems.

Hepatic response to aluminum toxicity: dyslipidemia and liver diseases.

Aluminum (Al) is a metal toxin that has been implicated in the etiology of a number of diseases including Alzheimer's, Parkinson's, dialysis encephalopathy, and osteomalacia. Al has been shown to exert its effects by disrupting lipid membrane fluidity, perturbing iron (Fe), magnesium, and calcium homeostasis, and causing oxidative stress. However, the exact molecular targets of aluminum's toxicity have remained elusive. In the present review, we describe how the use of a systems biology approach in cultured hepatoblastoma cells (HepG2) allowed the identification of the molecular targets of Al toxicity. Mitochondrial metabolism is the main site of the toxicological action of Al. Fe-dependent and redox sensitive enzymes in the tricarboxylic acid (TCA) cycle and oxidative phosphorylation (OXPHOS) are dramatically decreased by Al exposure. In an effort to compensate for diminished mitochondrial function, Al-treated cells stabilize hypoxia inducible factor-1α (HIF-1α) to increase ATP production by glycolysis. Additionally, Al toxicity leads to an increase in intracellular lipid accumulation due to enhanced lipogenesis and a decrease in the ß-oxidation of fatty acids. Central to these effects is the alteration of α-ketoglutarate (KG) homeostasis. In Al-exposed cells, KG is preferentially used to quench ROS leading to succinate accumulation and HIF-1α stabilization. Moreover, the channeling of KG to combat oxidative stress leads to a reduction of l-carnitine biosynthesis and a concomitant decrease in fatty acid oxidation. The fluidity and interaction of these metabolic modules and the implications of these findings in liver-related disorders are discussed herein.

A Metabolic Network Mediating the Cycling of Succinate, a Product of ROS Detoxification into α-Ketoglutarate, an Antioxidant.

Sulfur is an essential element for life. However, the soil microbe Pseudomonas (P.) fluorescens can survive in a low sulfur environment. When cultured in a sulfur-deficient medium, the bacterium reprograms its metabolic pathways to produce α-ketoglutarate (KG) and regenerate this keto-acid from succinate, a by-product of ROS detoxification. Succinate semialdehyde dehydrogenase (SSADH) and KG decarboxylase (KGDC) work in partnership to synthesize KG. This process is further aided by the increased activity of the enzymes glutamate decarboxylase (GDC) and γ-amino-butyrate transaminase (GABAT). The pool of succinate semialdehyde (SSA) generated is further channeled towards the formation of the antioxidant. Spectrophotometric analyses, HPLC experiments and electrophoretic studies with intact cells and cell-free extracts (CFE) pointed to the metabolites (succinate, SSA, GABA) and enzymes (SSADH, GDC, KGDC) contributing to this KG-forming metabolic machinery. Real-time polymerase chain reaction (RT-qPCR) revealed significant increase in transcripts of such enzymes as SSADH, GDC and KGDC. The findings of this study highlight a novel pathway involving keto-acids in ROS scavenging. The cycling of succinate into KG provides an efficient means of combatting an oxidative environment. Considering the central role of KG in biological processes, this metabolic network may be operative in other living systems.

Metabolic networks to combat oxidative stress in Pseudomonas fluorescens.

Oxidative stress is an unavoidable peril that aerobic organisms have to confront. Thus, it is not surprising that intricate strategies are deployed in an effort to fend the dangers associated with living in an O(2) environment. In the classical models of anti-oxidative defense mechanisms, a variety of stratagems including the reactive oxygen species (ROS) scavenging systems, the NADPH-generating enzymes and the DNA repair machineries are highlighted. However, it is becoming increasingly clear that metabolism may be intimately involved in anti-oxidative defence. Recent data show that metabolic reprogramming plays a pivotal role in the survival of organisms exposed to oxidative stress. Here, we describe how Pseudomonas fluorescens, the metabolically-versatile soil microbe, manipulates its metabolic networks in an effort to counter oxidative stress. An intricate link between metabolism and anti-oxidative defense is presented. P. fluorescens reconfigures its metabolic processes in an effort to satisfy its need for NADPH during oxidative insult. Seemingly, disparate metabolic modules appear to partner together to concomitantly fine-tune the levels of the anti-oxidant NADPH and the pro-oxidant NADH. Central to this shift in the metabolic production of the pyridine nucleotides is the increase in NAD kinase with the concomitant decrease in NADP phosphatase. The tricarboxylic acid cycle is tweaked in an effort to limit the formation of NADH. This metabolic redox-balancing act appears to afford a potent tool against oxidative challenge and may be a more widespread ROS-combating tactic than hitherto recognized.

Beyond mitochondria: Alternative energy-producing pathways from all strata of life.

It is well-established that mitochondria are the powerhouses of the cell, producing adenosine triphosphate (ATP), the universal energy currency. However, the most significant strengths of the electron transport chain (ETC), its intricacy and efficiency, are also its greatest downfalls. A reliance on metal complexes (FeS clusters, hemes), lipid moities such as cardiolipin, and cofactors including alpha-lipoic acid and quinones render oxidative phosphorylation vulnerable to environmental toxins, intracellular reactive oxygen species (ROS) and fluctuations in diet. To that effect, it is of interest to note that temporal disruptions in ETC activity in most organisms are rarely fatal, and often a redundant number of failsafes are in place to permit continued ATP production when needed. Here, we highlight the metabolic reconfigurations discovered in organisms ranging from parasitic Entamoeba to bacteria such as pseudomonads and then complex eukaryotic systems that allow these species to adapt to and occasionally thrive in harsh environments. The overarching aim of this review is to demonstrate the plasticity of metabolic networks and recognize that in times of duress, life finds a way.