Electron Transport Phosphorylation Research Articles

Unraveling response mechanism of electron transport mechanism to uranium-exposure in Bacillus sp. X02

With the development of the uranium mining industry, the remediation of uranium pollution is an urgent environmental problem to be solved. Bioreduction is currently recognized as a U(VI) bioremediation method with good application prospects. However, inadequate reporting of uranium-reducing bacteria and low bioreduction efficiency are the main reasons limiting the application of the bioreduction method. In this paper, we isolated an efficient uranium-reducing bacterium, Bacillus sp. X02, explored the effect of U(VI) on bioreduction efficiency through electron transport mechanisms. The results showed that the maximum removal and reduction rate were 98.35 % and 62.43 %, respectively. Further studies revealed that the efficiency of extracellular electron transport of Bacillus sp. X02 was enhanced under U(VI) exposure. But uranium stress down-regulated the expression of F-type ATPase, Cytochrome c reductase ISP, Cytochrome c oxidase, and Cytochrome bd complex, inhibited the processes of complex III, complex IV, and ATP synthesis in the oxidative phosphorylation electron transport pathway, decreasing the electron transfer and U(VI) reduction capacity of Bacillus sp. X02 ultimately. This work proposed a new electron transfer pathway for U(VI) bioreduction and provided new insights into the effect of uranium on bioreduction efficiency.

The Mysterious Multitude: Structural Perspective on the Accessory Subunits of Respiratory Complex I.

Complex I (CI) is the largest protein complex in the mitochondrial oxidative phosphorylation electron transport chain of the inner mitochondrial membrane and plays a key role in the transport of electrons from reduced substrates to molecular oxygen. CI is composed of 14 core subunits that are conserved across species and an increasing number of accessory subunits from bacteria to mammals. The fact that adding accessory subunits incurs costs of protein production and import suggests that these subunits play important physiological roles. Accordingly, knockout studies have demonstrated that accessory subunits are essential for CI assembly and function. Furthermore, clinical studies have shown that amino acid substitutions in accessory subunits lead to several debilitating and fatal CI deficiencies. Nevertheless, the specific roles of CI’s accessory subunits have remained mysterious. In this review, we explore the possible roles of each of mammalian CI’s 31 accessory subunits by integrating recent high-resolution CI structures with knockout, assembly, and clinical studies. Thus, we develop a framework of experimentally testable hypotheses for the function of the accessory subunits. We believe that this framework will provide inroads towards the complete understanding of mitochondrial CI physiology and help to develop strategies for the treatment of CI deficiencies.

Mitochondrial activities play a pivotal role in regulating cell cycle in response to doxorubicin

ABSTRACT Doxorubicin induces both DNA damage and metabolic interference. How these effects interact to modulate cellular toxicity is not completely understood but important given the widespread use of doxorubicin in cancer treatment. This study tests the hypothesis that cell cycle arrest and survival are affected by distinct mitochondrial activities during doxorubicin exposure. Parental and mutant S. cerevisiae strains deficient in selected genes with mitochondrial function were treated with doxorubicin and assayed for changes in proliferation rates, cell survival and cell cycle arrest kinetics. Mitochondrial DNA content was estimated using quantitative PCR. Mitochondrial function was assessed by measuring oxygen consumption with and without an uncoupler. Parental cells growing in a non-fermentable carbon source medium and mutants lacking mitochondria and grown in glucose medium both show abrupt cell cycle and proliferation arrest during doxorubicin exposure compared to parental cells grown in glucose. Mitochondrial DNA increases during doxorubicin exposure in S. cerevisiae and in human breast cancer cells. Yeast strains deficient in TCA cycle activity or electron transport both show more abrupt cell cycle arrest than parental cells when exposed to doxorubicin. Concurrent treatment with the mitochondrial uncoupler dinitrophenol facilitates cell cycle progression and proliferation during doxorubicin exposure. Doxorubicin exposure induces mitochondrial DNA synthesis with TCA cycle and oxidative phosphorylation activity having opposing effects on cell proliferation, survival and cell cycle kinetics. TCA cycle activity provides biosynthetic substrates to support cell cycle progression and cell proliferation while electron transport and oxidative phosphorylation facilitate cell cycle arrest and possibly increased cytotoxicity.

