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Sample records for eukaryotic strong inward-rectifier

  1. ‘Sleepy’ inward rectifier channels in guinea-pig cardiomyocytes are activated only during strong hyperpolarization

    Science.gov (United States)

    Liu, Gong Xin; Daut, Jürgen

    2002-01-01

    K+ channels of isolated guinea-pig cardiomyocytes were studied using the patch-clamp technique. At transmembrane potentials between −120 and −220 mV we observed inward currents through an apparently novel channel. The novel channel was strongly rectifying, no outward currents could be recorded. Between −200 and −160 mV it had a slope conductance of 42.8 ± 3.0 pS (s.d.; n = 96). The open probability (Po) showed a sigmoid voltage dependence and reached a maximum of 0.93 at −200 mV, half-maximal activation was approximately −150 mV. The voltage dependence of Po was not affected by application of 50 μm isoproterenol. The open-time distribution could be described by a single exponential function, the mean open time ranged between 73.5 ms at −220 mV and 1.4 ms at −160 mV. At least two exponential components were required to fit the closed time distribution. Experiments with different external Na+, K+ and Cl− concentrations suggested that the novel channel is K+ selective. Extracellular Ba2+ ions gave rise to a voltage-dependent reduction in Po by inducing long closed states; Cs+ markedly reduced mean open time at −200 mV. In cell-attached recordings the novel channel frequently converted to a classical inward rectifier channel, and vice versa. This conversion was not voltage dependent. After excision of the patch, the novel channel always converted to a classical inward rectifier channel within 0–3 min. This conversion was not affected by intracellular Mg2+, phosphatidylinositol (4,5)-bisphosphate or spermine. Taken together, our findings suggest that the novel K+ channel represents a different ‘mode’ of the classical inward rectifier channel in which opening occurs only at very negative potentials. PMID:11897847

  2. External K+ dependence of strong inward rectifier K+ channel conductance is caused not by K+ but by competitive pore blockade by external Na.

    Science.gov (United States)

    Ishihara, Keiko

    2018-06-15

    Strong inward rectifier K + (sKir) channels determine the membrane potentials of many types of excitable and nonexcitable cells, most notably the resting potentials of cardiac myocytes. They show little outward current during membrane depolarization (i.e., strong inward rectification) because of the channel blockade by cytoplasmic polyamines, which depends on the deviation of the membrane potential from the K + equilibrium potential ( V - E K ) when the extracellular K + concentration ([K + ] out ) is changed. Because their open - channel conductance is apparently proportional to the "square root" of [K + ] out , increases/decreases in [K + ] out enhance/diminish outward currents through sKir channels at membrane potentials near their reversal potential, which also affects, for example, the repolarization and action-potential duration of cardiac myocytes. Despite its importance, however, the mechanism underlying the [K + ] out dependence of the open sKir channel conductance has remained elusive. By studying Kir2.1, the canonical member of the sKir channel family, we first show that the outward currents of Kir2.1 are observed under the external K + -free condition when its inward rectification is reduced and that the complete inhibition of the currents at 0 [K + ] out results solely from pore blockade caused by the polyamines. Moreover, the noted square-root proportionality of the open sKir channel conductance to [K + ] out is mediated by the pore blockade by the external Na + , which is competitive with the external K + Our results show that external K + itself does not activate or facilitate K + permeation through the open sKir channel to mediate the apparent external K + dependence of its open channel conductance. The paradoxical increase/decrease in outward sKir channel currents during alternations in [K + ] out , which is physiologically relevant, is caused by competition from impermeant extracellular Na . © 2018 Ishihara.

  3. Boosting the signal: Endothelial inward rectifier K+ channels.

    Science.gov (United States)

    Jackson, William F

    2017-04-01

    Endothelial cells express a diverse array of ion channels including members of the strong inward rectifier family composed of K IR 2 subunits. These two-membrane spanning domain channels are modulated by their lipid environment, and exist in macromolecular signaling complexes with receptors, protein kinases and other ion channels. Inward rectifier K + channel (K IR ) currents display a region of negative slope conductance at membrane potentials positive to the K + equilibrium potential that allows outward current through the channels to be activated by membrane hyperpolarization, permitting K IR to amplify hyperpolarization induced by other K + channels and ion transporters. Increases in extracellular K + concentration activate K IR allowing them to sense extracellular K + concentration and transduce this change into membrane hyperpolarization. These properties position K IR to participate in the mechanism of action of hyperpolarizing vasodilators and contribute to cell-cell conduction of hyperpolarization along the wall of microvessels. The expression of K IR in capillaries in electrically active tissues may allow K IR to sense extracellular K + , contributing to functional hyperemia. Understanding the regulation of expression and function of microvascular endothelial K IR will improve our understanding of the control of blood flow in the microcirculation in health and disease and may provide new targets for the development of therapeutics in the future. © 2016 John Wiley & Sons Ltd.

  4. Inhibition of cardiac inward rectifier currents by cationic amphiphilic drugs.

    Science.gov (United States)

    van der Heyden, M A G; Stary-Weinzinger, A; Sanchez-Chapula, J A

    2013-09-01

    Cardiac inward rectifier channels belong to three different classes of the KIR channel protein family. The KIR2.x proteins generate the classical inward rectifier current, IK1, while KIR3 and KIR6 members are responsible for the acetylcholine responsive and ATP sensitive inward rectifier currents IKAch and IKATP, respectively. Aberrant function of these channels has been correlated with severe cardiac arrhythmias, indicating their significant contribution to normal cardiac electrophysiology. A common feature of inward rectifier channels is their dependence on the lipid phosphatidyl-4,5-bisphospate (PIP2) interaction for functional activity. Cationic amphiphilic drugs (CADs) are one of the largest classes of pharmaceutical compounds. Several widely used CADs have been associated with inward rectifier current disturbances, and recent evidence points to interference of the channel-PIP2 interaction as the underlying mechanism of action. Here, we will review how six of these well known drugs, used for treatment in various different conditions, interfere in cardiac inward rectifier functioning. In contrast, KIR channel inhibition by the anionic anesthetic thiopental is achieved by a different mechanism of channel-PIP2 interference. We will discuss the latest basic science insights of functional inward rectifier current characteristics, recently derived KIR channel structures and specific PIP2-receptor interactions at the molecular level and provide insight in how these drugs interfere in the structure-function relationships.

  5. Inward rectifier potassium currents in mammalian skeletal muscle fibres

    Science.gov (United States)

    DiFranco, Marino; Yu, Carl; Quiñonez, Marbella; Vergara, Julio L

    2015-01-01

    Inward rectifying potassium (Kir) channels play a central role in maintaining the resting membrane potential of skeletal muscle fibres. Nevertheless their role has been poorly studied in mammalian muscles. Immunohistochemical and transgenic expression were used to assess the molecular identity and subcellular localization of Kir channel isoforms. We found that Kir2.1 and Kir2.2 channels were targeted to both the surface andthe transverse tubular system membrane (TTS) compartments and that both isoforms can be overexpressed up to 3-fold 2 weeks after transfection. Inward rectifying currents (IKir) had the canonical features of quasi-instantaneous activation, strong inward rectification, depended on the external [K+], and could be blocked by Ba2+ or Rb+. In addition, IKir records show notable decays during large 100 ms hyperpolarizing pulses. Most of these properties were recapitulated by model simulations of the electrical properties of the muscle fibre as long as Kir channels were assumed to be present in the TTS. The model also simultaneously predicted the characteristics of membrane potential changes of the TTS, as reported optically by a fluorescent potentiometric dye. The activation of IKir by large hyperpolarizations resulted in significant attenuation of the optical signals with respect to the expectation for equal magnitude depolarizations; blocking IKir with Ba2+ (or Rb+) eliminated this attenuation. The experimental data, including the kinetic properties of IKir and TTS voltage records, and the voltage dependence of peak IKir, while measured at widely dissimilar bulk [K+] (96 and 24 mm), were closely predicted by assuming Kir permeability (PKir) values of ∼5.5 × 10−6 cm s−1 and equal distribution of Kir channels at the surface and TTS membranes. The decay of IKir records and the simultaneous increase in TTS voltage changes were mostly explained by K+ depletion from the TTS lumen. Most importantly, aside from allowing an accurate estimation of

  6. G-protein-coupled inward rectifier potassium current contributes to ventricular repolarization

    DEFF Research Database (Denmark)

    Liang, Bo; Nissen, Jakob D; Laursen, Morten

    2014-01-01

    The purpose of this study was to investigate the functional role of G-protein-coupled inward rectifier potassium (GIRK) channels in the cardiac ventricle.......The purpose of this study was to investigate the functional role of G-protein-coupled inward rectifier potassium (GIRK) channels in the cardiac ventricle....

  7. Long-pore Electrostatics in Inward-rectifier Potassium Channels

    Science.gov (United States)

    Robertson, Janice L.; Palmer, Lawrence G.; Roux, Benoît

    2008-01-01

    Inward-rectifier potassium (Kir) channels differ from the canonical K+ channel structure in that they possess a long extended pore (∼85 Å) for ion conduction that reaches deeply into the cytoplasm. This unique structural feature is presumably involved in regulating functional properties specific to Kir channels, such as conductance, rectification block, and ligand-dependent gating. To elucidate the underpinnings of these functional roles, we examine the electrostatics of an ion along this extended pore. Homology models are constructed based on the open-state model of KirBac1.1 for four mammalian Kir channels: Kir1.1/ROMK, Kir2.1/IRK, Kir3.1/GIRK, and Kir6.2/KATP. By solving the Poisson-Boltzmann equation, the electrostatic free energy of a K+ ion is determined along each pore, revealing that mammalian Kir channels provide a favorable environment for cations and suggesting the existence of high-density regions in the cytoplasmic domain and cavity. The contribution from the reaction field (the self-energy arising from the dielectric polarization induced by the ion's charge in the complex geometry of the pore) is unfavorable inside the long pore. However, this is well compensated by the electrostatic interaction with the static field arising from the protein charges and shielded by the dielectric surrounding. Decomposition of the static field provides a list of residues that display remarkable correspondence with existing mutagenesis data identifying amino acids that affect conduction and rectification. Many of these residues demonstrate interactions with the ion over long distances, up to 40 Å, suggesting that mutations potentially affect ion or blocker energetics over the entire pore. These results provide a foundation for understanding ion interactions in Kir channels and extend to the study of ion permeation, block, and gating in long, cation-specific pores. PMID:19001143

  8. Changes in Inward Rectifier K+ Channels in Hepatic Stellate Cells During Primary Culture

    Science.gov (United States)

    Lee, Dong Hyeon; Kong, In Deok; Lee, Joong-Woo

    2008-01-01

    Purpose This study examined the expression and function of inward rectifier K+ channels in cultured rat hepatic stellate cells (HSC). Materials and Methods The expression of inward rectifier K+ channels was measured using real-time RT-PCR, and electrophysiological properties were determined using the gramicidin-perforated patch-clamp technique. Results The dominant inward rectifier K+ channel subtypes were Kir2.1 and Kir6.1. These dominant K+ channel subtypes decreased significantly during the primary culture throughout activation process. HSC can be classified into two subgroups: one with an inward-rectifying K+ current (type 1) and the other without (type 2). The inward current was blocked by Ba2+ (100 µM) and enhanced by high K+ (140 mM), more prominently in type 1 HSC. There was a correlation between the amplitude of the Ba2+-sensitive current and the membrane potential. In addition, Ba2+ (300 µM) depolarized the membrane potential. After the culture period, the amplitude of the inward current decreased and the membrane potential became depolarized. Conclusion HSC express inward rectifier K+ channels, which physiologically regulate membrane potential and decrease during the activation process. These results will potentially help determine properties of the inward rectifier K+ channels in HSC as well as their roles in the activation process. PMID:18581597

  9. Inward Rectifier K+ Currents Are Regulated by CaMKII in Endothelial Cells of Primarily Cultured Bovine Pulmonary Arteries.

    Science.gov (United States)

    Qu, Lihui; Yu, Lei; Wang, Yanli; Jin, Xin; Zhang, Qianlong; Lu, Ping; Yu, Xiufeng; Zhong, Weiwei; Zheng, Xiaodong; Cui, Ningren; Jiang, Chun; Zhu, Daling

    2015-01-01

    Endothelium lines the interior surface of vascular walls and regulates vascular tones. The endothelial cells sense and respond to chemical and mechanical stimuli in the circulation, and couple the stimulus signals to vascular smooth muscles, in which inward rectifier K+ currents (Kir) play an important role. Here we applied several complementary strategies to determine the Kir subunit in primarily cultured pulmonary arterial endothelial cells (PAECs) that was regulated by the Ca2+/calmodulin (CaM)-dependent protein kinase II (CaMKII). In whole-cell voltage clamp, the Kir currents were sensitive to micromolar concentrations of extracellular Ba2+. In excised inside-out patches, an inward rectifier K+ current was observed with single-channel conductance 32.43 ± 0.45 pS and Popen 0.27 ± 0.04, which were consistent with known unitary conductance of Kir 2.1. RT-PCR and western blot results showed that expression of Kir 2.1 was significantly stronger than that of other subtypes in PAECs. Pharmacological analysis of the Kir currents demonstrated that insensitivity to intracellular ATP, pinacidil, glibenclamide, pH, GDP-β-S and choleratoxin suggested that currents weren't determined by KATP, Kir2.3, Kir2.4 and Kir3.x. The currents were strongly suppressed by exposure to CaMKII inhibitor W-7 and KN-62. The expression of Kir2.1 was inhibited by knocking down CaMKII. Consistently, vasodilation was suppressed by Ba2+, W-7 and KN-62 in isolated and perfused pulmonary arterial rings. These results suggest that the PAECs express an inward rectifier K+ current that is carried dominantly by Kir2.1, and this K+ channel appears to be targeted by CaMKII-dependent intracellular signaling systems.

  10. Inward Rectifier K+ Currents Are Regulated by CaMKII in Endothelial Cells of Primarily Cultured Bovine Pulmonary Arteries.

    Directory of Open Access Journals (Sweden)

    Lihui Qu

    Full Text Available Endothelium lines the interior surface of vascular walls and regulates vascular tones. The endothelial cells sense and respond to chemical and mechanical stimuli in the circulation, and couple the stimulus signals to vascular smooth muscles, in which inward rectifier K+ currents (Kir play an important role. Here we applied several complementary strategies to determine the Kir subunit in primarily cultured pulmonary arterial endothelial cells (PAECs that was regulated by the Ca2+/calmodulin (CaM-dependent protein kinase II (CaMKII. In whole-cell voltage clamp, the Kir currents were sensitive to micromolar concentrations of extracellular Ba2+. In excised inside-out patches, an inward rectifier K+ current was observed with single-channel conductance 32.43 ± 0.45 pS and Popen 0.27 ± 0.04, which were consistent with known unitary conductance of Kir 2.1. RT-PCR and western blot results showed that expression of Kir 2.1 was significantly stronger than that of other subtypes in PAECs. Pharmacological analysis of the Kir currents demonstrated that insensitivity to intracellular ATP, pinacidil, glibenclamide, pH, GDP-β-S and choleratoxin suggested that currents weren't determined by KATP, Kir2.3, Kir2.4 and Kir3.x. The currents were strongly suppressed by exposure to CaMKII inhibitor W-7 and KN-62. The expression of Kir2.1 was inhibited by knocking down CaMKII. Consistently, vasodilation was suppressed by Ba2+, W-7 and KN-62 in isolated and perfused pulmonary arterial rings. These results suggest that the PAECs express an inward rectifier K+ current that is carried dominantly by Kir2.1, and this K+ channel appears to be targeted by CaMKII-dependent intracellular signaling systems.

  11. Effect of ethanol at clinically relevant concentrations on atrial inward rectifier potassium current sensitive to acetylcholine

    Czech Academy of Sciences Publication Activity Database

    Bébarová, M.; Matejovič, P.; Pásek, Michal; Hořáková, Z.; Hošek, J.; Šimurdová, M.; Šimurda, J.

    2016-01-01

    Roč. 389, č. 10 (2016), s. 1049-1058 ISSN 0028-1298 Institutional support: RVO:61388998 Keywords : arrhythmias * atrial cardiomyocyte * inward rectifier potasssium current * ethanol * rat atrial cell model Subject RIV: BO - Biophysics Impact factor: 2.558, year: 2016

  12. Pharmacological Conversion of a Cardiac Inward Rectifier into an Outward Rectifier Potassium Channel.

    Science.gov (United States)

    Moreno-Galindo, Eloy G; Sanchez-Chapula, Jose A; Tristani-Firouzi, Martin; Navarro-Polanco, Ricardo A

    2016-09-01

    Potassium (K(+)) channels are crucial for determining the shape, duration, and frequency of action-potential firing in excitable cells. Broadly speaking, K(+) channels can be classified based on whether their macroscopic current outwardly or inwardly rectifies, whereby rectification refers to a change in conductance with voltage. Outwardly rectifying K(+) channels conduct greater current at depolarized membrane potentials, whereas inward rectifier channels conduct greater current at hyperpolarized membrane potentials. Under most circumstances, outward currents through inwardly rectifying K(+) channels are reduced at more depolarized potentials. However, the acetylcholine-gated K(+) channel (KACh) conducts current that inwardly rectifies when activated by some ligands (such as acetylcholine), and yet conducts current that outwardly rectifies when activated by other ligands (for example, pilocarpine and choline). The perplexing and paradoxical behavior of KACh channels is due to the intrinsic voltage sensitivity of the receptor that activates KACh channels, the M2 muscarinic receptor (M2R). Emerging evidence reveals that the affinity of M2R for distinct ligands varies in a voltage-dependent and ligand-specific manner. These intrinsic receptor properties determine whether current conducted by KACh channels inwardly or outwardly rectifies. This review summarizes the most recent concepts regarding the intrinsic voltage sensitivity of muscarinic receptors and the consequences of this intriguing behavior on cardiac physiology and pharmacology of KACh channels. Copyright © 2016 by The American Society for Pharmacology and Experimental Therapeutics.

  13. Specific residues of the cytoplasmic domains of cardiac inward rectifier potassium channels are effective antifibrillatory targets

    Science.gov (United States)

    Noujaim, Sami F.; Stuckey, Jeanne A.; Ponce-Balbuena, Daniela; Ferrer-Villada, Tania; López-Izquierdo, Angelica; Pandit, Sandeep; Calvo, Conrado J.; Grzeda, Krzysztof R.; Berenfeld, Omer; Sánchez Chapula, José A.; Jalife, José

    2010-01-01

    Atrial and ventricular tachyarrhythmias can be perpetuated by up-regulation of inward rectifier potassium channels. Thus, it may be beneficial to block inward rectifier channels under conditions in which their function becomes arrhythmogenic (e.g., inherited gain-of-function mutation channelopathies, ischemia, and chronic and vagally mediated atrial fibrillation). We hypothesize that the antimalarial quinoline chloroquine exerts potent antiarrhythmic effects by interacting with the cytoplasmic domains of Kir2.1 (IK1), Kir3.1 (IKACh), or Kir6.2 (IKATP) and reducing inward rectifier potassium currents. In isolated hearts of three different mammalian species, intracoronary chloroquine perfusion reduced fibrillatory frequency (atrial or ventricular), and effectively terminated the arrhythmia with resumption of sinus rhythm. In patch-clamp experiments chloroquine blocked IK1, IKACh, and IKATP. Comparative molecular modeling and ligand docking of chloroquine in the intracellular domains of Kir2.1, Kir3.1, and Kir6.2 suggested that chloroquine blocks or reduces potassium flow by interacting with negatively charged amino acids facing the ion permeation vestibule of the channel in question. These results open a novel path toward discovering antiarrhythmic pharmacophores that target specific residues of the cytoplasmic domain of inward rectifier potassium channels.—Noujaim, S. F., Stuckey, J. A., Ponce-Balbuena, D., Ferrer-Villada, T., López-Izquierdo, A., Pandit, S., Calvo, C. J., Grzeda, K. R., Berenfeld, O., Sánchez Chapula, J. A., Jalife, J. Specific residues of the cytoplasmic domains of cardiac inward rectifier potassium channels are effective antifibrillatory targets. PMID:20585026

  14. Inward rectifier potassium current IKir promotes intrinsic pacemaker activity of thalamocortical neurons.

    Science.gov (United States)

    Amarillo, Yimy; Tissone, Angela I; Mato, Germán; Nadal, Marcela S

    2018-06-01

    Slow repetitive burst firing by hyperpolarized thalamocortical (TC) neurons correlates with global slow rhythms (rectifier potassium current I Kir induces repetitive burst firing at slow and delta frequency bands. We demonstrate this in mouse TC neurons in brain slices by manipulating the Kir maximum conductance with dynamic clamp. We also performed a thorough theoretical analysis that explains how the unique properties of I Kir enable this current to induce slow periodic bursting in TC neurons. We describe a new ionic mechanism based on the voltage- and time-dependent interaction of I Kir and hyperpolarization-activated cationic current I h that endows TC neurons with the ability to oscillate spontaneously at very low frequencies, even below 0.5 Hz. Bifurcation analysis of conductance-based models of increasing complexity demonstrates that I Kir induces bistability of the membrane potential at the same time that it induces sustained oscillations in combination with I h and increases the robustness of low threshold-activated calcium current I T -mediated oscillations. NEW & NOTEWORTHY The strong inwardly rectifying potassium current I Kir of thalamocortical neurons displays a region of negative slope conductance in the current-voltage relationship that generates potassium currents activated by hyperpolarization. Bifurcation analysis shows that I Kir induces bistability of the membrane potential; generates sustained subthreshold oscillations by interacting with the hyperpolarization-activated cationic current I h ; and increases the robustness of oscillations mediated by the low threshold-activated calcium current I T . Upregulation of I Kir in thalamocortical neurons induces repetitive burst firing at slow and delta frequency bands (<4 Hz).

  15. The inward rectifier potassium channel Kir2.1 is expressed in mouse neutrophils from bone marrow and liver.

    Science.gov (United States)

    Masia, Ricard; Krause, Daniela S; Yellen, Gary

    2015-02-01

    Neutrophils are phagocytic cells that play a critical role in innate immunity by destroying bacterial pathogens. Channels belonging to the inward rectifier potassium channel subfamily 2 (Kir2 channels) have been described in other phagocytes (monocytes/macrophages and eosinophils) and in hematopoietic precursors of phagocytes. Their physiological function in these cells remains unclear, but some evidence suggests a role in growth factor-dependent proliferation and development. Expression of functional Kir2 channels has not been definitively demonstrated in mammalian neutrophils. Here, we show by RT-PCR that neutrophils from mouse bone marrow and liver express mRNA for the Kir2 subunit Kir2.1 but not for other subunits (Kir2.2, Kir2.3, and Kir2.4). In electrophysiological experiments, resting (unstimulated) neutrophils from mouse bone marrow and liver exhibit a constitutively active, external K(+)-dependent, strong inwardly rectifying current that constitutes the dominant current. The reversal potential is dependent on the external K(+) concentration in a Nernstian fashion, as expected for a K(+)-selective current. The current is not altered by changes in external or internal pH, and it is blocked by Ba(2+), Cs(+), and the Kir2-selective inhibitor ML133. The single-channel conductance is in agreement with previously reported values for Kir2.1 channels. These properties are characteristic of homomeric Kir2.1 channels. Current density in short-term cultures of bone marrow neutrophils is decreased in the absence of growth factors that are important for neutrophil proliferation [granulocyte colony-stimulating factor (G-CSF) and stem cell factor (SCF)]. These results demonstrate that mouse neutrophils express functional Kir2.1 channels and suggest that these channels may be important for neutrophil function, possibly in a growth factor-dependent manner. Copyright © 2015 the American Physiological Society.

  16. Dynamics of the inward rectifier K+ current during the action potential of guinea pig ventricular myocytes

    OpenAIRE

    Ibarra, J.; Morley, G.E.; Delmar, M.

    1991-01-01

    The potassium selective, inward rectifier current (IK1) is known to be responsible for maintaining the resting membrane potential of quiescent ventricular myocytes. However, the contribution of this current to the different phases of the cardiac action potential has not been adequately established. In the present study, we have used the action potential clamp (APC) technique to characterize the dynamic changes of a cesium-sensitive (i.e., Ik1) current which occur during the action potential. ...

  17. Inward rectifier potassium channels in the HL-1 cardiomyocyte-derived cell line.

    Science.gov (United States)

    Goldoni, Dana; Zhao, YouYou; Green, Brian D; McDermott, Barbara J; Collins, Anthony

    2010-11-01

    HL-1 is a line of immortalized cells of cardiomyocyte origin that are a useful complement to native cardiomyocytes in studies of cardiac gene regulation. Several types of ion channel have been identified in these cells, but not the physiologically important inward rectifier K(+) channels. Our aim was to identify and characterize inward rectifier K(+) channels in HL-1 cells. External Ba(2+) (100 µM) inhibited 44 ± 0.05% (mean ± s.e.m., n = 11) of inward current in whole-cell patch-clamp recordings. The reversal potential of the Ba(2+)-sensitive current shifted with external [K(+)] as expected for K(+)-selective channels. The slope conductance of the inward Ba(2+)-sensitive current increased with external [K(+)]. The apparent Kd for Ba(2+) was voltage dependent, ranging from 15 µM at -150  mV to 148 µM at -75  mV in 120  mM external K(+). This current was insensitive to 10 µM glybenclamide. A component of whole-cell current was sensitive to 150 µM 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS), although it did not correspond to the Ba(2+)-sensitive component. The effect of external 1 mM Cs(+) was similar to that of Ba(2+). Polymerase chain reaction using HL-1 cDNA as template and primers specific for the cardiac inward rectifier K(ir)2.1 produced a fragment of the expected size that was confirmed to be K(ir)2.1 by DNA sequencing. In conclusion, HL-1 cells express a current that is characteristic of cardiac inward rectifier K(+) channels, and express K(ir)2.1 mRNA. This cell line may have use as a system for studying inward rectifier gene regulation in a cardiomyocyte phenotype. © 2010 Wiley-Liss, Inc.

  18. Inhibition of the cardiac inward rectifier potassium currents by KB-R7943.

    Science.gov (United States)

    Abramochkin, Denis V; Alekseeva, Eugenia I; Vornanen, Matti

    2013-09-01

    KB-R7943 (2-[2-[4-(4-nitrobenzyloxy)phenyl]ethyl]isothiourea) was developed as a specific inhibitor of the sarcolemmal sodium-calcium exchanger (NCX) with potential experimental and therapeutic use. However, KB-R7943 is shown to be a potent blocker of several ion currents including inward and delayed rectifier K(+) currents of cardiomyocytes. To further characterize KB-R7943 as a blocker of the cardiac inward rectifiers we compared KB-R7943 sensitivity of the background inward rectifier (IK1) and the carbacholine-induced inward rectifier (IKACh) currents in mammalian (Rattus norvegicus; rat) and fish (Carassius carassius; crucian carp) cardiac myocytes. The basal IK1 of ventricular myocytes was blocked with apparent IC50-values of 4.6×10(-6) M and 3.5×10(-6) M for rat and fish, respectively. IKACh was almost an order of magnitude more sensitive to KB-R7943 than IK1 with IC50-values of 6.2×10(-7) M for rat and 2.5×10(-7) M for fish. The fish cardiac NCX current was half-maximally blocked at the concentration of 1.9-3×10(-6) M in both forward and reversed mode of operation. Thus, the sensitivity of three cardiac currents to KB-R7943 block increases in the order IK1~INCXrectifier potassium currents, in particular IKACh, should be taken into account when interpreting the data with this inhibitor from in vivo and in vitro experiments in both mammalian and fish models. © 2013.

  19. OSR1 regulates a subset of inward rectifier potassium channels via a binding motif variant.

    Science.gov (United States)

    Taylor, Clinton A; An, Sung-Wan; Kankanamalage, Sachith Gallolu; Stippec, Steve; Earnest, Svetlana; Trivedi, Ashesh T; Yang, Jonathan Zijiang; Mirzaei, Hamid; Huang, Chou-Long; Cobb, Melanie H

    2018-04-10

    The with-no-lysine (K) (WNK) signaling pathway to STE20/SPS1-related proline- and alanine-rich kinase (SPAK) and oxidative stress-responsive 1 (OSR1) kinase is an important mediator of cell volume and ion transport. SPAK and OSR1 associate with upstream kinases WNK 1-4, substrates, and other proteins through their C-terminal domains which interact with linear R-F-x-V/I sequence motifs. In this study we find that SPAK and OSR1 also interact with similar affinity with a motif variant, R-x-F-x-V/I. Eight of 16 human inward rectifier K + channels have an R-x-F-x-V motif. We demonstrate that two of these channels, Kir2.1 and Kir2.3, are activated by OSR1, while Kir4.1, which does not contain the motif, is not sensitive to changes in OSR1 or WNK activity. Mutation of the motif prevents activation of Kir2.3 by OSR1. Both siRNA knockdown of OSR1 and chemical inhibition of WNK activity disrupt NaCl-induced plasma membrane localization of Kir2.3. Our results suggest a mechanism by which WNK-OSR1 enhance Kir2.1 and Kir2.3 channel activity by increasing their plasma membrane localization. Regulation of members of the inward rectifier K + channel family adds functional and mechanistic insight into the physiological impact of the WNK pathway.

  20. Structural basis of control of inward rectifier Kir2 channel gating by bulk anionic phospholipids.

    Science.gov (United States)

    Lee, Sun-Joo; Ren, Feifei; Zangerl-Plessl, Eva-Maria; Heyman, Sarah; Stary-Weinzinger, Anna; Yuan, Peng; Nichols, Colin G

    2016-09-01

    Inward rectifier potassium (Kir) channel activity is controlled by plasma membrane lipids. Phosphatidylinositol-4,5-bisphosphate (PIP2) binding to a primary site is required for opening of classic inward rectifier Kir2.1 and Kir2.2 channels, but interaction of bulk anionic phospholipid (PL(-)) with a distinct second site is required for high PIP2 sensitivity. Here we show that introduction of a lipid-partitioning tryptophan at the second site (K62W) generates high PIP2 sensitivity, even in the absence of PL(-) Furthermore, high-resolution x-ray crystal structures of Kir2.2[K62W], with or without added PIP2 (2.8- and 2.0-Å resolution, respectively), reveal tight tethering of the C-terminal domain (CTD) to the transmembrane domain (TMD) in each condition. Our results suggest a refined model for phospholipid gating in which PL(-) binding at the second site pulls the CTD toward the membrane, inducing the formation of the high-affinity primary PIP2 site and explaining the positive allostery between PL(-) binding and PIP2 sensitivity. © 2016 Lee et al.

  1. G-protein-coupled inward rectifier potassium channels involved in corticostriatal presynaptic modulation.

    Science.gov (United States)

    Meneses, David; Mateos, Verónica; Islas, Gustavo; Barral, Jaime

    2015-09-01

    Presynaptic modulation has been associated mainly with calcium channels but recent data suggests that inward rectifier potassium channels (K(IR)) also play a role. In this work we set to characterize the role of presynaptic K(IR) channels in corticostriatal synaptic transmission. We elicited synaptic potentials in striatum by stimulating cortical areas and then determined the synaptic responses of corticostriatal synapsis by using paired pulse ratio (PPR) in the presence and absence of several potassium channel blockers. Unspecific potassium channels blockers Ba(2+) and Cs(+) reduced the PPR, suggesting that these channels are presynaptically located. Further pharmacological characterization showed that application of tertiapin-Q, a specific K(IR)3 channel family blocker, also induced a reduction of PPR, suggesting that K(IR)3 channels are present at corticostriatal terminals. In contrast, exposure to Lq2, a specific K(IR)1.1 inward rectifier potassium channel, did not induce any change in PPR suggesting the absence of these channels in the presynaptic corticostriatal terminals. Our results indicate that K(IR)3 channels are functionally expressed at the corticostriatal synapses, since blockage of these channels result in PPR decrease. Our results also help to explain how synaptic activity may become sensitive to extracellular signals mediated by G-protein coupled receptors. A vast repertoire of receptors may influence neurotransmitter release in an indirect manner through regulation of K(IR)3 channels. © 2015 Wiley Periodicals, Inc.

  2. Agonist of inward rectifier K+ channels enhances the protection of ischemic postconditioning in isolated rat hearts.

    Science.gov (United States)

    Liao, Z; Feng, Z; Long, C

    2014-07-01

    Selective inhibition of inward rectifier K + channels could abolish the protection mediated by ischemic preconditioning, but the roles of these channels in ischemic postconditioning have not been well characterized. Our study aims to evaluate the effect of inward rectifier K + channels on the protection induced by ischemic postconditioning. Langendorff-perfused rat hearts (n=8 per group) were split into four groups: postconditioning hearts (IPO group); ischemic postconditioning with BaCl 2 hearts (PB group); ischemic postconditioning with zacopride hearts (PZ group); and without ischemic postconditioning (CON group). After suffering 30 minutes of global ischemia, groups IPO, PB and PZ went through 10 seconds of ischemic postconditioning with three different perfusates: respectively, Krebs-Henseleit buffer (IPO group); 20 μmol/L BaCl 2 (antagonist of the channel, PB group); 1 μmol/L zacopride (agonist of the channel, PZ group). At the end of reperfusion, the myocardial performance was better preserved in the PZ group than the other three groups. The PB group showed no significant differences from the CON group. Our study has shown that the I K1 channel agonist zacopride is associated with the enhancement of ischemic postconditioning. © The Author(s) 2014.

  3. Structural determinants of PIP(2) regulation of inward rectifier K(ATP) channels.

    Science.gov (United States)

    Shyng, S L; Cukras, C A; Harwood, J; Nichols, C G

    2000-11-01

    Phosphatidylinositol 4,5-bisphosphate (PIP(2)) activates K(ATP) and other inward rectifier (Kir) channels. To determine residues important for PIP(2) regulation, we have systematically mutated each positive charge in the COOH terminus of Kir6.2 to alanine. The effects of these mutations on channel function were examined using (86)Rb efflux assays on intact cells and inside-out patch-clamp methods. Both methods identify essentially the same basic residues in two narrow regions (176-222 and 301-314) in the COOH terminus that are important for the maintenance of channel function and interaction with PIP(2). Only one residue (R201A) simultaneously affected ATP and PIP(2) sensitivity, which is consistent with the notion that these ligands, while functionally competitive, are unlikely to bind to identical sites. Strikingly, none of 13 basic residues in the terminal portion (residues 315-390) of the COOH terminus affected channel function when neutralized. The data help to define the structural requirements for PIP(2) sensitivity of K(ATP) channels. Moreover, the regions and residues defined in this study parallel those uncovered in recent studies of PIP(2) sensitivity in other inward rectifier channels, indicating a common structural basis for PIP(2) regulation.

  4. Inward rectifier potassium (Kir2.1) channels as end-stage boosters of endothelium-dependent vasodilators.

    Science.gov (United States)

    Sonkusare, Swapnil K; Dalsgaard, Thomas; Bonev, Adrian D; Nelson, Mark T

    2016-06-15

    Increase in endothelial cell (EC) calcium activates calcium-sensitive intermediate and small conductance potassium (IK and SK) channels, thereby causing hyperpolarization and endothelium-dependent vasodilatation. Endothelial cells express inward rectifier potassium (Kir) channels, but their role in endothelium-dependent vasodilatation is not clear. In the mesenteric arteries, only ECs, but not smooth muscle cells, displayed Kir currents that were predominantly mediated by the Kir2.1 isoform. Endothelium-dependent vasodilatations in response to muscarinic receptor, TRPV4 (transient receptor potential vanilloid 4) channel and IK/SK channel agonists were highly attenuated by Kir channel inhibitors and by Kir2.1 channel knockdown. These results point to EC Kir channels as amplifiers of vasodilatation in response to increases in EC calcium and IK/SK channel activation and suggest that EC Kir channels could be targeted to treat endothelial dysfunction, which is a hallmark of vascular disorders. Endothelium-dependent vasodilators, such as acetylcholine, increase intracellular Ca(2+) through activation of transient receptor potential vanilloid 4 (TRPV4) channels in the plasma membrane and inositol trisphosphate receptors in the endoplasmic reticulum, leading to stimulation of Ca(2+) -sensitive intermediate and small conductance K(+) (IK and SK, respectively) channels. Although strong inward rectifier K(+) (Kir) channels have been reported in the native endothelial cells (ECs) their role in EC-dependent vasodilatation is not clear. Here, we test the idea that Kir channels boost the EC-dependent vasodilatation of resistance-sized arteries. We show that ECs, but not smooth muscle cells, of small mesenteric arteries have Kir currents, which are substantially reduced in EC-specific Kir2.1 knockdown (EC-Kir2.1(-/-) ) mice. Elevation of extracellular K(+) to 14 mm caused vasodilatation of pressurized arteries, which was prevented by endothelial denudation and Kir channel

  5. Decreased inward rectifier potassium current IK1 in dystrophin-deficient ventricular cardiomyocytes.

    Science.gov (United States)

    Rubi, Lena; Koenig, Xaver; Kubista, Helmut; Todt, Hannes; Hilber, Karlheinz

    2017-03-04

    Kir2.x channels in ventricular cardiomyocytes (most prominently Kir2.1) account for the inward rectifier potassium current I K1 , which controls the resting membrane potential and the final phase of action potential repolarization. Recently it was hypothesized that the dystrophin-associated protein complex (DAPC) is important in the regulation of Kir2.x channels. To test this hypothesis, we investigated potential I K1 abnormalities in dystrophin-deficient ventricular cardiomyocytes derived from the hearts of Duchenne muscular dystrophy mouse models. We found that I K1 was substantially diminished in dystrophin-deficient cardiomyocytes when compared to wild type myocytes. This finding represents the first functional evidence for a significant role of the DAPC in the regulation of Kir2.x channels.