An energetic profile of Corynebacterium glutamicum underpinned by measured biomass yield on ATP

The supply and usage of energetic cofactors in metabolism is a central concern for systems metabolic engineering, particularly in case of energy intensive products. One of the most important parameters for systems wide balancing of energetic cofactors is the ATP requirement for biomass formation YATP/Biomass. Despite its fundamental importance, YATP/Biomass values for non-fermentative organisms are still rough estimates deduced from theoretical considerations. For the first time, we present an approach for the experimental determination of YATP/Biomass using comparative 13C metabolic flux analysis (13C MFA) of a wild type strain and an ATP synthase knockout mutant. We show that the energetic profile of a cell can then be deduced from a genome wide stoichiometric model and experimental maintenance data. Particularly, the contributions of substrate level phosphorylation (SLP) and electron transport phosphorylation (ETP) to ATP generation become available which enables the overall energetic efficiency of a cell to be characterized. As a model organism, the industrial platform organism Corynebacterium glutamicum is used. C. glutamicum uses a respiratory type of energy metabolism, implying that ATP can be synthesized either by SLP or by ETP with the membrane-bound F1FO-ATP synthase using the proton motive force (pmf) as driving force. The presence of two terminal oxidases, which differ in their proton translocation efficiency by a factor of three, further complicates energy balancing for this organism. By integration of experimental data and network models, we show that in the wild type SLP and ETP contribute equally to ATP generation. Thus, the role of ETP in respiring bacteria may have been overrated in the past. Remarkably, in the genome wide setting 65% of the pmf is actually not used for ATP synthesis. However, it turns out that, compared to other organisms C. glutamicum still uses its energy budget rather efficiently.

Overcoming Energetic Barriers in Acetogenic C1 Conversion.

Currently one of the biggest challenges for society is to combat global warming. A solution to this global threat is the implementation of a CO2-based bioeconomy and a H2-based bioenergy economy. Anaerobic lithotrophic bacteria such as the acetogenic bacteria are key players in the global carbon and H2 cycle and thus prime candidates as driving forces in a H2- and CO2-bioeconomy. Naturally, they convert two molecules of CO2 via the Wood-Ljungdahl pathway (WLP) to one molecule of acetyl-CoA which can be converted to different C2-products (acetate or ethanol) or elongated to C4 (butyrate) or C5-products (caproate). Since there is no net ATP generation from acetate formation, an electron-transport phosphorylation (ETP) module is hooked up to the WLP. ETP provides the cell with additional ATP, but the ATP gain is very low, only a fraction of an ATP per mol of acetate. Since acetogens live at the thermodynamic edge of life, metabolic engineering to obtain high-value products is currently limited by the low energy status of the cells that allows for the production of only a few compounds with rather low specificity. To set the stage for acetogens as production platforms for a wide range of bioproducts from CO2, the energetic barriers have to be overcome. This review summarizes the pathway, the energetics of the pathway and describes ways to overcome energetic barriers in acetogenic C1 conversion.