  6. Characterization of the chicken inward rectifier K+ channel IRK1/Kir2.1 gene

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    Locke Emily

    2004-11-01

    Full Text Available Abstract Background Inward rectifier potassium channels (IRK contribute to the normal function of skeletal and cardiac muscle cells. The chick inward rectifier K+ channel cIRK1/Kir2.1 is expressed in skeletal muscle, heart, brain, but not in liver; a distribution similar but not identical to that of mouse Kir2.1. We set out to explore regulatory domains of the cIRK1 promoter that enhance or inhibit expression of the gene in different cell types. Results We cloned and characterized the 5'-flanking region of cIRK1. cIRK1 contains two exons with splice sites in the 5'-untranslated region, a structure similar to mouse and human orthologs. cIRK1 has multiple transcription initiation sites, a feature also seen in mouse. However, while the chicken and mouse promoter regions share many regulatory motifs, cIRK1 possesses a GC-richer promoter and a putative TATA box, which appears to positively regulate gene expression. We report here the identification of several candidate cell/tissue specific cIRK1 regulatory domains by comparing promoter activities in expressing (Qm7 and non-expressing (DF1 cells using in vitro transcription assays. Conclusion While multiple transcription initiation sites and the combinatorial function of several domains in activating cIRK1 expression are similar to those seen in mKir2.1, the cIRK1 promoter differs by the presence of a putative TATA box. In addition, several domains that regulate the gene's expression differentially in muscle (Qm7 and fibroblast cells (DF1 were identified. These results provide fundamental data to analyze cIRK1 transcriptional mechanisms. The control elements identified here may provide clues to the tissue-specific expression of this K+ channel.

  7. Biophysical characterization of inwardly rectifying potassium currents (I(K1) I(K,ACh), I(K,Ca)) using sinus rhythm or atrial fibrillation action potential waveforms

    DEFF Research Database (Denmark)

    Tang, Chuyi; Skibsbye, Lasse; Yuan, Lei

    2015-01-01

    Although several physiological, pathophysiological and regulatory properties of classical inward rectifier K+ current I(K1), G-protein coupled inwardly-rectifying K+ current I(K,ACh) and the small-conductance Ca2+ activated K+ current I(K,Ca) have been identified, quantitative biophysical details...

  8. Role of inward rectifier potassium channels in salivary gland function and sugar feeding of the fruit fly, Drosophila melanogaster

    Science.gov (United States)

    The arthropod salivary gland is of critical importance for horizontal transmission of pathogens, yet a detailed understanding of the ion conductance pathways responsible for saliva production and excretion is lacking. A superfamily of potassium ion channels, known as inward rectifying potassium (Ki...

  9. Inward rectifier potassium (Kir2.1) channels as end‐stage boosters of endothelium‐dependent vasodilators

    Science.gov (United States)

    Dalsgaard, Thomas; Bonev, Adrian D.; Nelson, Mark T.

    2016-01-01

    Key points Increase in endothelial cell (EC) calcium activates calcium‐sensitive intermediate and small conductance potassium (IK and SK) channels, thereby causing hyperpolarization and endothelium‐dependent vasodilatation.Endothelial cells express inward rectifier potassium (Kir) channels, but their role in endothelium‐dependent vasodilatation is not clear.In the mesenteric arteries, only ECs, but not smooth muscle cells, displayed Kir currents that were predominantly mediated by the Kir2.1 isoform.Endothelium‐dependent vasodilatations in response to muscarinic receptor, TRPV4 (transient receptor potential vanilloid 4) channel and IK/SK channel agonists were highly attenuated by Kir channel inhibitors and by Kir2.1 channel knockdown.These results point to EC Kir channels as amplifiers of vasodilatation in response to increases in EC calcium and IK/SK channel activation and suggest that EC Kir channels could be targeted to treat endothelial dysfunction, which is a hallmark of vascular disorders. Abstract Endothelium‐dependent vasodilators, such as acetylcholine, increase intracellular Ca2+ through activation of transient receptor potential vanilloid 4 (TRPV4) channels in the plasma membrane and inositol trisphosphate receptors in the endoplasmic reticulum, leading to stimulation of Ca2+‐sensitive intermediate and small conductance K+ (IK and SK, respectively) channels. Although strong inward rectifier K+ (Kir) channels have been reported in the native endothelial cells (ECs) their role in EC‐dependent vasodilatation is not clear. Here, we test the idea that Kir channels boost the EC‐dependent vasodilatation of resistance‐sized arteries. We show that ECs, but not smooth muscle cells, of small mesenteric arteries have Kir currents, which are substantially reduced in EC‐specific Kir2.1 knockdown (EC‐Kir2.1 −/−) mice. Elevation of extracellular K+ to 14 mm caused vasodilatation of pressurized arteries, which was prevented by endothelial

  10. Inwardly rectifying potassium channels influence Drosophila wing morphogenesis by regulating Dpp release.

    Science.gov (United States)

    Dahal, Giri Raj; Pradhan, Sarala Joshi; Bates, Emily Anne

    2017-08-01

    Loss of embryonic ion channel function leads to morphological defects, but the underlying reason for these defects remains elusive. Here, we show that inwardly rectifying potassium (Irk) channels regulate release of the Drosophila bone morphogenetic protein Dpp in the developing fly wing and that this is necessary for developmental signaling. Inhibition of Irk channels decreases the incidence of distinct Dpp-GFP release events above baseline fluorescence while leading to a broader distribution of Dpp-GFP. Work by others in different cell types has shown that Irk channels regulate peptide release by modulating membrane potential and calcium levels. We found calcium transients in the developing wing, and inhibition of Irk channels reduces the duration and amplitude of calcium transients. Depolarization with high extracellular potassium evokes Dpp release. Taken together, our data implicate Irk channels as a requirement for regulated release of Dpp, highlighting the importance of the temporal pattern of Dpp presentation for morphogenesis of the wing. © 2017. Published by The Company of Biologists Ltd.

  11. Role of an inward rectifier K+ current and of hyperpolarization in human myoblast fusion

    Science.gov (United States)

    Liu, J-H; Bijlenga, P; Fischer-Lougheed, J; Occhiodoro, T; Kaelin, A; Bader, C R; Bernheim, L

    1998-01-01

    The role of K+ channels and membrane potential in myoblast fusion was evaluated by examining resting membrane potential and timing of expression of K+ currents at three stages of differentiation of human myogenic cells: undifferentiated myoblasts, fusion-competent myoblasts (FCMBs), and freshly formed myotubes. Two K+ currents contribute to a hyperpolarization of myoblasts prior to fusion: IK(NI), a non-inactivating delayed rectifier, and IK(IR), an inward rectifier. IK(NI) density is low in undifferentiated myoblasts, increases in FCMBs and declines in myotubes. On the other hand, IK(IR) is expressed in 28 % of the FCMBs and in all myotubes. IK(IR) is reversibly blocked by Ba2+ or Cs+. Cells expressing IK(IR) have resting membrane potentials of −65 mV. A block by Ba2+ or Cs+ induces a depolarization to a voltage determined by IK(NI) (−32 mV). Cs+ and Ba2+ ions reduce myoblast fusion. It is hypothesized that the IK(IR)-mediated hyperpolarization allows FCMBs to recruit Na+, K+ and T-type Ca2+ channels which are present in these cells and would otherwise be inactivated. FCMBs, rendered thereby capable of firing action potentials, could amplify depolarizing signals and may accelerate fusion. PMID:9705997

  12. The role of NH2-terminal positive charges in the activity of inward rectifier KATP channels.

    Science.gov (United States)

    Cukras, C A; Jeliazkova, I; Nichols, C G

    2002-09-01

    Approximately half of the NH(2) terminus of inward rectifier (Kir) channels can be deleted without significant change in channel function, but activity is lost when more than approximately 30 conserved residues before the first membrane spanning domain (M1) are removed. Systematic replacement of the positive charges in the NH(2) terminus of Kir6.2 with alanine reveals several residues that affect channel function when neutralized. Certain mutations (R4A, R5A, R16A, R27A, R39A, K47A, R50A, R54A, K67A) change open probability, whereas an overlapping set of mutants (R16A, R27A, K39A, K47A, R50A, R54A, K67A) change ATP sensitivity. Further analysis of the latter set differentiates mutations that alter ATP sensitivity as a consequence of altered open state stability (R16A, K39A, K67A) from those that may affect ATP binding directly (K47A, R50A, R54A). The data help to define the structural determinants of Kir channel function, and suggest possible structural motifs within the NH(2) terminus, as well as the relationship of the NH(2) terminus with the extended cytoplasmic COOH terminus of the channel.

  13. Activation of the Ca2+-sensing receptors increases currents through inward rectifier K+ channels via activation of phosphatidylinositol 4-kinase.

    Science.gov (United States)

    Liu, Chung-Hung; Chang, Hsueh-Kai; Lee, Sue-Ping; Shieh, Ru-Chi

    2016-11-01

    Inward rectifier K + channels are important for maintaining normal electrical function in many cell types. The proper function of these channels requires the presence of membrane phosphoinositide 4,5-bisphosphate (PIP 2 ). Stimulation of the Ca 2+ -sensing receptor CaR, a pleiotropic G protein-coupled receptor, activates both G q/11 , which decreases PIP 2 , and phosphatidylinositol 4-kinase (PI-4-K), which, conversely, increases PIP 2 . How membrane PIP 2 levels are regulated by CaR activation and whether these changes modulate inward rectifier K + are unknown. In this study, we found that activation of CaR by the allosteric agonist, NPSR568, increased inward rectifier K + current (I K1 ) in guinea pig ventricular myocytes and currents mediated by Kir2.1 channels exogenously expressed in HEK293T cells with a similar sensitivity. Moreover, using the fluorescent PIP 2 reporter tubby-R332H-cYFP to monitor PIP 2 levels, we found that CaR activation in HEK293T cells increased membrane PIP 2 concentrations. Pharmacological studies showed that both phospholipase C (PLC) and PI-4-K are activated by CaR stimulation with the latter played a dominant role in regulating membrane PIP 2 and, thus, Kir currents. These results provide the first direct evidence that CaR activation upregulates currents through inward rectifier K + channels by accelerating PIP 2 synthesis. The regulation of I K1 plays a critical role in the stability of the electrical properties of many excitable cells, including cardiac myocytes and neurons. Further, synthetic allosteric modulators that increase CaR activity have been used to treat hyperparathyroidism, and negative CaR modulators are of potential importance in the treatment of osteoporosis. Thus, our results provide further insight into the roles played by CaR in the cardiovascular system and are potentially valuable for heart disease treatment and drug safety.

  14. Inhibition of G protein-activated inwardly rectifying K+ channels by different classes of antidepressants.

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    Toru Kobayashi

    Full Text Available Various antidepressants are commonly used for the treatment of depression and several other neuropsychiatric disorders. In addition to their primary effects on serotonergic or noradrenergic neurotransmitter systems, antidepressants have been shown to interact with several receptors and ion channels. However, the molecular mechanisms that underlie the effects of antidepressants have not yet been sufficiently clarified. G protein-activated inwardly rectifying K(+ (GIRK, Kir3 channels play an important role in regulating neuronal excitability and heart rate, and GIRK channel modulation has been suggested to have therapeutic potential for several neuropsychiatric disorders and cardiac arrhythmias. In the present study, we investigated the effects of various classes of antidepressants on GIRK channels using the Xenopus oocyte expression assay. In oocytes injected with mRNA for GIRK1/GIRK2 or GIRK1/GIRK4 subunits, extracellular application of sertraline, duloxetine, and amoxapine effectively reduced GIRK currents, whereas nefazodone, venlafaxine, mianserin, and mirtazapine weakly inhibited GIRK currents even at toxic levels. The inhibitory effects were concentration-dependent, with various degrees of potency and effectiveness. Furthermore, the effects of sertraline were voltage-independent and time-independent during each voltage pulse, whereas the effects of duloxetine were voltage-dependent with weaker inhibition with negative membrane potentials and time-dependent with a gradual decrease in each voltage pulse. However, Kir2.1 channels were insensitive to all of the drugs. Moreover, the GIRK currents induced by ethanol were inhibited by sertraline but not by intracellularly applied sertraline. The present results suggest that GIRK channel inhibition may reveal a novel characteristic of the commonly used antidepressants, particularly sertraline, and contributes to some of the therapeutic effects and adverse effects.

  15. Inward-rectifying potassium (Kir) channels regulate pacemaker activity in spinal nociceptive circuits during early life

    Science.gov (United States)

    Li, Jie; Blankenship, Meredith L.; Baccei, Mark L.

    2013-01-01

    Pacemaker neurons in neonatal spinal nociceptive circuits generate intrinsic burst-firing and are distinguished by a lower “leak” membrane conductance compared to adjacent, non-bursting neurons. However, little is known about which subtypes of leak channels regulate the level of pacemaker activity within the developing rat superficial dorsal horn (SDH). Here we demonstrate that a hallmark feature of lamina I pacemaker neurons is a reduced conductance through inward-rectifying potassium (Kir) channels at physiological membrane potentials. Differences in the strength of inward rectification between pacemakers and non-pacemakers indicate the presence of functionally distinct Kir currents in these two populations at room temperature. However, Kir currents in both groups showed high sensitivity to block by extracellular Ba2+ (IC50 ~ 10 µM), which suggests the presence of ‘classical’ Kir (Kir2.x) channels in the neonatal SDH. The reduced Kir conductance within pacemakers is unlikely to be explained by an absence of particular Kir2.x isoforms, as immunohistochemical analysis revealed the expression of Kir2.1, Kir2.2 and Kir2.3 within spontaneously bursting neurons. Importantly, Ba2+ application unmasked rhythmic burst-firing in ~42% of non-bursting lamina I neurons, suggesting that pacemaker activity is a latent property of a sizeable population of SDH cells during early life. In addition, the prevalence of spontaneous burst-firing within lamina I was enhanced in the presence of high internal concentrations of free Mg2+, consistent with its documented ability to block Kir channels from the intracellular side. Collectively, the results indicate that Kir channels are key modulators of pacemaker activity in newborn central pain networks. PMID:23426663

  16. Effect of ethanol at clinically relevant concentrations on atrial inward rectifier potassium current sensitive to acetylcholine.

    Science.gov (United States)

    Bébarová, Markéta; Matejovič, Peter; Pásek, Michal; Hořáková, Zuzana; Hošek, Jan; Šimurdová, Milena; Šimurda, Jiří

    2016-10-01

    Alcohol intoxication tends to induce arrhythmias, most often the atrial fibrillation. To elucidate arrhythmogenic mechanisms related to alcohol consumption, the effect of ethanol on main components of the ionic membrane current is investigated step by step. Considering limited knowledge, we aimed to examine the effect of clinically relevant concentrations of ethanol (0.8-80 mM) on acetylcholine-sensitive inward rectifier potassium current I K(Ach). Experiments were performed by the whole-cell patch clamp technique at 23 ± 1 °C on isolated rat and guinea-pig atrial myocytes, and on expressed human Kir3.1/3.4 channels. Ethanol induced changes of I K(Ach) in the whole range of concentrations applied; the effect was not voltage dependent. The constitutively active component of I K(Ach) was significantly increased by ethanol with the maximum effect (an increase by ∼100 %) between 8 and 20 mM. The changes were comparable in rat and guinea-pig atrial myocytes and also in expressed human Kir3.1/3.4 channels (i.e., structural correlate of I K(Ach)). In the case of the acetylcholine-induced component of I K(Ach), a dual ethanol effect was apparent with a striking heterogeneity of changes in individual cells. The effect correlated with the current magnitude in control: the current was increased by eth-anol in the cells showing small current in control and vice versa. The average effect peaked at 20 mM ethanol (an increase of the current by ∼20 %). Observed changes of action potential duration agreed well with the voltage clamp data. Ethanol significantly affected both components of I K(Ach) even in concentrations corresponding to light alcohol consumption.

  17. Structural basis of drugs that increase cardiac inward rectifier Kir2.1 currents.

    Science.gov (United States)

    Gómez, Ricardo; Caballero, Ricardo; Barana, Adriana; Amorós, Irene; De Palm, Sue-Haida; Matamoros, Marcos; Núñez, Mercedes; Pérez-Hernández, Marta; Iriepa, Isabel; Tamargo, Juan; Delpón, Eva

    2014-11-01

    We hypothesize that some drugs, besides flecainide, increase the inward rectifier current (IK1) generated by Kir2.1 homotetramers (IKir2.1) and thus, exhibit pro- and/or antiarrhythmic effects particularly at the ventricular level. To test this hypothesis, we analysed the effects of propafenone, atenolol, dronedarone, and timolol on Kir2.x channels. Currents were recorded with the patch-clamp technique using whole-cell, inside-out, and cell-attached configurations. Propafenone (0.1 nM-1 µM) did not modify either IK1 recorded in human right atrial myocytes or the current generated by homo- or heterotetramers of Kir2.2 and 2.3 channels recorded in transiently transfected Chinese hamster ovary cells. On the other hand, propafenone increased IKir2.1 (EC50 = 12.0 ± 3.0 nM) as a consequence of its interaction with Cys311, an effect which decreased inward rectification of the current. Propafenone significantly increased mean open time and opening frequency at all the voltages tested, resulting in a significant increase of the mean open probability of the channel. Timolol, which interacted with Cys311, was also able to increase IKir2.1. On the contrary, neither atenolol nor dronedarone modified IKir2.1. Molecular modelling of the Kir2.1-drugs interaction allowed identification of the pharmacophore of drugs that increase IKir2.1. Kir2.1 channels exhibit a binding site determined by Cys311 that is responsible for drug-induced IKir2.1 increase. Drug binding decreases channel affinity for polyamines and current rectification, and can be a mechanism of drug-induced pro- and antiarrhythmic effects not considered until now. Published on behalf of the European Society of Cardiology. All rights reserved. © The Author 2014. For permissions please email: journals.permissions@oup.com.

  18. Nicotine at clinically relevant concentrations affects atrial inward rectifier potassium current sensitive to acetylcholine.

    Science.gov (United States)

    Bébarová, Markéta; Matejovič, Peter; Švecová, Olga; Kula, Roman; Šimurdová, Milena; Šimurda, Jiří

    2017-05-01

    Nicotine abuse is associated with variety of diseases including arrhythmias, most often atrial fibrillation (AF). Altered inward rectifier potassium currents including acetylcholine-sensitive current I K(Ach) are known to be related to AF pathogenesis. Since relevant data are missing, we aimed to investigate I K(Ach) changes at clinically relevant concentrations of nicotine. Experiments were performed by the whole cell patch clamp technique at 23 ± 1 °C on isolated rat atrial myocytes. Nicotine was applied at following concentrations: 4, 40 and 400 nM; ethanol at 20 mM (∼0.09%). Nicotine at 40 and 400 nM significantly activated constitutively active component of I K(Ach) with the maximum effect at 40 nM (an increase by ∼100%); similar effect was observed at -110 and -50 mV. Changes at 4 nM nicotine were negligible on average. Coapplication of 40 nM nicotine and 20 mM ethanol (which is also known to activate this current) did not show cumulative effect. In the case of acetylcholine-induced component of I K(Ach) , a dual effect of nicotine and its correlation with the current magnitude in control were apparent: the current was increased by nicotine in the cells showing small current in control and vice versa. The effect of 40 and 400 nM nicotine on acetylcholine-induced component of I K(Ach) was significantly different at -110 and -50 mV. We conclude that nicotine at clinically relevant concentrations significantly increased constitutively active component of I K(Ach) and showed a dual effect on its acetylcholine-induced component, similarly as ethanol. Synchronous application of nicotine and ethanol did not cause additive effect.