Abstract IA31: Modeling and targeting double-hit lymphoma

Abstract High-grade B-cell lymphoma with concurrent activation of MYC and BCL2, also known as double-hit lymphoma (DHL), shows dismal prognosis with current front-line therapies (e.g., R-CHOP), calling for the development of new therapeutic regimens. The synergy between MYC and BCL2 in lymphomagenesis is explained by the ability of BCL2 to block the proapoptotic activity of MYC, while leaving intact its proliferative potential; on this basis, we reasoned that compounds that exacerbate MYC-induced apoptosis might cooperate with BCL2 inhibitors in killing DHL cells. We previously observed that genes encoding components of the mitochondrial ribosome were coordinately activated in MYC-driven lymphoma (1) and were critical for tumor maintenance in transgenic mice (2). In line with the genetic data, inhibition of mitochondrial translation with the antibiotic tigecycline was synthetic-lethal with MYC activation and extended lifespan in lymphoma-bearing mice (2). Combining tigecycline with the BCL2 inhibitor venetoclax revealed synergy in the induction of apoptosis in human DHL cell lines, as well as marked antitumoral activity in xenografted mice (3). These preclinical data warrant the repurposing of venetoclax and tigecycline for the treatment of refractory and/or relapsed DHL. By blocking the synthesis of mitochondrially encoded proteins, tigecycline impairs the assembly of the electron-transport chain (ETC) complexes and oxidative phosphorylation. We thus hypothesized that direct ETC inhibitors might also preferentially kill MYC-overexpressing cells and synergize with venetoclax against DHL. We will present data that verify these predictions for IACS-010759, a small-molecule inhibitor of mitochondrial complex I. A current limitation for further preclinical development and mechanistic analysis lies in the lack of a bona fide mouse model of DHL. While the cooperation between MYC and BCL2 in lymphomagenesis was amply documented over almost three decades, the models described so far led to tumor onset at early stages of B-cell ontogeny; as a consequence, no model is available that reliably reproduces the development of DHL from germinal center (GC) B-cells, as observed in the clinic. In order to overcome this limitation, we combined CRE-activated alleles of MYC and BCL2 with Cgamma1-CRE, a CRE-expressing transgene that is specifically expressed in GC B cells. Concerted activation of both oncogenes led to increased expansion of GC B cells upon immunization, followed within a few months by the onset of aggressive, invasive B-cell lymphomas. Our progress in characterizing this disease model will be presented at the meeting.

The Rnf complex is a Na+ coupled respiratory enzyme in a fermenting bacterium, Thermotoga maritima

rnf genes are widespread in bacteria and biochemical and genetic data are in line with the hypothesis that they encode a membrane-bound enzyme that oxidizes reduced ferredoxin and reduces NAD and vice versa, coupled to ion transport across the cytoplasmic membrane. The Rnf complex is of critical importance in many bacteria for energy conservation but also for reverse electron transport to drive ferredoxin reduction. However, the enzyme has never been purified and thus, ion transport could not be demonstrated yet. Here, we have purified the Rnf complex from the anaerobic, fermenting thermophilic bacterium Thermotoga maritima and show that is a primary Na+ pump. These studies provide the proof that the Rnf complex is indeed an ion (Na+) translocating, respiratory enzyme. Together with a Na+-F1FO ATP synthase it builds a simple, two-limb respiratory chain in T. maritima. The physiological role of electron transport phosphorylation in a fermenting bacterium is discussed.

Abstract 359: Cardiac-specific Deletion of Prohibitin-1 Leads to Fetal Demise, Gross Mitochondrial Abnormalities and Heart Failure in Adolescent Mice

Prohibitins (PHB1, 2) are lipid raft-associated proteins which form a multimeric, alternating ring-like structure in cellular membranes, and are particularly abundant in mitochondria and plasma membranes, but have also been found in nuclei. These pleiotropic proteins are associated with numerous signaling pathways responsible for cell differentiation, growth, and metabolism. In mitochondria, PHBs are thought to act as scaffold-like chaperone proteins responsible for stabilizing enzymes responsible for electron transport and oxidative phosphorylation (OXPHOS), minimizing reactive oxygen species (ROS), and coordinating fission/fusion. However, the extent to which PHB1 regulates mitochondria and electromechanical function of the heart, is not known. We established a cardiac-specific PHB1-deficient mouse strain by crossing floxxed phb1 mice with mice expressing Cre recombinase under control of myosin heavy chain (MHC) promoter. The vast majority of cardiac-specific PHB1-deficient mice (cPHB1ko) died in utero during pregnancy (E12-E15). Uterine sonography of the expired cPHB1ko embryos showed significantly decreased crown-rump length and increased placenta thickness with large amount of fluids, restricted uterine artery flow and detachment between amnion and chorion compared with normal embryos (P < 0.001). The few that were born develop severe dilated cardiomyopathy, resulting in mortality by 8-9 weeks of age. Heart weight/body weight ratio is ~2-fold greater in the KO mice vs. WT. Echocardiography reveals severely enlarged right atria and ventricles in cPHB1ko mice, corresponding to significantly decreased ejection faction (EF), increased end diastolic volume (EDV) and end systolic volume (ESV) compared to wild-type littermates (P < 0.05). In mitochondria, cPHB1ko mice have derangements in OXPHOS as evidenced by decreased pyruvate and palmitoylcarnitine-supported respiration and ATP production. Moreover, both the rate of calcium uptake (P = 0.05) and the total calcium retention capacity (P < 0.01) were diminished in the KO mice. These preliminary findings implicate PHB1 as a critical regulator of myocardial development and function, due in part to its role in maintaining OXPHOS and controlling metabolism.

Mitochondria of transformed cell as a target of antitumor influence

Mitochondria are intracellular organelles in eukaryotic cells that participate in bioenergy metabolism and cell homeostasis, including ATP generation through electron transport and oxidative phosphorylation in combination with oxidation of metabolites by the tricarboxylic acid cycle and fatty acid catabolism via β-oxidation. the production of reactive oxygen species, as well as the initiation and implementation of apoptosis. Mitochondria play a crucial role in cellular energy metabolism and the regulation of programmed cell death. mitochondria activate numerous signaling pathways associated with cell death. Mitochondria have the ability to control the activation of programmed cell death by regulating the translocation of proapoptotic proteins from the intermediate space of mitochondria to the cytosol. This is the reason for the emergence of a new discipline — mitochondrial medicine. The review examined and analyzed scientific publications on the role of mitochondria in the life support of transformed cells, the study of their functioning and structurally functional dysfunctions, as part of mitochondrial medicine. Mitochondrial medicine is a developing discipline whose significance stems from the central function of mitochondria in the production of adenosine triphosphate, the generation of reactive oxygen species, and cell death due to necrosis or apoptosis. Consequently, mitochondrial dysfunction plays an important role in the pathophysiology of cancer, many other common diseases and side effects of drugs. Perhaps the combined use of modulators of mitochondrial metabolism and antitumor therapy will contribute to the emergence of a new direction in antitumor treatment, which will significantly increase the effectiveness of cancer treatment.

Dietary Energy Level Promotes Rumen Microbial Protein Synthesis by Improving the Energy Productivity of the Ruminal Microbiome.

Improving the yield of rumen microbial protein (MCP) has significant importance in the promotion of animal performance and the reduction of protein feed waste. The amount of energy supplied to rumen microorganisms is an important factor affecting the amount of protein nitrogen incorporated into rumen MCP. Substrate-level phosphorylation (SLP) and electron transport phosphorylation (ETP) are two major mechanisms of energy generation within microbial cells. However, the way that energy and protein levels in the diet impact the energy productivity of the ruminal microbiome and, thereafter, rumen MCP yields is not known yet. In present study, we have investigated, by animal experiments and metagenome shotgun sequencing, the effects of energy-rich and protein-rich diets on rumen MCP yields, as well as SLP-coupled and ETP-coupled energy productivity of the ruminal microbiome. We have found that an energy-rich diet induces a significant increase in rumen MCP yield, whereas a protein-rich diet has no significant impacts on it. Based on 10 reconstructed pathways related to the energy metabolism of the ruminal microbiome, we have determined that the energy-rich diet induces significant increases in the total abundance of SLP enzymes coupled to the nicotinamide adenine dinucleotide (NADH) oxidation in the glucose fermentation and F-type ATPase of the electron transporter chain, whereas the protein-rich diet has no significant impact in the abundance of these enzymes. At the species level, the energy-rich diet induces significant increases in the total abundance of 15 ETP-related genera and 40 genera that have SLP-coupled fermentation pathways, whereas the protein-rich diet has no significant impact on the total abundance of these genera. Our results suggest that an increase in dietary energy levels promotes rumen energy productivity and MCP yield by improving levels of ETP and SLP coupled to glucose fermentation in the ruminal microbiome. But, an increase in dietary protein level has no such effects.