  19. EFFECT OF ETHANOL AND ACETALDEHYDE AT CLINICALLY RELEVANT CONCENTRATIONS ON ATRIAL INWARD RECTIFIER POTASSIUM CURRENT I-K1 SEPARATE AND COMBINED EFFECT

    Czech Academy of Sciences Publication Activity Database

    Hořáková, Z.; Matejovič, P.; Pásek, Michal; Hošek, J.; Šimurdová, M.; Šimurda, J.; Bébarová, M.

    2016-01-01

    Roč. 67, č. 3 (2016), s. 339-351 ISSN 0867-5910 Institutional support: RVO:61388998 Keywords : arrhythmias * cardiomyocyte * inward rectifier potassium current * ethanol * mathematical model Subject RIV: BO - Biophysics Impact factor: 2.883, year: 2016

  20. Inward rectifier potassium current (I K1) and Kir2 composition of the zebrafish (Danio rerio) heart.

    Science.gov (United States)

    Hassinen, Minna; Haverinen, Jaakko; Hardy, Matt E; Shiels, Holly A; Vornanen, Matti

    2015-12-01

    Electrophysiological properties and molecular background of the zebrafish (Danio rerio) cardiac inward rectifier current (IK1) were examined. Ventricular myocytes of zebrafish have a robust (-6.7 ± 1.2 pA pF(-1) at -120 mV) strongly rectifying and Ba(2+)-sensitive (IC50 = 3.8 μM) IK1. Transcripts of six Kir2 channels (drKir2.1a, drKir2.1b, drKir2.2a, drKir2.2b, drKir2.3, and drKir2.4) were expressed in the zebrafish heart. drKir2.4 and drKir2.2a were the dominant isoforms in both the ventricle (92.9 ± 1.5 and 6.3 ± 1.5%) and the atrium (28.9 ± 2.9 and 64.7 ± 3.0%). The remaining four channels comprised together less than 1 and 7 % of the total transcripts in ventricle and atrium, respectively. The four main gene products (drKir2.1a, drKir2.2a, drKir2.2b, drKir2.4) were cloned, sequenced, and expressed in HEK cells for electrophysiological characterization. drKir2.1a was the most weakly rectifying (passed more outward current) and drKir2.2b the most strongly rectifying (passed less outward current) channel, whilst drKir2.2a and drKir2.4 were intermediate between the two. In regard to sensitivity to Ba(2+) block, drKir2.4 was the most sensitive (IC50 = 1.8 μM) and drKir2.1a the least sensitive channel (IC50 = 132 μM). These findings indicate that the Kir2 isoform composition of the zebrafish heart markedly differs from that of mammalian hearts. Furthermore orthologous Kir2 channels (Kir2.1 and Kir2.4) of zebrafish and mammals show striking differences in Ba(2+)-sensitivity. Structural and functional differences needs to be taken into account when zebrafish is used as a model for human cardiac electrophysiology, cardiac diseases, and in screening cardioactive substances.

  1. Opening of the inward rectifier potassium channel alleviates maladaptive tissue repair following myocardial infarction.

    Science.gov (United States)

    Liu, Chengfang; Liu, Enli; Luo, Tiane; Zhang, Weifang; He, Rongli

    2016-08-01

    Activation of the inward rectifier potassium current (IK1) channel has been reported to be associated with suppression of ventricular arrhythmias. In this study, we tested the hypothesis that opening of the IK1 channel with zacopride (ZAC) was involved in the modulation of tissue repair after myocardial infarction. Sprague-Dawley rats were subject to coronary artery ligation and ZAC was administered intraperitoneally (15 µg/kg/day) for 28 days. Compared with the ischemia group, treatment with ZAC significantly reduced the ratio of heart/body weight and the cross-sectional area of cardiomyocytes, suggesting less cardiac hypertrophy. ZAC reduced the accumulation of collagen types I and III, accompanied with decrease of collagen area, which were associated with a reduction of collagen deposition in the fibrotic myocardium. Echocardiography showed improved cardiac function, evidenced by the reduced left ventricular end-diastolic dimension and left ventricular end-systolic dimension, and the increased ejection fraction and fractional shortening in ZAC-treated animals (all P < 0.05 vs. ischemia group). In coincidence with these changes, ZAC up-regulated the protein level of the IK1 channel and down-regulated the phosphorylation of mammalian target of rapamycin (mTOR) and 70-kDa ribosomal protein S6 (p70S6) kinase. Administration of chloroquine alone, an IK1 channel antagonist, had no effect on all the parameters measured, but significantly blocked the beneficial effects of ZAC on cardiac repair. In conclusion, opening of the IK1 channel with ZAC inhibits maladaptive tissue repair and improves cardiac function, potentially mediated by the inhibition of ischemia-activated mTOR-p70S6 signaling pathway via the IK1 channel. So the development of pharmacological agents specifically targeting the activation of the IK1 channel may protect the heart against myocardial ischemia-induced cardiac dysfunction. © The Author 2016. Published by Oxford University Press on behalf of

  2. Expression of G-protein inwardly rectifying potassium channels (GIRKs in lung cancer cell lines

    Directory of Open Access Journals (Sweden)

    Schuller Hildegard M

    2005-08-01

    Full Text Available Abstract Background Previous data from our laboratory has indicated that there is a functional link between the β-adrenergic receptor signaling pathway and the G-protein inwardly rectifying potassium channel (GIRK1 in human breast cancer cell lines. We wanted to determine if GIRK channels were expressed in lung cancers and if a similar link exists in lung cancer. Methods GIRK1-4 expression and levels were determined by reverse transcription polymerase chain reaction (RT-PCR and real-time PCR. GIRK protein levels were determined by western blots and cell proliferation was determined by a 5-bromo-2'-deoxyuridine (BrdU assay. Results GIRK1 mRNA was expressed in three of six small cell lung cancer (SCLC cell lines, and either GIRK2, 3 or 4 mRNA expression was detected in all six SCLC cell lines. Treatment of NCI-H69 with β2-adrenergic antagonist ICI 118,551 (100 μM daily for seven days led to slight decreases of GIRK1 mRNA expression levels. Treatment of NCI-H69 with the β-adrenergic agonist isoproterenol (10 μM decreased growth rates in these cells. The GIRK inhibitor U50488H (2 μM also inhibited proliferation, and this decrease was potentiated by isoproterenol. In the SCLC cell lines that demonstrated GIRK1 mRNA expression, we also saw GIRK1 protein expression. We feel these may be important regulatory pathways since no expression of mRNA of the GIRK channels (1 & 2 was found in hamster pulmonary neuroendocrine cells, a suggested cell of origin for SCLC, nor was GIRK1 or 2 expression found in human small airway epithelial cells. GIRK (1,2,3,4 mRNA expression was also seen in A549 adenocarcinoma and NCI-H727 carcinoid cell lines. GIRK1 mRNA expression was not found in tissue samples from adenocarcinoma or squamous cancer patients, nor was it found in NCI-H322 or NCI-H441 adenocarcinoma cell lines. GIRK (1,3,4 mRNA expression was seen in three squamous cell lines, GIRK2 was only expressed in one squamous cell line. However, GIRK1 protein

  3. Activation of the Ca2+-sensing receptors increases currents through inward rectifier K+ channels via activation of phosphatidylinositol 4-kinase

    OpenAIRE

    Liu, Chung-Hung; Chang, Hsueh-Kai; Lee, Sue-Ping; Shieh, Ru-Chi

    2016-01-01

    Inward rectifier K+ channels are important for maintaining normal electrical function in many cell types. The proper function of these channels requires the presence of membrane phosphoinositide 4,5-bisphosphate (PIP2). Stimulation of the Ca2+-sensing receptor CaR, a pleiotropic G protein-coupled receptor, activates both Gq/11, which decreases PIP2, and phosphatidylinositol 4-kinase (PI-4-K), which, conversely, increases PIP2. How membrane PIP2 levels are regulated by CaR activation and wheth...

  4. Inhibition by acrolein of light-induced stomatal opening through inhibition of inward-rectifying potassium channels in Arabidopsis thaliana.

    Science.gov (United States)

    Islam, Md Moshiul; Ye, Wenxiu; Matsushima, Daiki; Khokon, Md Atiqur Rahman; Munemasa, Shintaro; Nakamura, Yoshimasa; Murata, Yoshiyuki

    2015-01-01

    Acrolein is a reactive α,β-unsaturated aldehyde derived from lipid peroxides, which are produced in plants under a variety of stress. We investigated effects of acrolein on light-induced stomatal opening using Arabidopsis thaliana. Acrolein inhibited light-induced stomatal opening in a dose-dependent manner. Acrolein at 100 μM inhibited plasma membrane inward-rectifying potassium (Kin) channels in guard cells. Acrolein at 100 μM inhibited Kin channel KAT1 expressed in a heterologous system using Xenopus leaves oocytes. These results suggest that acrolein inhibits light-induced stomatal opening through inhibition of Kin channels in guard cells.

  5. Strong eukaryotic IRESs have weak secondary structure.

    Directory of Open Access Journals (Sweden)

    Xuhua Xia

    Full Text Available BACKGROUND: The objective of this work was to investigate the hypothesis that eukaryotic Internal Ribosome Entry Sites (IRES lack secondary structure and to examine the generality of the hypothesis. METHODOLOGY/PRINCIPAL FINDINGS: IRESs of the yeast and the fruit fly are located in the 5'UTR immediately upstream of the initiation codon. The minimum folding energy (MFE of 60 nt RNA segments immediately upstream of the initiation codons was calculated as a proxy of secondary structure stability. MFE of the reverse complements of these 60 nt segments was also calculated. The relationship between MFE and empirically determined IRES activity was investigated to test the hypothesis that strong IRES activity is associated with weak secondary structure. We show that IRES activity in the yeast and the fruit fly correlates strongly with the structural stability, with highest IRES activity found in RNA segments that exhibit the weakest secondary structure. CONCLUSIONS: We found that a subset of eukaryotic IRESs exhibits very low secondary structure in the 5'-UTR sequences immediately upstream of the initiation codon. The consistency in results between the yeast and the fruit fly suggests a possible shared mechanism of cap-independent translation initiation that relies on an unstructured RNA segment.

  6. Short- and long-term inhibition of cardiac inward-rectifier potassium channel current by an antiarrhythmic drug bepridil.

    Science.gov (United States)

    Ma, Fangfang; Takanari, Hiroki; Masuda, Kimiko; Morishima, Masaki; Ono, Katsushige

    2016-07-01

    Bepridil is an effective antiarrhythmic drug on supraventricular and ventricular arrhythmias, and inhibitor of calmodulin. Recent investigations have been elucidating that bepridil exerts antiarrhythmic effects through its acute and chronic application for patients. The aim of this study was to identify the efficacy and the potential mechanism of bepridil on the inward-rectifier potassium channel in neonatal rat cardiomyocytes in acute- and long-term conditions. Bepridil inhibited inward-rectifier potassium current (I K1) as a short-term effect with IC50 of 17 μM. Bepridil also reduced I K1 of neonatal cardiomyocytes when applied for 24 h in the culture medium with IC50 of 2.7 μM. Both a calmodulin inhibitor (W-7) and an inhibitor of calmodulin-kinase II (KN93) reduced I K1 when applied for 24 h as a long-term effect in the same fashion, suggesting that the long-term application of bepridil inhibits I K1 more potently than that of the short-term application through the inhibition of calmodulin kinase II pathway in cardiomyocytes.

  7. Diadenosine pentaphosphate affects electrical activity in guinea pig atrium via activation of potassium acetylcholine-dependent inward rectifier.

    Science.gov (United States)

    Abramochkin, Denis V; Karimova, Viktoria M; Filatova, Tatiana S; Kamkin, Andre

    2017-07-01

    Diadenosine pentaphosphate (Ap5A) belongs to the family of diadenosine polyphosphates, endogenously produced compounds that affect vascular tone and cardiac performance when released from platelets. The previous findings indicate that Ap5A shortens action potentials (APs) in rat myocardium via activation of purine P2 receptors. The present study demonstrates alternative mechanism of Ap5A electrophysiological effects found in guinea pig myocardium. Ap5A (10 -4  M) shortens APs in guinea pig working atrial myocardium and slows down pacemaker activity in the sinoatrial node. P1 receptors antagonist DPCPX (10 -7  M) or selective GIRK channels blocker tertiapin (10 -6  M) completely abolished all Ap5A effects, while P2 blocker PPADS (10 -4  M) was ineffective. Patch-clamp experiments revealed potassium inward rectifier current activated by Ap5A in guinea pig atrial myocytes. The current was abolished by DPCPX or tertiapin and therefore was considered as potassium acetylcholine-dependent inward rectifier (I KACh ). Thus, unlike rat, in guinea pig atrium Ap5A produces activation of P1 receptors and subsequent opening of KACh channels leading to negative effects on cardiac electrical activity.

  8. Isolation of proflavine as a blocker of G protein-gated inward rectifier potassium channels by a cell growth-based screening system.

    Science.gov (United States)

    Kawada, Hitoshi; Inanobe, Atsushi; Kurachi, Yoshihisa

    2016-10-01

    The overexpression of Kir3.2, a subunit of the G protein-gated inwardly rectifying K(+) channel, is implicated in some of the neurological phenotypes of Down syndrome (DS). Chemical compounds that block Kir3.2 are expected to improve the symptoms of DS. The purpose of this study is to develop a cell-based screening system to identify Kir3.2 blockers and then investigate the mode of action of the blocker. Chemical screening was carried out using a K(+) transporter-deficient yeast strain that expressed a constitutively active Kir3.2 mutant. The mode of action of an effective blocker was electrophysiologically analyzed using Kir channels expressed in Xenopus oocytes. Proflavine was identified to inhibit the growth of Kir3.2-transformant cells and Kir3.2 activity in a concentration-dependent manner. The current inhibition was strong when membrane potentials (Vm) was above equilibrium potential of K(+) (EK). When Vm was below EK, the blockage apparently depended on the difference between Vm and [K(+)]. Furthermore, the inhibition became stronger by lowering extracellular [K(+)]. These results indicated that the yeast strain serves as a screening system to isolate Kir3.2 blockers and proflavine is a prototype of a pore blocker of Kir3.2. Copyright © 2016 Elsevier Ltd. All rights reserved.

  9. The function and molecular identity of inward rectifier channels in vestibular hair cells of the mouse inner ear

    Science.gov (United States)

    Levin, Michaela E.

    2012-01-01

    Inner ear hair cells respond to mechanical stimuli with graded receptor potentials. These graded responses are modulated by a host of voltage-dependent currents that flow across the basolateral membrane. Here, we examine the molecular identity and the function of a class of voltage-dependent ion channels that carries the potassium-selective inward rectifier current known as IK1. IK1 has been identified in vestibular hair cells of various species, but its molecular composition and functional contributions remain obscure. We used quantitative RT-PCR to show that the inward rectifier gene, Kir2.1, is highly expressed in mouse utricle between embryonic day 15 and adulthood. We confirmed Kir2.1 protein expression in hair cells by immunolocalization. To examine the molecular composition of IK1, we recorded voltage-dependent currents from type II hair cells in response to 50-ms steps from −124 to −54 in 10-mV increments. Wild-type cells had rapidly activating inward currents with reversal potentials close to the K+ equilibrium potential and a whole-cell conductance of 4.8 ± 1.5 nS (n = 46). In utricle hair cells from Kir2.1-deficient (Kir2.1−/−) mice, IK1 was absent at all stages examined. To identify the functional contribution of Kir2.1, we recorded membrane responses in current-clamp mode. Hair cells from Kir2.1−/− mice had significantly (P < 0.001) more depolarized resting potentials and larger, slower membrane responses than those of wild-type cells. These data suggest that Kir2.1 is required for IK1 in type II utricle hair cells and contributes to hyperpolarized resting potentials and fast, small amplitude receptor potentials in response to current inputs, such as those evoked by hair bundle deflections. PMID:22496522

  10. N-(2-methoxyphenyl) benzenesulfonamide, a novel regulator of neuronal G protein-gated inward rectifier K+ channels.

    Science.gov (United States)

    Walsh, Kenneth B; Gay, Elaine A; Blough, Bruce E; Geurkink, David W

    2017-11-15

    G protein-gated inward rectifier K + (GIRK) channels are members of the super-family of proteins known as inward rectifier K + (Kir) channels and are expressed throughout the peripheral and central nervous systems. Neuronal GIRK channels are the downstream targets of a number of neuromodulators including opioids, somatostatin, dopamine and cannabinoids. Previous studies have demonstrated that the ATP-sensitive K + channel, another member of the Kir channel family, is regulated by sulfonamide drugs. Therefore, to determine if sulfonamides also modulate GIRK channels, we screened a library of arylsulfonamide compounds using a GIRK channel fluorescent assay that utilized pituitary AtT20 cells expressing GIRK channels along with the somatostatin type-2 and -5 receptors. Enhancement of the GIRK channel fluorescent signal by one compound, N-(2-methoxyphenyl) benzenesulfonamide (MPBS), was dependent on the activation of the channel by somatostatin. In whole-cell patch clamp experiments, application of MPBS both shifted the somatostatin concentration-response curve (EC 50 = 3.5nM [control] vs.1.0nM [MPBS]) for GIRK channel activation and increased the maximum GIRK current measured with 100nM somatostatin. However, GIRK channel activation was not observed when MPBS was applied to the cells in the absence of somatostatin. While the MPBS structural analog 4-fluoro-N-(2-methoxyphenyl) benzenesulfonamide also augmented the somatostatin-induced GIRK fluorescent signal, no increase in the signal was observed with the sulfonamides tolbutamide, sulfapyridine and celecoxib. In conclusion, MPBS represents a novel prototypic GPCR-dependent regulator of neuronal GIRK channels. Copyright © 2017 Elsevier B.V. All rights reserved.

  11. LRET Determination of Molecular Distances during pH Gating of the Mammalian Inward Rectifier Kir1.1b.

    Science.gov (United States)

    Nanazashvili, Mikheil; Sánchez-Rodríguez, Jorge E; Fosque, Ben; Bezanilla, Francisco; Sackin, Henry

    2018-01-09

    Gating of the mammalian inward rectifier Kir1.1 at the helix bundle crossing (HBC) by intracellular pH is believed to be mediated by conformational changes in the C-terminal domain (CTD). However, the exact motion of the CTD during Kir gating remains controversial. Crystal structures and single-molecule fluorescence resonance energy transfer of KirBac channels have implied a rigid body rotation and/or a contraction of the CTD as possible triggers for opening of the HBC gate. In our study, we used lanthanide-based resonance energy transfer on single-Cys dimeric constructs of the mammalian renal inward rectifier, Kir1.1b, incorporated into anionic liposomes plus PIP 2 , to determine unambiguous, state-dependent distances between paired Cys residues on diagonally opposite subunits. Functionality and pH dependence of our proteoliposome channels were verified in separate electrophysiological experiments. The lanthanide-based resonance energy transfer distances measured in closed (pH 6) and open (pH 8) conditions indicated neither expansion nor contraction of the CTD during gating, whereas the HBC gate widened by 8.8 ± 4 Å, from 6.3 ± 2 to 15.1 ± 6 Å, during opening. These results are consistent with a Kir gating model in which rigid body rotation of the large CTD around the permeation axis is correlated with opening of the HBC hydrophobic gate, allowing permeation of a 7 Å hydrated K ion. Copyright © 2017 Biophysical Society. Published by Elsevier Inc. All rights reserved.