Physiology and central carbon metabolism of the gut bacterium Prevotella copri

The human gut microbiota is a crucial factor for the host's physiology with respect to health and disease. Metagenomic shotgun sequencing of microbial gut communities revealed that Prevotella copri is one of the most important players in the gastrointestinal tract of many individuals. Because of the importance of this bacterium we analyzed the growth behavior and the central metabolic pathways of P. copri. Bioinformatic data, transcriptome profiling and enzyme activity measurements indicated that the major pathways are based on glycolysis and succinate production from fumarate. In addition, pyruvate can be degraded to acetate and formate. Electron transport phosphorylation depends on fumarate respiration with NADH and reduced ferredoxin as electron donors. In contrast to Bacteroides vulgatus, P. copri showed a more pronounced dependency on the addition of CO2 or bicarbonate for biomass formation, which is a remarkable difference between P. copri and Bacteroides spp. with important implication in the context of gut microbial competition. The analysis of substrate consumption and product concentrations from many P. copri cultures with different optical densities allowed a prediction of the carbon and electron flow in the central metabolism and a detailed calculation of growth yields as well as carbon and redox balances.

Clarifying the supercomplex: the higher-order organization of the mitochondrial electron transport chain.

The oxidative phosphorylation electron transport chain (OXPHOS-ETC) of the inner mitochondrial membrane is composed of five large protein complexes, named CI-CV. These complexes convert energy from the food we eat into ATP, a small molecule used to power a multitude of essential reactions throughout the cell. OXPHOS-ETC complexes are organized into supercomplexes (SCs) of defined stoichiometry: CI forms a supercomplex with CIII2 and CIV (SC I+III2+IV, known as the respirasome), as well as with CIII2 alone (SC I+III2). CIII2 forms a supercomplex with CIV (SC III2+IV) and CV forms dimers (CV2). Recent cryo-EM studies have revealed the structures of SC I+III2+IV and SC I+III2. Furthermore, recent work has shed light on the assembly and function of the SCs. Here we review and compare these recent studies and discuss how they have advanced our understanding of mitochondrial electron transport.

Abstract 5893: Bioenergetic signature from cocultures of pancreatic tumor cell lines and fibroblasts

Abstract Pancreatic ductal adenocarcinoma (PDAC) is one of the tumors with greater invasiveness and metastasis. PDAC is associated with a large desmoplastic reaction, characterized by fibroblastic proliferation and extracellular matrix secretion. Besides, cellular bioenergetics has become a central issue of investigation in cancer biology, because the altered energy metabolism of cancer cells has been proposed as a potential target for cancer treatment. We have established co-cultures between PDAC cell lines (Capan-1 or PL-45) and fibroblasts (LC5). A reverse phase protein microarray (RPMA) approach has been applied to quantify proteins of energy metabolism in theses co-cultures, with the aim of identifying potential biomarkers in PDACs. The fifteen proteins of energy metabolism studied included members of the mitochondrial oxidation of pyruvate, the tricarboxylic acid cycle, β-oxidation of fatty acids, electron transport and oxidative phosphorylation, glycogen metabolism, glycolysis and oxidative stress using highly specific antibodies. Co-cultures modified proteins expression of energy metabolism, respect to monocultures. Capan-1 in co-culture increased several proteins belonging to OXPHOS (NDUFS3, SDHB, CORE2 and COXII), the antioxidant SOD2, the mitochondrial HSP60, and G6PDH. The expression of the glycolitic PKM2 decreased. Their fibroblasts partners increased the expression of SDHB and also decreased PKM2. PL-45 in co-culture, experimented an increase in HADHA. Their fibroblasts partners showed a decrease in PKM2, LDHA, IF1, PDHe and HSP60.Capann-1 and PL-45 in co-culture with fibroblast, differ en their energy metabolism phenotype. Overall, the results indicate that the quantification of bioenergetic signature offers potential biomarkers that could be implemented to refine the understanding of the biological principles of PDAC and, eventually, the management of patients with these tumors. Citation Format: Mª Teresa Agulló-Ortuño, Elena Prieto-García, C. Vanesa Díaz-García, Irene Otero Blas, Inmaculada García-Ruíz, José A. López-Martín. Bioenergetic signature from cocultures of pancreatic tumor cell lines and fibroblasts [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5893. doi:10.1158/1538-7445.AM2017-5893

Effect of phytotoxic secondary metabolites and semisynthetic compounds from endophytic fungus Xylaria feejeensis strain SM3e-1b on spinach chloroplast photosynthesis

We investigated the mechanism of action on the photosynthesis light reactions of three major secondary metabolites produced by the endophytic fungus Xylaria feejeensis strain SM3e-1b, isolated from Sapium macrocarpum; and four novel derivatives of coriloxine, a major compound produced by X. feejeensis. The natural phytotoxins include one epoxycyclohexenone derivative, coriloxine (1), and two quinone derivatives (2–3). The semisynthetic derivatives of coriloxine are two cyclohexenone (4–6) and two quinone compounds (5–7). Cyclohexenone (4), (4R,5S,6R)-6-chloro-4,5-dihydroxy-3-methoxy-5-methylcyclohex-2-enone, inhibited ATP synthesis in freshly lysed spinach chloroplasts from water to MV; it also partly inhibited the basal and uncoupled photosynthetic electron transport, and significantly enhanced the phosphorylating electron transport and Mg2+-ATPase activity, thus demonstrating its action as an uncoupler agent. On the other hand, quinone (7), 2-((4-butylphenyl)amino)-5-methoxy-3-methylcyclohexa-2,5-diene-1,4-dione, inhibited ATP synthesis, and non-cyclic electron transport from water to MV in basal, phosphorylating and uncoupled conditions in a concentration-dependent manner. Hence, (7) behaves as a Hill reaction inhibitor at the PSII electron transport on the water splitting enzyme (OEC), and on the acceptor side between P680 and QA. This mechanism of action was confirmed by chlorophyll a fluorescence measurements. These results indicate that coriloxine derivatives 4 and 7 could work as prototype structures for the development of new herbicides. Contrastingly, natural products 1–3, and derivatives 5 and 6 did not show a significant inhibitory effect on ATP synthesis.

Organohalid-Atmung bei Mikroorganismen

Reductive dehalogenation is coupled to energy conservation via electron transport phosphorylation in organohalide-respiring bacteria. The electron transfer to the terminal reductase, the corrinoid-containing reductive dehalogenase, leads to the formation of a proton gradient across the membrane. The architecture of the electron transfer chains differs between different phyla. Organohalide respiration is an important part of the global halogen cycle and of high relevance in bioremediation.

Bacterial Electron Transfer Chains Primed by Proteomics.

Electron transport phosphorylation is the central mechanism for most prokaryotic species to harvest energy released in the respiration of their substrates as ATP. Microorganisms have evolved incredible variations on this principle, most of these we perhaps do not know, considering that only a fraction of the microbial richness is known. Besides these variations, microbial species may show substantial versatility in using respiratory systems. In connection herewith, regulatory mechanisms control the expression of these respiratory enzyme systems and their assembly at the translational and posttranslational levels, to optimally accommodate changes in the supply of their energy substrates. Here, we present an overview of methods and techniques from the field of proteomics to explore bacterial electron transfer chains and their regulation at levels ranging from the whole organism down to the Ångstrom scales of protein structures. From the survey of the literature on this subject, it is concluded that proteomics, indeed, has substantially contributed to our comprehending of bacterial respiratory mechanisms, often in elegant combinations with genetic and biochemical approaches. However, we also note that advanced proteomics offers a wealth of opportunities, which have not been exploited at all, or at best underexploited in hypothesis-driving and hypothesis-driven research on bacterial bioenergetics. Examples obtained from the related area of mitochondrial oxidative phosphorylation research, where the application of advanced proteomics is more common, may illustrate these opportunities.