  12. The inward rectifier current inhibitor PA-6 terminates atrial fibrillation and does not cause ventricular arrhythmias in goat and dog models

    NARCIS (Netherlands)

    Ji, Yuan; Varkevisser, Rosanne; Opacic, Dragan; Bossu, Alexandre; Kuiper, Marion; Beekman, Jet D.M.; Yang, Sihyung; Khan, Azinwi Phina; Dobrev, Dobromir; Voigt, Niels; Wang, Michael Zhuo; Verheule, Sander; Vos, Marc A.; van der Heyden, Marcel A.G.

    2017-01-01

    Background and Purpose: The density of the inward rectifier current (IK1) increases in atrial fibrillation (AF), shortening effective refractory period and thus promoting atrial re-entry. The synthetic compound pentamidine analogue 6 (PA-6) is a selective and potent IK1 inhibitor. We tested PA-6 for

  13. Left-to-Right Atrial Inward Rectifier Potassium Current Gradients in Patients With Paroxysmal Versus Chronic Atrial Fibrillation

    Science.gov (United States)

    Voigt, Niels; Trausch, Anne; Knaut, Michael; Matschke, Klaus; Varró, András; Van Wagoner, David R.; Nattel, Stanley; Ravens, Ursula; Dobrev, Dobromir

    2018-01-01

    Background Recent evidence suggests that atrial fibrillation (AF) is maintained by high-frequency reentrant sources with a left-to-right–dominant frequency gradient, particularly in patients with paroxysmal AF (pAF). Unequal left-to-right distribution of inward rectifier K+ currents has been suggested to underlie this dominant frequency gradient, but this hypothesis has never been tested in humans. Methods and Results Currents were measured with whole-cell voltage-clamp in cardiomyocytes from right atrial (RA) and left (LA) atrial appendages of patients in sinus rhythm (SR) and patients with AF undergoing cardiac surgery. Western blot was used to quantify protein expression of IK1 (Kir2.1 and Kir2.3) and IK,ACh (Kir3.1 and Kir3.4) subunits. Basal current was ≈2-fold larger in chronic AF (cAF) versus SR patients, without RA-LA differences. In pAF, basal current was ≈2-fold larger in LA versus RA, indicating a left-to-right atrial gradient. In both atria, Kir2.1 expression was ≈2-fold greater in cAF but comparable in pAF versus SR. Kir2.3 levels were unchanged in cAF and RA-pAF but showed a 51% decrease in LA-pAF. In SR, carbachol-activated (2 μmol/L) IK,ACh was 70% larger in RA versus LA. This right-to-left atrial gradient was decreased in pAF and cAF caused by reduced IK,ACh in RA only. Similarly, in SR, Kir3.1 and Kir3.4 proteins were greater in RA versus LA and decreased in RA of pAF and cAF. Kir3.1 and Kir3.4 expression was unchanged in LA of pAF and cAF. Conclusions Our results support the hypothesis that a left-to-right gradient in inward rectifier background current contributes to high-frequency sources in LA that maintain pAF. These findings have potentially important implications for development of atrial-selective therapeutic approaches. PMID:20657029

  14. Inverse agonist-like action of cadmium on G-protein-gated inward-rectifier K+ channels

    International Nuclear Information System (INIS)

    Inanobe, Atsushi; Matsuura, Takanori; Nakagawa, Atsushi; Kurachi, Yoshihisa

    2011-01-01

    Highlights: → We examined allosteric control of K + channel gating. → We identified a high-affinity site for Cd 2+ to inhibit Kir3.2 activity. → The 6-coordination geometry supports the binding. → Cd 2+ inhibits Kir3.2 by trapping the conformation in the closed state. -- Abstract: The gate at the pore-forming domain of potassium channels is allosterically controlled by a stimulus-sensing domain. Using Cd 2+ as a probe, we examined the structural elements responsible for gating in an inward-rectifier K + channel (Kir3.2). One of four endogenous cysteines facing the cytoplasm contributes to a high-affinity site for inhibition by internal Cd 2+ . Crystal structure of its cytoplasmic domain in complex with Cd 2+ reveals that octahedral coordination geometry supports the high-affinity binding. This mode of action causes the tethering of the N-terminus to CD loop in the stimulus-sensing domain, suggesting that their conformational changes participate in gating and Cd 2+ inhibits Kir3.2 by trapping the conformation in the closed state like 'inverse agonist'.

  15. Inwardly Rectifying Potassium (Kir) Channels Represent a Critical Ion Conductance Pathway in the Nervous Systems of Insects.

    Science.gov (United States)

    Chen, Rui; Swale, Daniel R

    2018-01-25

    A complete understanding of the physiological pathways critical for proper function of the insect nervous system is still lacking. The recent development of potent and selective small-molecule modulators of insect inward rectifier potassium (Kir) channels has enabled the interrogation of the physiological role and toxicological potential of Kir channels within various insect tissue systems. Therefore, we aimed to highlight the physiological and functional role of neural Kir channels the central nervous system, muscular system, and neuromuscular system through pharmacological and genetic manipulations. Our data provide significant evidence that Drosophila neural systems rely on the inward conductance of K + ions for proper function since pharmacological inhibition and genetic ablation of neural Kir channels yielded dramatic alterations of the CNS spike discharge frequency and broadening and reduced amplitude of the evoked EPSP at the neuromuscular junction. Based on these data, we conclude that neural Kir channels in insects (1) are critical for proper function of the insect nervous system, (2) represents an unexplored physiological pathway that is likely to shape the understanding of neuronal signaling, maintenance of membrane potentials, and maintenance of the ionic balance of insects, and (3) are capable of inducing acute toxicity to insects through neurological poisoning.

  16. Synergistic activation of G protein-gated inwardly rectifying potassium channels by cholesterol and PI(4,5)P2.

    Science.gov (United States)

    Bukiya, Anna N; Rosenhouse-Dantsker, Avia

    2017-07-01

    G-protein gated inwardly rectifying potassium (GIRK or Kir3) channels play a major role in the control of the heart rate, and require the membrane phospholipid phosphatidylinositol-bis-phosphate (PI(4,5)P 2 ) for activation. Recently, we have shown that the activity of the heterotetrameric Kir3.1/Kir3.4 channel that underlies atrial K ACh currents was enhanced by cholesterol. Similarly, the activities of both the Kir3.4 homomer and its active pore mutant Kir3.4* (Kir3.4_S143T) were also enhanced by cholesterol. Here we employ planar lipid bilayers to investigate the crosstalk between PI(4,5)P 2 and cholesterol, and demonstrate that these two lipids act synergistically to activate Kir3.4* currents. Further studies using the Xenopus oocytes heterologous expression system suggest that PI(4,5)P 2 and cholesterol act via distinct binding sites. Whereas PI(4,5)P 2 binds to the cytosolic domain of the channel, the putative binding region of cholesterol is located at the center of the transmembrane domain overlapping the central glycine hinge region of the channel. Together, our data suggest that changes in the levels of two key membrane lipids - cholesterol and PI(4,5)P 2 - could act in concert to provide fine-tuning of Kir3 channel function. Copyright © 2017 Elsevier B.V. All rights reserved.

  17. Molecular and functional characterization of Anopheles gambiae inward rectifier potassium (Kir1) channels: a novel role in egg production.

    Science.gov (United States)

    Raphemot, Rene; Estévez-Lao, Tania Y; Rouhier, Matthew F; Piermarini, Peter M; Denton, Jerod S; Hillyer, Julián F

    2014-08-01

    Inward rectifier potassium (Kir) channels play essential roles in regulating diverse physiological processes. Although Kir channels are encoded in mosquito genomes, their functions remain largely unknown. In this study, we identified the members of the Anopheles gambiae Kir gene family and began to investigate their function. Notably, we sequenced the A. gambiae Kir1 (AgKir1) gene and showed that it encodes all the canonical features of a Kir channel: an ion pore that is composed of a pore helix and a selectivity filter, two transmembrane domains that flank the ion pore, and the so-called G-loop. Heterologous expression of AgKir1 in Xenopus oocytes revealed that this gene encodes a functional, barium-sensitive Kir channel. Quantitative RT-PCR experiments then showed that relative AgKir1 mRNA levels are highest in the pupal stage, and that AgKir1 mRNA is enriched in the adult ovaries. Gene silencing of AgKir1 by RNA interference did not affect the survival of female mosquitoes following a blood meal, but decreased their egg output. These data provide evidence for a new role of Kir channels in mosquito fecundity, and further validates them as promising molecular targets for the development of a new class of mosquitocides to be used in vector control. Copyright © 2014 Elsevier Ltd. All rights reserved.

  18. Inverse agonist-like action of cadmium on G-protein-gated inward-rectifier K{sup +} channels

    Energy Technology Data Exchange (ETDEWEB)

    Inanobe, Atsushi, E-mail: inanobe@pharma2.med.osaka-u.ac.jp [Department of Pharmacology, Graduate School of Medicine, Osaka University, Osaka (Japan); Center for Advanced Medical Engineering and Informatics, Osaka University, Osaka (Japan); Matsuura, Takanori [Laboratory of Protein Informatics, Institute for Protein Research, Osaka University, Osaka (Japan); Nakagawa, Atsushi [Laboratory of Supramolecular Crystallography, Institute for Protein Research, Osaka University, Osaka (Japan); Kurachi, Yoshihisa, E-mail: ykurachi@pharma2.med.osaka-u.ac.jp [Department of Pharmacology, Graduate School of Medicine, Osaka University, Osaka (Japan); Center for Advanced Medical Engineering and Informatics, Osaka University, Osaka (Japan)

    2011-04-08

    Highlights: {yields} We examined allosteric control of K{sup +} channel gating. {yields} We identified a high-affinity site for Cd{sup 2+} to inhibit Kir3.2 activity. {yields} The 6-coordination geometry supports the binding. {yields} Cd{sup 2+} inhibits Kir3.2 by trapping the conformation in the closed state. -- Abstract: The gate at the pore-forming domain of potassium channels is allosterically controlled by a stimulus-sensing domain. Using Cd{sup 2+} as a probe, we examined the structural elements responsible for gating in an inward-rectifier K{sup +} channel (Kir3.2). One of four endogenous cysteines facing the cytoplasm contributes to a high-affinity site for inhibition by internal Cd{sup 2+}. Crystal structure of its cytoplasmic domain in complex with Cd{sup 2+} reveals that octahedral coordination geometry supports the high-affinity binding. This mode of action causes the tethering of the N-terminus to CD loop in the stimulus-sensing domain, suggesting that their conformational changes participate in gating and Cd{sup 2+} inhibits Kir3.2 by trapping the conformation in the closed state like 'inverse agonist'.

  19. Kir 4.1 inward rectifier potassium channel is upregulated in astrocytes in a murine multiple sclerosis model.

    Science.gov (United States)

    Mercado, Francisco; Almanza, Angélica; Rubio, Nazario; Soto, Enrique

    2018-06-11

    Multiple sclerosis (MS) is a high prevalence degenerative disease characterized at the cellular level by glial and neuronal cell death. The causes of cell death during the disease course are not fully understood. In this work we demonstrate that in a MS model induced by Theiler's murine encephalomyelitis virus (TMEV) infection, the inward rectifier (Kir) 4.1 potassium channel subunit is overexpressed in astrocytes. In voltage clamp experiments the inward current density from TMEV-infected astrocytes was significantly larger than in mock-infected ones. The cRNA hybridization analysis from mock- and TMEV-infected cells showed an upregulation of a potassium transport channel coding sequence. We validated this mRNA increase by RT-PCR and quantitative PCR using Kir 4.1 specific primers. Western blotting experiments confirmed the upregulation of Kir 4.1, and alignment between sequences provided the demonstration that the over-expressed gene encodes for a Kir family member. Flow cytometry showed that the Kir 4.1 protein is located mainly in the cell membrane in mock and TMEV-infected astrocytes. Our results demonstrate an increase in K + inward current in TMEV-infected glial cells, this increment may reduce the neuronal depolarization, contributing to cell resilience mechanisms. Copyright © 2018 Elsevier B.V. All rights reserved.

  20. Low Resting Membrane Potential and Low Inward Rectifier Potassium Currents Are Not Inherent Features of hiPSC-Derived Cardiomyocytes.

    Science.gov (United States)

    Horváth, András; Lemoine, Marc D; Löser, Alexandra; Mannhardt, Ingra; Flenner, Frederik; Uzun, Ahmet Umur; Neuber, Christiane; Breckwoldt, Kaja; Hansen, Arne; Girdauskas, Evaldas; Reichenspurner, Hermann; Willems, Stephan; Jost, Norbert; Wettwer, Erich; Eschenhagen, Thomas; Christ, Torsten

    2018-03-13

    Human induced pluripotent stem cell (hiPSC) cardiomyocytes (CMs) show less negative resting membrane potential (RMP), which is attributed to small inward rectifier currents (I K1 ). Here, I K1 was measured in hiPSC-CMs (proprietary and commercial cell line) cultured as monolayer (ML) or 3D engineered heart tissue (EHT) and, for direct comparison, in CMs from human right atrial (RA) and left ventricular (LV) tissue. RMP was measured in isolated cells and intact tissues. I K1 density in ML- and EHT-CMs from the proprietary line was similar to LV and RA, respectively. I K1 density in EHT-CMs from the commercial line was 2-fold smaller than in the proprietary line. RMP in EHT of both lines was similar to RA and LV. Repolarization fraction and I K,ACh response discriminated best between RA and LV and indicated predominantly ventricular phenotype in hiPSC-CMs/EHT. The data indicate that I K1 is not necessarily low in hiPSC-CMs, and technical issues may underlie low RMP in hiPSC-CMs. Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.

  1. Effects of different components of serum after radiation, burn and combined radiation-burn injury on inward rectifier potassium channel of myocardial cells

    International Nuclear Information System (INIS)

    Ye Benlan; Cheng Tianmin; Xiao Jiasi

    1997-01-01

    Objective: To study the effects of different components of serum in rats inflicted with radiation, burn and combined radiation-burn injury on inward rectifier potassium channel of cultured myocardial cells. Method: Using patch clamp method to study the action of single ion channel. Results: The low molecular and lipid components of serum after different injuries models could all activate the inward rectifier potassium channel in cultured myocardial cells. The components of serum after combined radiation-burn injury showed the most significant effect, and the way of this effect was different from that from single injury. Conclusion: The serum components post injury altered the electric characteristic of myocardial cells, which may play a role in the combined effect of depressed cardiac function after combined radiation-burn injury

  2. Inhibition of Inwardly Rectifying Potassium (Kir 4.1 Channels Facilitates Brain-Derived Neurotrophic Factor (BDNF Expression in Astrocytes

    Directory of Open Access Journals (Sweden)

    Masato Kinboshi

    2017-12-01

    Full Text Available Inwardly rectifying potassium (Kir 4.1 channels in astrocytes regulate neuronal excitability by mediating spatial potassium buffering. Although dysfunction of astrocytic Kir4.1 channels is implicated in the development of epileptic seizures, the functional mechanisms of Kir4.1 channels in modulating epileptogenesis remain unknown. We herein evaluated the effects of Kir4.1 inhibition (blockade and knockdown on expression of brain-derived neurotrophic factor (BDNF, a key modulator of epileptogenesis, in the primary cultures of mouse astrocytes. For blockade of Kir4.1 channels, we tested several antidepressant agents which reportedly bound to and blocked Kir4.1 channels in a subunit-specific manner. Treatment of astrocytes with fluoxetine enhanced BDNF mRNA expression in a concentration-dependent manner and increased the BDNF protein level. Other antidepressants (e.g., sertraline and imipramine also increased the expression of BDNF mRNA with relative potencies similar to those for inhibition of Kir4.1 channels. In addition, suppression of Kir4.1 expression by the transfection of small interfering RNA (siRNA targeting Kir4.1 significantly increased the mRNA and protein levels of BDNF. The BDNF induction by Kir4.1 siRNA transfection was suppressed by the MEK1/2 inhibitor U0126, but not by the p38 MAPK inhibitor SB202190 or the JNK inhibitor SP600125. The present results demonstrated that inhibition of Kir4.1 channels facilitates BDNF expression in astrocytes primarily by activating the Ras/Raf/MEK/ERK pathway, which may be linked to the development of epilepsy and other neuropsychiatric disorders.

  3. Inhibition of G-Protein-Activated Inwardly Rectifying K+ Channels by the Selective Norepinephrine Reuptake Inhibitors Atomoxetine and Reboxetine

    Science.gov (United States)

    Kobayashi, Toru; Washiyama, Kazuo; Ikeda, Kazutaka

    2010-01-01

    Atomoxetine and reboxetine are commonly used as selective norepinephrine reuptake inhibitors (NRIs) for the treatment of attention-deficit/hyperactivity disorder and depression, respectively. Furthermore, recent studies have suggested that NRIs may be useful for the treatment of several other psychiatric disorders. However, the molecular mechanisms underlying the various effects of NRIs have not yet been sufficiently clarified. G-protein-activated inwardly rectifying K+ (GIRK or Kir3) channels have an important function in regulating neuronal excitability and heart rate, and GIRK channel modulation has been suggested to be a potential treatment for several neuropsychiatric disorders and cardiac arrhythmias. In this study, we investigated the effects of atomoxetine and reboxetine on GIRK channels using the Xenopus oocyte expression assay. In oocytes injected with mRNA for GIRK1/GIRK2, GIRK2, or GIRK1/GIRK4 subunits, extracellular application of atomoxetine or reboxetine reversibly reduced GIRK currents. The inhibitory effects were concentration-dependent, but voltage-independent, and time-independent during each voltage pulse. However, Kir1.1 and Kir2.1 channels were insensitive to atomoxetine and reboxetine. Atomoxetine and reboxetine also inhibited GIRK currents induced by activation of cloned A1 adenosine receptors or by intracellularly applied GTPγS, a nonhydrolyzable GTP analogue. Furthermore, the GIRK currents induced by ethanol were concentration-dependently inhibited by extracellularly applied atomoxetine but not by intracellularly applied atomoxetine. The present results suggest that atomoxetine and reboxetine inhibit brain- and cardiac-type GIRK channels, revealing a novel characteristic of clinically used NRIs. GIRK channel inhibition may contribute to some of the therapeutic effects of NRIs and adverse side effects related to nervous system and heart function. PMID:20393461

  4. Cholesterol up-regulates neuronal G protein-gated inwardly rectifying potassium (GIRK) channel activity in the hippocampus.