Abstract B25: Novel hepatic phenotypes caused by conditional c-Myc deletion

Abstract The transcription factor c-Myc (hereafter Myc) is among the most frequently deregulated oncoproteins. Inhibition of Myc triggers proliferative arrest of transformed cells and promotes tumor regression and/or apoptosis. Myc is also developmentally necessary and myc-/- embryos die at E9.5-10.5. However, Myc's role in the maintenance of specific tissues has been shown to be of variable importance and necessity. For example, several studies of Myc's role in promoting liver regeneration following partial hepatectomy (PH) have given conflicting results, although all show Myc to be generally dispensable for this function. We have used a conditional murine knockout (KO) model of Myc to study its role in liver regeneration. By employing a mycfl/fl;Alb-Cre+ model in which loss of Myc occurs perinatally, we studied non-oncogenic liver proliferation and metabolism in the absence of Myc signaling. We employed basic metabolic benchmarks of liver function including measurements of triglyceride levels, oxidative phosphorylation, and TCA cycle and electron transport chain function. At the molecular level, RNAseq was performed on isolated hepatocytes and the mitochondrial proteome was evaluated by both differential and unbiased mass spectrometry. At the time of active Myc excision, myc-/- mice had a significantly lower liver: body weight ratios relative to myc+/+ controls. However, this was reversed in older mice and was associated with the hepatic accumulation of neutral lipids, cholesterol and increased fatty acid β-oxidation in myc-/- mice. RNAseq on hepatocytes and Ingenuity Pathway analyses showed differences in 105 transcripts (q<0.05), the major pathways encoding ribosomal proteins, members of the cytochrome p450 family and enzymes involved in cholesterol and bile metabolism. These findings correlated with abnormalities in fatty acid and sterol metabolism and storage in liver samples. PH may provide an insufficiently sustained proliferative challenge to allow adequate evaluation of Myc's potential role in liver regeneration. We therefore utilized a mouse model of hereditary tyrosinemia in which knockout of the gene encoding fumarylacetoacetate hydrolase (FAH) leads to hepatocellular death that can be rescued by the infusion of fah+/+ hepatocytes, which expand and eventually replace the fah-/- recipient hepatocytes. FAH-deficient animals could be rescued equally well by both myc+/+ and myc-/- hepatocytes. However, livers from the latter group showed excessive neutral lipid accumulation and fibrosis, reminiscent of non-alcoholic steatohepatitis (NASH). Taken together, our results provide unequivocal evidence that Myc is dispensable for long-term hepatic regeneration but is necessary to maintain proper lipid and steroid metabolism. In Myc's absence the excessive accumulation of these intermediates predisposes to the development of a relatively mild pathology mimicking non-alcoholic fatty liver disease, which under the duress of chronic proliferation, progresses to a more severe NASH-like picture of end-stage liver disease. Our studies thus reveal a heretofore unappreciated role for Myc in hepatic metabolic homeostasis. Citation Format: Lia R. Edmunds, Lokendra Sharma, Peter Anthony Otero, Sonia D'Souza, James M. Dolezal, Xuemei Zeng, Ying Ding, Fei Ding, Megan E. Beck, Lisa E. Kratz, Jerry Vockley, Eric Goetzman, Donald Scott, Nathan Yates, Andrew W. Duncan, Edward V. Prochownik. Novel hepatic phenotypes caused by conditional c-Myc deletion. [abstract]. In: Proceedings of the AACR Special Conference: Metabolism and Cancer; Jun 7-10, 2015; Bellevue, WA. Philadelphia (PA): AACR; Mol Cancer Res 2016;14(1_Suppl):Abstract nr B25.

Identification of a new marine algal species Pyropia nitida sp. nov. (Bangiales: Rhodophyta) from Monterey, California

An unidentified marine red algal species classified in Pyropia J. Agardh was discovered from Monterey, CA. Morphological, barcode, and complete mitochondrial genome analysis of the alga support its recognition as a new species, Pyropia nitida sp. nov. The species is a high-intertidal, winter annual that is lanceolate in shape, monostromatic, and dioecious. Based on CO1 sequences, P. nitida is closely allied with the P. nereocystis clade. The mitogenome of P. nitida is 35 313 bp in length and contains 53 genes, including two ribosomal RNAs, 24 transfer RNAs, four ribosomal proteins, two ymfs, four ORFs, and 17 genes involved in electron transport and oxidative phosphorylation. The results support the recognition of P. nitida as distinct from the morphologically similar P. lanceolata.

Energy Conservation Associated with Ethanol Formation from H2 and CO2 in Clostridium autoethanogenum Involving Electron Bifurcation.