    Science.gov (United States)

    Bukiya, Anna N; Durdagi, Serdar; Noskov, Sergei; Rosenhouse-Dantsker, Avia

    2017-04-14

    Hypercholesterolemia is a well known risk factor for the development of neurodegenerative disease. However, the underlying mechanisms are mostly unknown. In recent years, it has become increasingly evident that cholesterol-driven effects on physiology and pathophysiology derive from its ability to alter the function of a variety of membrane proteins including ion channels. Yet, the effect of cholesterol on G protein-gated inwardly rectifying potassium (GIRK) channels expressed in the brain is unknown. GIRK channels mediate the actions of inhibitory brain neurotransmitters. As a result, loss of GIRK function can enhance neuron excitability, whereas gain of GIRK function can reduce neuronal activity. Here we show that in rats on a high-cholesterol diet, cholesterol levels in hippocampal neurons are increased. We also demonstrate that cholesterol plays a critical role in modulating neuronal GIRK currents. Specifically, cholesterol enrichment of rat hippocampal neurons resulted in enhanced channel activity. In accordance, elevated currents upon cholesterol enrichment were also observed in Xenopus oocytes expressing GIRK2 channels, the primary GIRK subunit expressed in the brain. Furthermore, using planar lipid bilayers, we show that although cholesterol did not affect the unitary conductance of GIRK2, it significantly enhanced the frequency of channel openings. Last, combining computational and functional approaches, we identified two putative cholesterol-binding sites in the transmembrane domain of GIRK2. These findings establish that cholesterol plays a critical role in modulating GIRK activity in the brain. Because up-regulation of GIRK function can reduce neuronal activity, our findings may lead to novel approaches for prevention and therapy of cholesterol-driven neurodegenerative disease. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  5. Role of suppression of the inward rectifier current in terminal action potential repolarization in the failing heart.

    Science.gov (United States)

    Klein, Michael G; Shou, Matie; Stohlman, Jayna; Solhjoo, Soroosh; Haigney, Myles; Tidwell, Richard R; Goldstein, Robert E; Flagg, Thomas P; Haigney, Mark C

    2017-08-01

    The failing heart exhibits an increased arrhythmia susceptibility that is often attributed to action potential (AP) prolongation due to significant ion channel remodeling. The inwardly rectifying K + current (I K1 ) has been reported to be reduced, but its contribution to shaping the AP waveform and cell excitability in the failing heart remains unclear. The purpose of this study was to define the effect of I K1 suppression on the cardiac AP and excitability in the normal and failing hearts. We used electrophysiological and pharmacological approaches to investigate I K1 function in a swine tachy-pacing model of heart failure (HF). Terminal repolarization of the AP (TRAP; the time constant of the exponential fit to terminal repolarization) was markedly prolonged in both myocytes and arterially perfused wedges from animals with HF. TRAP was increased by 54.1% in HF myocytes (P < .001) and 26.2% in HF wedges (P = .014). The increase in TRAP was recapitulated by the potent and specific I K1 inhibitor, PA-6 (pentamidine analog 6), indicating that I K1 is the primary determinant of the final phase of repolarization. Moreover, we find that I K1 suppression reduced the ratio of effective refractory period to AP duration at 90% of repolarization, permitting re-excitation before full repolarization, reduction of AP upstroke velocity, and likely promotion of slow conduction. Using an objective measure of terminal repolarization, we conclude that I K1 is the major determinant of the terminal repolarization time course. Moreover, suppression of I K1 prolongs repolarization and reduces postrepolarization refractoriness without marked effects on the overall AP duration. Collectively, these findings demonstrate how I K1 suppression may contribute to arrhythmogenesis in the failing heart. Published by Elsevier Inc.

  6. Acute desensitization of acetylcholine and endothelin-1 activated inward rectifier K+ current in myocytes from the cardiac atrioventricular node.

    Science.gov (United States)

    Choisy, Stéphanie C M; James, Andrew F; Hancox, Jules C

    2012-07-06

    The atrioventricular node (AVN) is a vital component of the pacemaker-conduction system of the heart, co-ordinating conduction of electrical excitation from cardiac atria to ventricles and acting as a secondary pacemaker. The electrical behaviour of the AVN is modulated by vagal activity via activation of muscarinic potassium current, IKACh. However, it is not yet known if this response exhibits 'fade' or desensitization in the AVN, as established for the heart's primary pacemaker--the sinoatrial node. In this study, acute activation of IKACh in rabbit single AVN cells was investigated using whole-cell patch clamp at 37 °C. 0.1-1 μM acetylcholine (ACh) rapidly activated a robust IKACh in AVN myocytes during a descending voltage-ramp protocol. This response was inhibited by tertiapin-Q (TQ; 300 nM) and by the M2 muscarinic ACh receptor antagonist AFDX-116 (1 μM). During sustained ACh exposure the elicited IKACh exhibited bi-exponential fade (τf of 2.0 s and τs 76.9 s at -120 mV; 1 μM ACh). 10 nM ET-1 elicited a current similar to IKACh, which faded with a mono-exponential time-course (τ of 52.6 s at -120 mV). When ET-1 was applied following ACh, the ET-1 activated response was greatly attenuated, demonstrating that ACh could desensitize the response to ET-1. For neither ACh nor ET-1 was the rate of current fade dependent upon the initial response magnitude, which is inconsistent with K+ flux mediated changes in electrochemical driving force as the underlying mechanism. Collectively, these findings demonstrate that TQ sensitive inwardly rectifying K+ current in cardiac AVN cells, elicited by M2 muscarinic receptor or ET-1 receptor activation, exhibits fade due to rapid desensitization. Copyright © 2012 Elsevier Inc. All rights reserved.

  7. Expression of inwardly rectifying potassium channels (GIRKs) and beta-adrenergic regulation of breast cancer cell lines

    International Nuclear Information System (INIS)

    Plummer, Howard K III; Yu, Qiang; Cakir, Yavuz; Schuller, Hildegard M

    2004-01-01

    Previous research has indicated that at various organ sites there is a subset of adenocarcinomas that is regulated by beta-adrenergic and arachidonic acid-mediated signal transduction pathways. We wished to determine if this regulation exists in breast adenocarcinomas. Expression of mRNA that encodes a G-protein coupled inwardly rectifying potassium channel (GIRK1) has been shown in tissue samples from approximately 40% of primary human breast cancers. Previously, GIRK channels have been associated with beta-adrenergic signaling. Breast cancer cell lines were screened for GIRK channels by RT-PCR. Cell cultures of breast cancer cells were treated with beta-adrenergic agonists and antagonists, and changes in gene expression were determined by both relative competitive and real time PCR. Potassium flux was determined by flow cytometry and cell signaling was determined by western blotting. Breast cancer cell lines MCF-7, MDA-MB-361 MDA-MB 453, and ZR-75-1 expressed mRNA for the GIRK1 channel, while MDA-MB-468 and MDA-MB-435S did not. GIRK4 was expressed in all six breast cancer cell lines, and GIRK2 was expressed in all but ZR-75-1 and MDA-MB-435. Exposure of MDA-MB-453 cells for 6 days to the beta-blocker propranolol (1 μM) increased the GIRK1 mRNA levels and decreased beta 2 -adrenergic mRNA levels, while treatment for 30 minutes daily for 7 days had no effect. Exposure to a beta-adrenergic agonist and antagonist for 24 hours had no effect on gene expression. The beta adrenergic agonist, formoterol hemifumarate, led to increases in K + flux into MDA-MB-453 cells, and this increase was inhibited by the GIRK channel inhibitor clozapine. The tobacco carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), a high affinity agonist for beta-adrenergic receptors stimulated activation of Erk 1/2 in MDA-MB-453 cells. Our data suggests β-adrenergic receptors and GIRK channels may play a role in breast cancer

  8. Divalent ion block of inward rectifier current in human capillary endothelial cells and effects on resting membrane potential

    Science.gov (United States)

    Jow, Flora; Numann, Randy

    1998-01-01

    Cultured human capillary endothelial cells (HCEC) contain a large inward rectifier current, IK(IR), that can be abolished by removing external K+ or by adding 50 μm Ba2+.We show that IK(IR) is responsible for maintaining the hyperpolarized potential (−60.6 ± 0.5 mV, n = 83) of HCEC. Blocking IK(IR) with 50 μm Ba2+ shifts the zero current level and depolarizes HCEC by 36.5 ± 1.3 mV (n = 4).Increasing external Ca2+ concentration ([Ca2+]o) from 0.5 to 7 mm reduces the magnitude of IK(IR) by 36.5 ± 2.3 % (n = 5) and depolarizes the cells by 10.33 ± 2.4 mV (n = 3), whereas decreasing [Ca2+]o from 1.8 to 0.5 mm increases the amplitude of IK(IR) by 6.9 ± 1.9 % (n = 4). The relationship between [Ca2+]o and the percentage block of IK(IR) gives a Kd value of 5.4 ± 0.6 mm at −120 mV.IK(IR) is also blocked by other divalent ions, with Ba2+ >> Sr2+ > Mg2+ > Mn2+= Ca2+, and the block of peak current at −120 mV being 85.3 ± 3.2 % (n = 5) for 50 μm Ba2+, 62.9 ± 2.2 % (n = 5) for 5 mm Sr2+, 40.7 ± 2.5 % (n = 9) for 5 mm Mg2+, 33.4 ± 2.1 % (n = 5) for 5 mm Mn2+ and 32.9 ± 2.1 % (n = 5) for 5 mm Ca2+.The voltage dependence of Sr2+ block of peak IK(IR) occurred with a Kd value of 1.0 ± 0.09 mm for −140 mV, 1.9 ± 0.16 mm for −130 mV, 3.1 ± 0.28 mm for −120 mV, 4.6 ± 0.34 mm for −110 mV and 6.4 ± 0.5 mm for −100 mV (n = 5), with a calculated electrical distance (δ) of 0.44 from the outside. PMID:9729622

  9. Mechanism underlying selective regulation of G protein-gated inwardly rectifying potassium channels by the psychostimulant-sensitive sorting nexin 27

    Science.gov (United States)

    Balana, Bartosz; Maslennikov, Innokentiy; Kwiatkowski, Witek; Stern, Kalyn M.; Bahima, Laia; Choe, Senyon; Slesinger, Paul A.

    2011-01-01

    G protein-gated inwardly rectifying potassium (GIRK) channels are important gatekeepers of neuronal excitability. The surface expression of neuronal GIRK channels is regulated by the psychostimulant-sensitive sorting nexin 27 (SNX27) protein through a class I (-X-Ser/Thr-X-Φ, where X is any residue and Φ is a hydrophobic amino acid) PDZ-binding interaction. The G protein-insensitive inward rectifier channel (IRK1) contains the same class I PDZ-binding motif but associates with a different synaptic PDZ protein, postsynaptic density protein 95 (PSD95). The mechanism by which SNX27 and PSD95 discriminate these channels was previously unclear. Using high-resolution structures coupled with biochemical and functional analyses, we identified key amino acids upstream of the channel's canonical PDZ-binding motif that associate electrostatically with a unique structural pocket in the SNX27-PDZ domain. Changing specific charged residues in the channel's carboxyl terminus or in the PDZ domain converts the selective association and functional regulation by SNX27. Elucidation of this unique interaction site between ion channels and PDZ-containing proteins could provide a therapeutic target for treating brain diseases. PMID:21422294

  10. Investigations on contribution of glial inwardly-rectifying K+ current to membrane potential and ion flux: An experimental and theoretical study

    Directory of Open Access Journals (Sweden)

    Sheng-Nan Wu

    2015-01-01

    Full Text Available The inwardly rectifying K+ current [IK(IR] allows large inward K+ currents at potentials negative to K+ equilibrium potential (EK and it becomes small outward K+ currents at those positive to EK. How changes of such currents enriched in glial cells can influence the functions of glial cell, neurons, or both is not clearly defined, although mutations of Kir4.1 channels have been demonstrated to cause serious neurological disorders. In this study, we identified the presence of IK(IR in human glioma cells (U373 and U87 cells. The amplitude of IK(IR in U373 cells was subject to inhibition by amitriptyline, arecoline, or BaCl2. The activity of inwardly rectifying K+ channels was also clearly detected, and single-channel conductance of these channels was calculated to be around 23 pS. Moreover, based on a simulation model derived from neuron–glial interaction mediated by ion flux, we further found out that incorporation of glial IK(IR conductance into the model can significantly contribute to regulation of extracellular K+ concentrations and glial resting potential, particularly during high-frequency stimulation. Glial cells and neurons can mutually modulate their expression of ion channels through K+ ions released into the extracellular space. It is thus anticipated that glial IK(IR may be a potential target utilized to influence the activity of neuronal and glial cells as well as their interaction.

  11. Impaired Na⁺-dependent regulation of acetylcholine-activated inward-rectifier K⁺ current modulates action potential rate dependence in patients with chronic atrial fibrillation.

    Science.gov (United States)

    Voigt, Niels; Heijman, Jordi; Trausch, Anne; Mintert-Jancke, Elisa; Pott, Lutz; Ravens, Ursula; Dobrev, Dobromir

    2013-08-01

    Shortened action-potential duration (APD) and blunted APD rate adaptation are hallmarks of chronic atrial fibrillation (cAF). Basal and muscarinic (M)-receptor-activated inward-rectifier K(+) currents (IK1 and IK,ACh, respectively) contribute to regulation of human atrial APD and are subject to cAF-dependent remodeling. Intracellular Na(+) ([Na(+)]i) enhances IK,ACh in experimental models but the effect of [Na(+)]i-dependent regulation of inward-rectifier K(+) currents on APD in human atrial myocytes is currently unknown. Here, we report a [Na(+)]i-dependent inhibition of outward IK1 in atrial myocytes from sinus rhythm (SR) or cAF patients. In contrast, IK,ACh activated by carbachol, a non-selective M-receptor agonist, increased with elevation of [Na(+)]i in SR. This [Na(+)]i-dependent IK,ACh regulation was absent in cAF. Including [Na(+)]i dependence of IK1 and IK,ACh in a recent computational model of the human atrial myocyte revealed that [Na(+)]i accumulation at fast rates inhibits IK1 and blunts physiological APD rate dependence in both groups. [Na(+)]i-dependent IK,ACh augmentation at fast rates increased APD rate dependence in SR, but not in cAF. These results identify impaired Na(+)-sensitivity of IK,ACh as one potential mechanism contributing to the blunted APD rate dependence in patients with cAF. This article is part of a Special Issue entitled "Na(+) Regulation in Cardiac Myocytes". Copyright © 2013 The Authors. Published by Elsevier Ltd.. All rights reserved.

  12. From in silico to in vitro: a trip to reveal flavonoid binding on the Rattus norvegicus Kir6.1 ATP-sensitive inward rectifier potassium channel.

    Science.gov (United States)

    Trezza, Alfonso; Cicaloni, Vittoria; Porciatti, Piera; Langella, Andrea; Fusi, Fabio; Saponara, Simona; Spiga, Ottavia

    2018-01-01

    ATP-sensitive inward rectifier potassium channels (Kir), are a potassium channel family involved in many physiological processes. K ATP dysfunctions are observed in several diseases such as hypoglycaemia, hyperinsulinemia, Prinzmetal angina-like symptoms, cardiovascular diseases. A broader view of the K ATP mechanism is needed in order to operate on their regulation, and in this work we clarify the structure of the Rattus norvegicus ATP-sensitive inward rectifier potassium channel 8 (Kir6.1), which has been obtained through a homology modelling procedure. Due to the medical use of flavonoids, a considerable increase in studies on their influence on human health has recently been observed, therefore our aim is to study, through computational methods, the three-dimensional (3D) conformation together with mechanism of action of Kir6.1 with three flavonoids. Computational analysis by performing molecular dynamics (MD) and docking simulation on rat 3D modelled structure have been completed, in its closed and open conformation state and in complex with Quercetin, 5-Hydroxyflavone and Rutin flavonoids. Our study showed that only Quercetin and 5-Hydroxyflavone were responsible for a significant down-regulation of the Kir6.1 activity, stabilising it in a closed conformation. This hypothesis was supported by in vitro experiments demonstrating that Quercetin and 5-Hydroxyflavone were capable to inhibit K ATP currents of rat tail main artery myocytes recorded by the patch-clamp technique. Combined methodological approaches, such as molecular modelling, docking and MD simulations of Kir6.1 channel, used to elucidate flavonoids intrinsic mechanism of action, are introduced, revealing a new potential druggable protein site.

  13. Two modes of polyamine block regulating the cardiac inward rectifier K+ current IK1 as revealed by a study of the Kir2.1 channel expressed in a human cell line.

    Science.gov (United States)

    Ishihara, Keiko; Ehara, Tsuguhisa

    2004-04-01

    The strong inward rectifier K(+) current, I(K1), shows significant outward current amplitude in the voltage range near the reversal potential and thereby causes rapid repolarization at the final phase of cardiac action potentials. However, the mechanism that generates the outward I(K1) is not well understood. We recorded currents from the inside-out patches of HEK 293T cells that express the strong inward rectifier K(+) channel Kir2.1 and studied the blockage of the currents caused by cytoplasmic polyamines, namely, spermine and spermidine. The outward current-voltage (I-V) relationships of Kir2.1, obtained with 5-10 microm spermine or 10-100 microm spermidine, were similar to the steady-state outward I-V relationship of I(K1), showing a peak at a level that is approximately 20 mV more positive than the reversal potential, with a negative slope at more positive voltages. The relationships exhibited a plateau or a double-hump shape with 1 microm spermine/spermidine or 0.1 microm spermine, respectively. In the chord conductance-voltage relationships, there were extra conductances in the positive voltage range, which could not be described by the Boltzmann relations fitting the major part of the relationships. The extra conductances, which generated most of the outward currents in the presence of 5-10 microm spermine or 10-100 microm spermidine, were quantitatively explained by a model that considered two populations of Kir2.1 channels, which were blocked by polyamines in either a high-affinity mode (Mode 1 channel) or a low-affinity mode (Mode 2 channel). Analysis of the inward tail currents following test pulses indicated that the relief from the spermine block of Kir2.1 consisted of an exponential component and a virtually instantaneous component. The fractions of the two components nearly agreed with the fractions of the blockages in Mode 1 and Mode 2 calculated by the model. The estimated proportion of Mode 1 channels to total channels was 0.9 with 0.1-10 microm

  14. G-protein inwardly rectifying potassium channel 1 (GIRK 1) gene expression correlates with tumor progression in non-small cell lung cancer

    International Nuclear Information System (INIS)

    Takanami, Iwao; Inoue, Yoshimasa; Gika, Masatoshi

    2004-01-01

    G-protein inwardly rectifying potassium channel 1 (GIRK1) is thought to play a role in cell proliferation in cancer, and GIRK1 gene expression level may define a more aggressive phenotype. We detected GIRK1 expression in tissue specimens from patients with non-small cell lung cancers (NSCLCs) and assessed their clinical characteristics. Using reverse transcription-polymerase chain reaction (RT-PCR) analyses, we quantified the expression of GIRK1 in 72 patients with NSCLCs to investigate the relationship between GIRK1 expression and clinicopathologic factors and prognosis. In 72 NSCLC patients, 50 (69%) samples were evaluated as having high GIRK1 gene expression, and 22 (31%) were evaluated as having low GIRK1 gene expression. GIRK1 gene expression was significantly associated with lymph node metastasis, stage (p = 0.0194 for lymph node metastasis; p = 0.0207 for stage). The overall and stage I survival rates for patients with high GIRK1 gene expressed tumors was significantly worse than for those individuals whose tumors had low GIRK1 expression (p = 0.0004 for the overall group; p = 0.0376 for stage I). These data indicate that GIRK1 may contribute to tumor progression and GIRK1 gene expression can serve as a useful prognostic marker in the overall and stage I NSCLCs

  15. Over-Expression of Dopamine D2 Receptor and Inwardly Rectifying Potassium Channel Genes in Drug-Naive Schizophrenic Peripheral Blood Lymphocytes as Potential Diagnostic Markers

    Directory of Open Access Journals (Sweden)

    Ágnes Zvara

    2005-01-01

    Full Text Available Schizophrenia is one of the most common neuropsychiatric disorders affecting nearly 1% of the human population. Current diagnosis of schizophrenia is based on complex clinical symptoms. The use of easily detectable peripheral molecular markers could substantially help the diagnosis of psychiatric disorders. Recent studies showed that peripheral blood lymphocytes (PBL express subtypes of D1 and D2 subclasses of dopamine receptors. Recently, dopamine receptor D3 (DRD3 was found to be over-expressed in schizophrenic PBL and proposed to be a diagnostic and follow-up marker for schizophrenia. In this study we screened PBL of 13 drug-naive/drug-free schizophrenic patients to identify additional markers of schizophrenia. One of the benefits of our study is the use of blood samples of non-medicated, drug-naive patients. This excludes the possibility that changes detected in gene expression levels might be attributed to the medication rather than to the disorder itself. Among others, genes for dopamine receptor D2 (DRD2 and the inwardly rectifying potassium channel (Kir2.3 were found to be over-expressed in microarray analysis. Increased mRNA levels were confirmed by quantitative real-time PCR (QRT-PCR using the SybrGreen method and dual labeled TaqMan probes. The use of both molecular markers allows a more rapid and precise prediction of schizophrenia and might help find the optimal medication for schizophrenic patients.

  16. Inward rectifier K+ channel and T-type Ca2+ channel contribute to enhancement of GABAergic transmission induced by β1-adrenoceptor in the prefrontal cortex.

    Science.gov (United States)

    Luo, Fei; Zheng, Jian; Sun, Xuan; Tang, Hua

    2017-02-01

    The functions of prefrontal cortex (PFC) are sensitive to norepinephrine (NE). Endogenously released NE influences synaptic transmission through activation of different subtypes of adrenergic receptors in PFC including α 1 , α 2 , β 1 or β 2 -adrenoceptor. Our recent study has revealed that β 1 -adrenoceptor (β 1 -AR) activation modulates glutamatergic transmission in the PFC, whereas the roles of β 1 -AR in GABAergic transmission are elusive. In the current study, we probed the effects of the β 1 -AR agonist dobutamine (Dobu) on GABAergic transmission onto pyramidal neurons in the PFC of juvenile rats. Dobu increased both the frequency and amplitude of miniature IPSCs (mIPSCs). Ca 2+ influx through T-type voltage-gated Ca 2+ channel was required for Dobu-enhanced mIPSC frequency. We also found that Dobu facilitated GABA release probability and the number of releasable vesicles through regulating T-type Ca 2+ channel. Dobu depolarized GABAergic fast-spiking (FS) interneurons with no effects on the firing rate of action potentials (APs) of interneurons. Dobu-induced depolarization of FS interneurons required inward rectifier K + channel (Kir). Our results suggest that Dobu increase GABA release via inhibition of Kir, which further depolarizes FS interneurons resulting in Ca 2+ influx via T-type Ca 2+ channel. Copyright © 2016 Elsevier Inc. All rights reserved.

  17. Effects of aquaporin 4 and inward rectifier potassium channel 4.1 on medullospinal edema after methylprednisolone treatment to suppress acute spinal cord injury in rats.

    Science.gov (United States)

    Li, Ye; Hu, Haifeng; Liu, Jingchen; Zhu, Qingsan; Gu, Rui

    2018-02-01

    To investigate the effects of aquaporin 4 (AQP4) and inward rectifier potassium channel 4.1 (Kir4.1) on medullospinal edema after treatment with methylprednisolone (MP) to suppress acute spinal cord injury (ASCI) in rats. Sprague Dawley rats were randomly divided into control, sham, ASCI, and MP-treated ASCI groups. After the induction of ASCI, we injected 30 mg/kg MP via the tail vein at various time points. The Tarlov scoring method was applied to evaluate neurological symptoms, and the wet-dry weights method was applied to measure the water content of the spinal cord. The motor function score of the ASCI group was significantly lower than that of the sham group, and the spinal water content was significantly increased. In addition, the levels of AQP4 and Kir4.1 were significantly increased, as was their degree of coexpression. Compared with that in the ASCI group, the motor function score and the water content were significantly increased in the MP group; in addition, the expression and coexpression of AQP4 and Kir4.1 were significantly reduced. Methylprednisolone inhibited medullospinal edema in rats with acute spinal cord injury, possibly by reducing the coexpression of aquaporin 4 and Kir4.1 in medullospinal tissues.

  18. Expression and contributions of the Kir2.1 inward-rectifier K+ channel to proliferation, migration and chemotaxis of microglia in unstimulated and anti-inflammatory states

    Directory of Open Access Journals (Sweden)

    Doris eLam

    2015-05-01

    Full Text Available When microglia respond to CNS damage, they can range from pro-inflammatory (classical, M1 to anti-inflammatory, alternative (M2 and acquired deactivation states. It is important to determine how microglial functions are affected by these activation states, and to identify molecules that regulate their behavior. Microglial proliferation and migration are crucial during development and following damage in the adult, and both functions are Ca2+-dependent. In many cell types, the membrane potential and driving force for Ca2+ influx are regulated by inward-rectifier K+ channels, including Kir2.1, which is prevalent in microglia. However, it is not known whether Kir2.1 expression and contributions are altered in anti-inflammatory states. We tested the hypothesis that Kir2.1 contributes to Ca2+ entry, proliferation and migration of rat microglia. Kir2.1 (KCNJ2 transcript expression, current amplitude, and proliferation were comparable in unstimulated microglia and following alternative activation (IL-4 stimulated and acquired deactivation (IL-10 stimulated. To examine functional roles of Kir2.1 in microglia, we first determined that ML133 was more effective than the commonly used blocker, Ba2+; i.e., ML133 was potent (IC50=3.5 M and voltage independent. Both blockers slightly increased proliferation in unstimulated or IL-4 (but not IL-10-stimulated microglia. Stimulation with IL-4 or IL-10 increased migration and ATP-induced chemotaxis, and blocking Kir2.1 greatly reduced both but ML133 was more effective. In all three activation states, blocking Kir2.1 with ML133 dramatically reduced Ca2+ influx through Ca2+-release-activated Ca2+ (CRAC channels. Thus, Kir2.1 channel activity is necessary for microglial Ca2+ signaling and migration under resting and anti-inflammatory states but the channel weakly inhibits proliferation.

  19. Regulation of inward rectifier potassium current ionic channel remodeling by AT1 -Calcineurin-NFAT signaling pathway in stretch-induced hypertrophic atrial myocytes.

    Science.gov (United States)

    He, Jionghong; Xu, Yanan; Yang, Long; Xia, Guiling; Deng, Na; Yang, Yongyao; Tian, Ye; Fu, Zenan; Huang, Yongqi

    2018-05-02

    Previous studies have shown that the activation of angiotensin II receptor type I (AT 1 ) is attributed to cardiac remodeling stimulated by increased heart load, and that it is followed by the activation of the calcineurin-nuclear factor of activated T-cells (NFAT) signaling pathway. Additionally, AT 1 has been found to be a regulator of cardiocyte ionic channel remodeling, and calcineurin-NFAT signals participate in the regulation of cardiocyte ionic channel expression. A hypothesis therefore follows that stretch stimulation may regulate cardiocyte ionic channel remodeling by activating the AT 1 -calcineurin-NFAT pathway. Here, we investigated the role of the AT 1 -calcineurin-NFAT pathway in the remodeling of inward rectifier potassium (I k1 ) channel, in addition to its role in changing action potential, in stretch-induced hypertrophic atrial myocytes of neonatal rats. Our results showed that increased stretch significantly led to atrial myocytes hypertrophy; it also increased the activity of calcineurin enzymatic activity, which was subsequently attenuated by telmisartan or cyclosporine-A. The level of NFAT 3 protein in nuclear extracts, the mRNA and protein expression of Kir2.1 in whole cell extracts, and the density of I k1 were noticeably increased in stretched samples. Stretch stimulation significantly shortened the action potential duration (APD) of repolarization at the 50% and 90% level. Telmisartan, cyclosporine-A, and 11R-VIVIT attenuated stretch-induced alterations in the levels of NFAT 3 , mRNA and protein expression of Kir2.1, the density of I k1 , and the APD. Our findings suggest that the AT 1 -calcineurin-NFAT signaling pathway played an important role in regulating I k1 channel remodeling and APD change in stretch-induced hypertrophic atrial myocytes of neonatal rats. This article is protected by copyright. All rights reserved.

  20. Histamine facilitates GABAergic transmission in the rat entorhinal cortex: Roles of H1 and H2 receptors, Na+ -permeable cation channels, and inward rectifier K+ channels.

    Science.gov (United States)

    Cilz, Nicholas I; Lei, Saobo

    2017-05-01

    In the brain, histamine (HA) serves as a neuromodulator and a neurotransmitter released from the tuberomammillary nucleus (TMN). HA is involved in wakefulness, thermoregulation, energy homeostasis, nociception, and learning and memory. The medial entorhinal cortex (MEC) receives inputs from the TMN and expresses HA receptors (H 1 , H 2 , and H 3 ). We investigated the effects of HA on GABAergic transmission in the MEC and found that HA significantly increased the frequency of spontaneous inhibitory postsynaptic currents (sIPSCs) with an EC 50 of 1.3 µM, but failed to significantly alter sIPSC amplitude. HA-induced increases in sIPSC frequency were sensitive to tetrodotoxin (TTX), required extracellular Ca 2+ , and persisted when GDP-β-S, a G-protein inactivator, was applied postsynaptically via the recording pipettes, indicating that HA increased GABA release by facilitating the excitability of GABAergic interneurons in the MEC. Recordings from local MEC interneurons revealed that HA significantly increased their excitability as determined by membrane depolarization, generation of an inward current at -65 mV, and augmentation of action potential firing frequency. Both H 1 and H 2 receptors were involved in HA-induced increases in sIPSCs and interneuron excitability. Immunohistochemical staining showed that both H 1 and H 2 receptors are expressed on GABAergic interneurons in the MEC. HA-induced depolarization of interneurons involved a mixed ionic mechanism including activation of a Na + -permeable cation channel and inhibition of a cesium-sensitive inward rectifier K + channel, although HA also inhibited the delayed rectifier K + channels. Our results may provide a cellular mechanism, at least partially, to explain the roles of HA in the brain. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  1. Neuronal and glial expression of inward rectifier potassium channel subunits Kir2.x in rat dorsal root ganglion and spinal cord.

    Science.gov (United States)

    Murata, Yuzo; Yasaka, Toshiharu; Takano, Makoto; Ishihara, Keiko

    2016-03-23

    Inward rectifier K(+) channels of the Kir2.x subfamily play important roles in controlling the neuronal excitability. Although their cellular localization in the brain has been extensively studied, only a few studies have examined their expression in the spinal cord and peripheral nervous system. In this study, immunohistochemical analyses of Kir2.1, Kir2.2, and Kir2.3 expression were performed in rat dorsal root ganglion (DRG) and spinal cord using bright-field and confocal microscopy. In DRG, most ganglionic neurons expressed Kir2.1, Kir2.2 and Kir2.3, whereas satellite glial cells chiefly expressed Kir2.3. In the spinal cord, Kir2.1, Kir2.2 and Kir2.3 were all expressed highly in the gray matter of dorsal and ventral horns and moderately in the white matter also. Within the gray matter, the expression was especially high in the substantia gelatinosa (lamina II). Confocal images obtained using markers for neuronal cells, NeuN, and astrocytes, Sox9, showed expression of all three Kir2 subunits in both neuronal somata and astrocytes in lamina I-III of the dorsal horn and the lateral spinal nucleus of the dorsolateral funiculus. Immunoreactive signals other than those in neuronal and glial somata were abundant in lamina I and II, which probably located mainly in nerve fibers or nerve terminals. Colocalization of Kir2.1 and 2.3 and that of Kir2.2 and 2.3 were present in neuronal and glial somata. In the ventral horn, motor neurons and interneurons were also immunoreactive with the three Kir2 subunits. Our study suggests that Kir2 channels composed of Kir2.1-2.3 subunits are expressed in neuronal and glial cells in the DRG and spinal cord, contributing to sensory transduction and motor control. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  2. Tissue Distribution of Kir7.1 Inwardly Rectifying K+ Channel Probed in a Knock-in Mouse Expressing a Haemagglutinin-Tagged Protein

    Directory of Open Access Journals (Sweden)

    Isabel Cornejo

    2018-04-01

    Full Text Available Kir7.1 encoded by the Kcnj13 gene in the mouse is an inwardly rectifying K+ channel present in epithelia where it shares membrane localization with the Na+/K+-pump. Further investigations of the localisation and function of Kir7.1 would benefit from the availability of a knockout mouse, but perinatal mortality attributed to cleft palate in the neonate has thwarted this research. To facilitate localisation studies we now use CRISPR/Cas9 technology to generate a knock-in mouse, the Kir7.1-HA that expresses the channel tagged with a haemagglutinin (HA epitope. The availability of antibodies for the HA epitope allows for application of western blot and immunolocalisation methods using widely available anti-HA antibodies with WT tissues providing unambiguous negative control. We demonstrate that Kir7.1-HA cloned from the choroid plexus of the knock-in mouse has the electrophysiological properties of the native channel, including characteristically large Rb+ currents. These large Kir7.1-mediated currents are accompanied by abundant apical membrane Kir7.1-HA immunoreactivity. WT-controlled western blots demonstrate the presence of Kir7.1-HA in the eye and the choroid plexus, trachea and lung, and intestinal epithelium but exclusively in the ileum. In the kidney, and at variance with previous reports in the rat and guinea-pig, Kir7.1-HA is expressed in the inner medulla but not in the cortex or outer medulla. In isolated tubules immunoreactivity was associated with inner medulla collecting ducts but not thin limbs of the loop of Henle. Kir7.1-HA shows basolateral expression in the respiratory tract epithelium from trachea to bronchioli. The channel also appears basolateral in the epithelium of the nasal cavity and nasopharynx in newborn animals. We show that HA-tagged Kir7.1 channel introduced in the mouse by a knock-in procedure has functional properties similar to the native protein and the animal thus generated has clear advantages in localisation

  3. Tissue Distribution of Kir7.1 Inwardly Rectifying K+ Channel Probed in a Knock-in Mouse Expressing a Haemagglutinin-Tagged Protein.

    Science.gov (United States)

    Cornejo, Isabel; Villanueva, Sandra; Burgos, Johanna; López-Cayuqueo, Karen I; Chambrey, Régine; Julio-Kalajzić, Francisca; Buelvas, Neudo; Niemeyer, María I; Figueiras-Fierro, Dulce; Brown, Peter D; Sepúlveda, Francisco V; Cid, L P

    2018-01-01

    Kir7.1 encoded by the Kcnj13 gene in the mouse is an inwardly rectifying K + channel present in epithelia where it shares membrane localization with the Na + /K + -pump. Further investigations of the localisation and function of Kir7.1 would benefit from the availability of a knockout mouse, but perinatal mortality attributed to cleft palate in the neonate has thwarted this research. To facilitate localisation studies we now use CRISPR/Cas9 technology to generate a knock-in mouse, the Kir7.1-HA that expresses the channel tagged with a haemagglutinin (HA) epitope. The availability of antibodies for the HA epitope allows for application of western blot and immunolocalisation methods using widely available anti-HA antibodies with WT tissues providing unambiguous negative control. We demonstrate that Kir7.1-HA cloned from the choroid plexus of the knock-in mouse has the electrophysiological properties of the native channel, including characteristically large Rb + currents. These large Kir7.1-mediated currents are accompanied by abundant apical membrane Kir7.1-HA immunoreactivity. WT-controlled western blots demonstrate the presence of Kir7.1-HA in the eye and the choroid plexus, trachea and lung, and intestinal epithelium but exclusively in the ileum. In the kidney, and at variance with previous reports in the rat and guinea-pig, Kir7.1-HA is expressed in the inner medulla but not in the cortex or outer medulla. In isolated tubules immunoreactivity was associated with inner medulla collecting ducts but not thin limbs of the loop of Henle. Kir7.1-HA shows basolateral expression in the respiratory tract epithelium from trachea to bronchioli. The channel also appears basolateral in the epithelium of the nasal cavity and nasopharynx in newborn animals. We show that HA-tagged Kir7.1 channel introduced in the mouse by a knock-in procedure has functional properties similar to the native protein and the animal thus generated has clear advantages in localisation studies. It

  4. Microbial eukaryote plankton communities of high-mountain lakes from three continents exhibit strong biogeographic patterns.

    Science.gov (United States)

    Filker, Sabine; Sommaruga, Ruben; Vila, Irma; Stoeck, Thorsten

    2016-05-01

    Microbial eukaryotes hold a key role in aquatic ecosystem functioning. Yet, their diversity in freshwater lakes, particularly in high-mountain lakes, is relatively unknown compared with the marine environment. Low nutrient availability, low water temperature and high ultraviolet radiation make most high-mountain lakes extremely challenging habitats for life and require specific molecular and physiological adaptations. We therefore expected that these ecosystems support a plankton diversity that differs notably from other freshwater lakes. In addition, we hypothesized that the communities under study exhibit geographic structuring. Our rationale was that geographic dispersal of small-sized eukaryotes in high-mountain lakes over continental distances seems difficult. We analysed hypervariable V4 fragments of the SSU rRNA gene to compare the genetic microbial eukaryote diversity in high-mountain lakes located in the European Alps, the Chilean Altiplano and the Ethiopian Bale Mountains. Microbial eukaryotes were not globally distributed corroborating patterns found for bacteria, multicellular animals and plants. Instead, the plankton community composition emerged as a highly specific fingerprint of a geographic region even on higher taxonomic levels. The intraregional heterogeneity of the investigated lakes was mirrored in shifts in microbial eukaryote community structure, which, however, was much less pronounced compared with interregional beta-diversity. Statistical analyses revealed that on a regional scale, environmental factors are strong predictors for plankton community structures in high-mountain lakes. While on long-distance scales (>10 000 km), isolation by distance is the most plausible scenario, on intermediate scales (up to 6000 km), both contemporary environmental factors and historical contingencies interact to shift plankton community structures. © 2016 John Wiley & Sons Ltd.