Most acetogens can reduce CO2 with H2 to acetic acid via the Wood-Ljungdahl pathway, in which the ATP required for formate activation is regenerated in the acetate kinase reaction. However, a few acetogens, such as Clostridium autoethanogenum, Clostridium ljungdahlii, and Clostridium ragsdalei, also form large amounts of ethanol from CO2 and H2. How these anaerobes with a growth pH optimum near 5 conserve energy has remained elusive. We investigated this question by determining the specific activities and cofactor specificities of all relevant oxidoreductases in cell extracts of H2/CO2-grown C. autoethanogenum. The activity studies were backed up by transcriptional and mutational analyses. Most notably, despite the presence of six hydrogenase systems of various types encoded in the genome, the cells appear to contain only one active hydrogenase. The active [FeFe]-hydrogenase is electron bifurcating, with ferredoxin and NADP as the two electron acceptors. Consistently, most of the other active oxidoreductases rely on either reduced ferredoxin and/or NADPH as the electron donor. An exception is ethanol dehydrogenase, which was found to be NAD specific. Methylenetetrahydrofolate reductase activity could only be demonstrated with artificial electron donors. Key to the understanding of this energy metabolism is the presence of membrane-associated reduced ferredoxin:NAD(+) oxidoreductase (Rnf), of electron-bifurcating and ferredoxin-dependent transhydrogenase (Nfn), and of acetaldehyde:ferredoxin oxidoreductase, which is present with very high specific activities in H2/CO2-grown cells. Based on these findings and on thermodynamic considerations, we propose metabolic schemes that allow, depending on the H2 partial pressure, the chemiosmotic synthesis of 0.14 to 1.5 mol ATP per mol ethanol synthesized from CO2 and H2. Ethanol formation from syngas (H2, CO, and CO2) and from H2 and CO2 that is catalyzed by bacteria is presently a much-discussed process for sustainable production of biofuels. Although the process is already in use, its biochemistry is only incompletely understood. The most pertinent question is how the bacteria conserve energy for growth during ethanol formation from H2 and CO2, considering that acetyl coenzyme A (acetyl-CoA), is an intermediate. Can reduction of the activated acetic acid to ethanol with H2 be coupled with the phosphorylation of ADP? Evidence is presented that this is indeed possible, via both substrate-level phosphorylation and electron transport phosphorylation. In the case of substrate-level phosphorylation, acetyl-CoA reduction to ethanol proceeds via free acetic acid involving acetaldehyde:ferredoxin oxidoreductase (carboxylate reductase).

Electron transport phosphorylation in rumen butyrivibrios: unprecedented ATP yield for glucose fermentation to butyrate

From a genomic analysis of rumen butyrivibrios (Butyrivibrio and Pseudobutyrivibrio sp.), we have re-evaluated the contribution of electron transport phosphorylation (ETP) to ATP formation in this group. This group is unique in that most (76%) genomes were predicted to possess genes for both Ech and Rnf transmembrane ion pumps. These pumps act in concert with the NifJ and Bcd-Etf to form a electrochemical potential (ΔμH+ and ΔμNa+), which drives ATP synthesis by ETP. Of the 62 total butyrivibrio genomes currently available from the Hungate 1000 project, all 62 were predicted to possess NifJ, which reduces oxidized ferredoxin (Fdox) during pyruvate conversion to acetyl-CoA. All 62 possessed all subunits of Bcd-Etf, which reduces Fdox and oxidizes reduced NAD during crotonyl-CoA reduction. Additionally, 61 genomes possessed all subunits of the Rnf, which generates ΔμH+ or ΔμNa+ from oxidation of reduced Fd (Fdred) and reduction of oxidized NAD. Further, 47 genomes possessed all six subunits of the Ech, which generates ΔμH+ from oxidation of Fdred. For glucose fermentation to butyrate and H2, the electrochemical potential established should drive synthesis of ∼1.5 ATP by the F0F1-ATP synthase (possessed by all 62 genomes). The total yield is ∼4.5 ATP/glucose after accounting for three ATP formed by classic substrate-level phosphorylation, and it is one the highest yields for any glucose fermentation. The yield was the same when unsaturated fatty acid bonds, not H+, served as the electron acceptor (as during biohydrogenation). Possession of both Ech and Rnf had been previously documented in only a few sulfate-reducers, was rare in other rumen prokaryotic genomes in our analysis, and may confer an energetic advantage to rumen butyrivibrios. This unique energy conservation system might enhance the butyrivibrios’ ability to overcome growth inhibition by unsaturated fatty acids, as postulated herein.