  5. Discovery, characterization and structure-activity relationships of an inhibitor of inward rectifier potassium (Kir channels with preference for Kir2.3, Kir3.X and Kir7.1

    Directory of Open Access Journals (Sweden)

    Jerod S Denton

    2011-11-01

    Full Text Available The inward rectifier family of potassium (Kir channels is comprised of at least 16 family members exhibiting broad and often overlapping cellular, tissue or organ distributions. The discovery of disease-causing mutations in humans and experiments on knockout mice has underscored the importance of Kir channels in physiology and in some cases raised questions about their potential as drug targets. However, the paucity of potent and selective small-molecule modulators targeting specific family members has with few exceptions mired efforts to understand their physiology and assess their therapeutic potential. A growing body of evidence suggests that GIRK (G protein-regulated inward rectifier K channels of the Kir3.X subfamily may represent novel targets for the treatment of atrial fibrillation. In an effort to expand the molecular pharmacology of GIRK, we performed a thallium (Tl+ flux-based high-throughput screen (HTS of a Kir1.1 inhibitor library for modulators of GIRK. One compound, termed VU573, exhibited 10-fold selectivity for GIRK over Kir1.1 (IC50 = 1.9 M and 19 M, respectively and was therefore selected for further study. In electrophysiological experiments performed on Xenopus laevis oocytes and mammalian cells, VU573 inhibited Kir3.1/3.2 (neuronal GIRK and Kir3.1/3.4 (cardiac GIRK channels with equal potency and preferentially inhibited GIRK, Kir2.3 and Kir7.1 over Kir1.1 and Kir2.1. Tl+ flux assays were established for Kir2.3 and the M125R pore mutant of Kir7.1 to support medicinal chemistry efforts to develop more potent and selective analogs for these channels. The structure-activity relationships of VU573 revealed few analogs with improved potency, however two compounds retained most of their activity toward GIRK and Kir2.3 and lost activity toward Kir7.1. We anticipate that the VU573 series will be useful for exploring the physiology and structure-function relationships of these Kir channels.

  6. Regulation of the instantaneous inward rectifier and the delayed outward rectifier potassium channels by Captopril and Angiotensin II via the Phosphoinositide-3 kinase pathway in volume-overload-induced hypertrophied cardiac myocytes.

    Science.gov (United States)

    Alvin, Zikiar V; Laurence, Graham G; Coleman, Bernell R; Zhao, Aiqiu; Hajj-Moussa, Majd; Haddad, Georges E

    2011-07-01

    Early development of cardiac hypertrophy may be beneficial but sustained hypertrophic activation leads to myocardial dysfunction. Regulation of the repolarizing currents can be modulated by the activation of humoral factors, such as angiotensin II (ANG II) through protein kinases. The aim of this work is to assess the regulation of IK and IK1 by ANG II through the PI3-K pathway in hypertrophied ventricular myocytes. Cardiac eccentric hypertrophy was induced through volume-overload in adult male rats by aorto-caval shunt (3 weeks). After one week half of the rats were given captopril (2 weeks; 0.5 g/l/day) and the other half served as control. The voltage-clamp and western blot techniques were used to measure the delayed outward rectifier potassium current (IK) and the instantaneous inward rectifier potassium current (IK1) and Akt activity, respectively. Hypertrophied cardiomyocytes showed reduction in IK and IK1. Treatment with captopril alleviated this difference seen between sham and shunt cardiomyocytes. Acute administration of ANG II (10-6M) to cardiocytes treated with captopril reduced IK and IK1 in shunts, but not in sham. Captopril treatment reversed ANG II effects on IK and IK1 in a PI3-K-independent manner. However in the absence of angiotensin converting enzyme inhibition, ANG II increased both IK and IK1 in a PI3-K-dependent manner in hypertrophied cardiomyocytes. Thus, captopril treatment reveals a negative effect of ANG II on IK and IK1, which is PI3-K independent, whereas in the absence of angiotensin converting enzyme inhibition IK and IK1 regulation is dependent upon PI3-K.

  7. Evalutating Inward Rectifier Current Inhibiton for Treatment of Atrial Fibrillation

    NARCIS (Netherlands)

    Ji, Yuan

    2017-01-01

    Atrial Fibrillation (AF) is one of the most common forms of cardiac arrhythmia and affects a large percentage of the human population, especially in the elderly. Currently, more than 6 million Europeans suffer from AF, and due to ageing this number will at least double in the next 50 years.

  8. A conductance maximum observed in an inward-rectifier potassium channel

    OpenAIRE

    1994-01-01

    One prediction of a multi-ion pore is that its conductance should reach a maximum and then begin to decrease as the concentration of permeant ion is raised equally on both sides of the membrane. A conductance maximum has been observed at the single-channel level in gramicidin and in a Ca(2+)-activated K+ channel at extremely high ion concentration (> 1,000 mM) (Hladky, S. B., and D. A. Haydon. 1972. Biochimica et Biophysica Acta. 274:294-312; Eisenmam, G., J. Sandblom, and E. Neher. 1977. In ...

  9. Discovery and characterization of a potent and selective inhibitor of Aedes aegypti inward rectifier potassium channels.

    Directory of Open Access Journals (Sweden)

    Rene Raphemot

    Full Text Available Vector-borne diseases such as dengue fever and malaria, which are transmitted by infected female mosquitoes, affect nearly half of the world's population. The emergence of insecticide-resistant mosquito populations is reducing the effectiveness of conventional insecticides and threatening current vector control strategies, which has created an urgent need to identify new molecular targets against which novel classes of insecticides can be developed. We previously demonstrated that small molecule inhibitors of mammalian Kir channels represent promising chemicals for new mosquitocide development. In this study, high-throughput screening of approximately 30,000 chemically diverse small-molecules was employed to discover potent and selective inhibitors of Aedes aegypti Kir1 (AeKir1 channels heterologously expressed in HEK293 cells. Of 283 confirmed screening 'hits', the small-molecule inhibitor VU625 was selected for lead optimization and in vivo studies based on its potency and selectivity toward AeKir1, and tractability for medicinal chemistry. In patch clamp electrophysiology experiments of HEK293 cells, VU625 inhibits AeKir1 with an IC50 value of 96.8 nM, making VU625 the most potent inhibitor of AeKir1 described to date. Furthermore, electrophysiology experiments in Xenopus oocytes revealed that VU625 is a weak inhibitor of AeKir2B. Surprisingly, injection of VU625 failed to elicit significant effects on mosquito behavior, urine excretion, or survival. However, when co-injected with probenecid, VU625 inhibited the excretory capacity of mosquitoes and was toxic, suggesting that the compound is a substrate of organic anion and/or ATP-binding cassette (ABC transporters. The dose-toxicity relationship of VU625 (when co-injected with probenecid is biphasic, which is consistent with the molecule inhibiting both AeKir1 and AeKir2B with different potencies. This study demonstrates proof-of-concept that potent and highly selective inhibitors of mosquito Kir channels can be developed using conventional drug discovery approaches. Furthermore, it reinforces the notion that the physical and chemical properties that determine a compound's bioavailability in vivo will be critical in determining the efficacy of Kir channel inhibitors as insecticides.

  10. Dual effect of ethanol on inward rectifier potassium current IK1 in rat ventricular myocytes

    Czech Academy of Sciences Publication Activity Database

    Bébarová, M.; Matejovič, P.; Pásek, Michal; Šimurdová, M.; Šimurda, J.

    2014-01-01

    Roč. 65, č. 4 (2014), s. 497-509 ISSN 0867-5910 Grant - others:GA MZd NT14301 Institutional support: RVO:61388998 Keywords : ethanol * rat ventricular myocyte * rat ventricular action potential model Subject RIV: BO - Biophysics Impact factor: 2.386, year: 2014

  11. Modeling removal of accumulated potassium from T-tubules by inward rectifier potassium channels

    NARCIS (Netherlands)

    Wallinga, W.; Vliek, M.; Wienk, E.D.; Alberink, M.J.; Ypey, D.L.; Ypey, D.L.

    1996-01-01

    The membrane models of Cannon et al. (1993) and Alberink et al. (1995) for mammalian skeletal muscle fibers are based upon Hodgkin-Huxley descriptions of sodium, potassium delayed rectifier and leak conductances and the capacitive current taking into account fast inactivation of sodium channels. Now

  12. Autophagy in unicellular eukaryotes

    NARCIS (Netherlands)

    Kiel, J.A.K.W.

    2010-01-01

    Cells need a constant supply of precursors to enable the production of macromolecules to sustain growth and survival. Unlike metazoans, unicellular eukaryotes depend exclusively on the extracellular medium for this supply. When environmental nutrients become depleted, existing cytoplasmic components

  13. Eukaryotic Cell Panorama

    Science.gov (United States)

    Goodsell, David S.

    2011-01-01

    Diverse biological data may be used to create illustrations of molecules in their cellular context. This report describes the scientific results that support an illustration of a eukaryotic cell, enlarged by one million times to show the distribution and arrangement of macromolecules. The panoramic cross section includes eight panels that extend…

  14. Eukaryotic cell flattening

    Science.gov (United States)

    Bae, Albert; Westendorf, Christian; Erlenkamper, Christoph; Galland, Edouard; Franck, Carl; Bodenschatz, Eberhard; Beta, Carsten

    2010-03-01

    Eukaryotic cell flattening is valuable for improving microscopic observations, ranging from bright field to total internal reflection fluorescence microscopy. In this talk, we will discuss traditional overlay techniques, and more modern, microfluidic based flattening, which provides a greater level of control. We demonstrate these techniques on the social amoebae Dictyostelium discoideum, comparing the advantages and disadvantages of each method.

  15. Comparative Genomics of Eukaryotes.

    NARCIS (Netherlands)

    Noort, V. van

    2007-01-01

    This thesis focuses on developing comparative genomics methods in eukaryotes, with an emphasis on applications for gene function prediction and regulatory element detection. In the past, methods have been developed to predict functional associations between gene pairs in prokaryotes. The challenge

  16. DNA to DNA transcription might exist in eukaryotic cells

    OpenAIRE

    Li, Gao-De

    2016-01-01

    Till now, in biological sciences, the term, transcription, mainly refers to DNA to RNA transcription. But our recently published experimental findings obtained from Plasmodium falciparum strongly suggest the existence of DNA to DNA transcription in the genome of eukaryotic cells, which could shed some light on the functions of certain noncoding DNA in the human and other eukaryotic genomes.

  17. 5-HT modulation of multiple inward rectifiers in motoneurons in intact preparations of the neonatal rat spinal cord

    DEFF Research Database (Denmark)

    Kjaerulff, Ole; Kiehn, Ole

    2001-01-01

    This study introduces novel aspects of inward rectification in neonatal rat spinal motoneurons (MNs) and its modulation by serotonin (5-HT). Whole cell tight-seal recordings were made from MNs in an isolated lumbar spinal cord preparation from rats 1-2 days of age. In voltage clamp, hyperpolarizi...

  18. Structural and functional determinants of conserved lipid interaction domains of inward rectifying Kir6.2 channels.

    Science.gov (United States)

    Cukras, Catherine A; Jeliazkova, Iana; Nichols, Colin G

    2002-06-01

    All members of the inward rectifiier K(+) (Kir) channel family are activated by phosphoinositides and other amphiphilic lipids. To further elucidate the mechanistic basis, we examined the membrane association of Kir6.2 fragments of K(ATP) channels, and the effects of site-directed mutations of these fragments and full-length Kir6.2 on membrane association and K(ATP) channel activity, respectively. GFP-tagged Kir6.2 COOH terminus and GFP-tagged pleckstrin homology domain from phospholipase C delta1 both associate with isolated membranes, and association of each is specifically reduced by muscarinic m1 receptor-mediated phospholipid depletion. Kir COOH termini are predicted to contain multiple beta-strands and a conserved alpha-helix (residues approximately 306-311 in Kir6.2). Systematic mutagenesis of D307-F315 reveals a critical role of E308, I309, W311 and F315, consistent with residues lying on one side of a alpha-helix. Together with systematic mutation of conserved charges, the results define critical determinants of a conserved domain that underlies phospholipid interaction in Kir channels.

  19. Eukaryotic DNA Replication Fork.

    Science.gov (United States)

    Burgers, Peter M J; Kunkel, Thomas A

    2017-06-20

    This review focuses on the biogenesis and composition of the eukaryotic DNA replication fork, with an emphasis on the enzymes that synthesize DNA and repair discontinuities on the lagging strand of the replication fork. Physical and genetic methodologies aimed at understanding these processes are discussed. The preponderance of evidence supports a model in which DNA polymerase ε (Pol ε) carries out the bulk of leading strand DNA synthesis at an undisturbed replication fork. DNA polymerases α and δ carry out the initiation of Okazaki fragment synthesis and its elongation and maturation, respectively. This review also discusses alternative proposals, including cellular processes during which alternative forks may be utilized, and new biochemical studies with purified proteins that are aimed at reconstituting leading and lagging strand DNA synthesis separately and as an integrated replication fork.

  20. Morphological and ecological complexity in early eukaryotic ecosystems.

    Science.gov (United States)

    Javaux, E J; Knoll, A H; Walter, M R

    2001-07-05

    Molecular phylogeny and biogeochemistry indicate that eukaryotes differentiated early in Earth history. Sequence comparisons of small-subunit ribosomal RNA genes suggest a deep evolutionary divergence of Eukarya and Archaea; C27-C29 steranes (derived from sterols synthesized by eukaryotes) and strong depletion of 13C (a biogeochemical signature of methanogenic Archaea) in 2,700 Myr old kerogens independently place a minimum age on this split. Steranes, large spheroidal microfossils, and rare macrofossils of possible eukaryotic origin occur in Palaeoproterozoic rocks. Until now, however, evidence for morphological and taxonomic diversification within the domain has generally been restricted to very late Mesoproterozoic and Neoproterozoic successions. Here we show that the cytoskeletal and ecological prerequisites for eukaryotic diversification were already established in eukaryotic microorganisms fossilized nearly 1,500 Myr ago in shales of the early Mesoproterozoic Roper Group in northern Australia.

  1. Eukaryotic DNA Replicases

    KAUST Repository

    Zaher, Manal S.; Oke, Muse; Hamdan, Samir

    2014-01-01

    The current model of the eukaryotic DNA replication fork includes three replicative DNA polymerases, polymerase α/primase complex (Pol α), polymerase δ (Pol δ), and polymerase ε (Pol ε). The primase synthesizes 8–12 nucleotide RNA primers that are extended by the DNA polymerization activity of Pol α into 30–35 nucleotide RNA-DNA primers. Replication factor C (RFC) opens the polymerase clamp-like processivity factor, proliferating cell nuclear antigen (PCNA), and loads it onto the primer-template. Pol δ utilizes PCNA to mediate highly processive DNA synthesis, while Pol ε has intrinsic high processivity that is modestly stimulated by PCNA. Pol ε replicates the leading strand and Pol δ replicates the lagging strand in a division of labor that is not strict. The three polymerases are comprised of multiple subunits and share unifying features in their large catalytic and B subunits. The remaining subunits are evolutionarily not related and perform diverse functions. The catalytic subunits are members of family B, which are distinguished by their larger sizes due to inserts in their N- and C-terminal regions. The sizes of these inserts vary among the three polymerases, and their functions remain largely unknown. Strikingly, the quaternary structures of Pol α, Pol δ, and Pol ε are arranged similarly. The catalytic subunits adopt a globular structure that is linked via its conserved C-terminal region to the B subunit. The remaining subunits are linked to the catalytic and B subunits in a highly flexible manner.

  2. Eukaryotic DNA Replicases

    KAUST Repository

    Zaher, Manal S.

    2014-11-21

    The current model of the eukaryotic DNA replication fork includes three replicative DNA polymerases, polymerase α/primase complex (Pol α), polymerase δ (Pol δ), and polymerase ε (Pol ε). The primase synthesizes 8–12 nucleotide RNA primers that are extended by the DNA polymerization activity of Pol α into 30–35 nucleotide RNA-DNA primers. Replication factor C (RFC) opens the polymerase clamp-like processivity factor, proliferating cell nuclear antigen (PCNA), and loads it onto the primer-template. Pol δ utilizes PCNA to mediate highly processive DNA synthesis, while Pol ε has intrinsic high processivity that is modestly stimulated by PCNA. Pol ε replicates the leading strand and Pol δ replicates the lagging strand in a division of labor that is not strict. The three polymerases are comprised of multiple subunits and share unifying features in their large catalytic and B subunits. The remaining subunits are evolutionarily not related and perform diverse functions. The catalytic subunits are members of family B, which are distinguished by their larger sizes due to inserts in their N- and C-terminal regions. The sizes of these inserts vary among the three polymerases, and their functions remain largely unknown. Strikingly, the quaternary structures of Pol α, Pol δ, and Pol ε are arranged similarly. The catalytic subunits adopt a globular structure that is linked via its conserved C-terminal region to the B subunit. The remaining subunits are linked to the catalytic and B subunits in a highly flexible manner.

  3. DNA mismatch repair and its many roles in eukaryotic cells

    DEFF Research Database (Denmark)

    Liu, Dekang; Keijzers, Guido; Rasmussen, Lene Juel

    2017-01-01

    in the clinic, and as a biomarker of cancer susceptibility in animal model systems. Prokaryotic MMR is well-characterized at the molecular and mechanistic level; however, MMR is considerably more complex in eukaryotic cells than in prokaryotic cells, and in recent years, it has become evident that MMR plays...... novel roles in eukaryotic cells, several of which are not yet well-defined or understood. Many MMR-deficient human cancer cells lack mutations in known human MMR genes, which strongly suggests that essential eukaryotic MMR components/cofactors remain unidentified and uncharacterized. Furthermore......, the mechanism by which the eukaryotic MMR machinery discriminates between the parental (template) and the daughter (nascent) DNA strand is incompletely understood and how cells choose between the EXO1-dependent and the EXO1–independent subpathways of MMR is not known. This review summarizes recent literature...

  4. Gonococcal attachment to eukaryotic cells

    International Nuclear Information System (INIS)

    James, J.F.; Lammel, C.J.; Draper, D.L.; Brown, D.A.; Sweet, R.L.; Brooks, G.F.

    1983-01-01

    The attachment of Neisseria gonorrhoeae to eukaryotic cells grown in tissue culture was analyzed by use of light and electron microscopy and by labeling of the bacteria with [ 3 H]- and [ 14 C]adenine. Isogenic piliated and nonpiliated N. gonorrhoeae from opaque and transparent colonies were studied. The results of light microscopy studies showed that the gonococci attached to cells of human origin, including Flow 2000, HeLa 229, and HEp 2. Studies using radiolabeled gonococci gave comparable results. Piliated N. gonorrhoeae usually attached in larger numbers than nonpiliated organisms, and those from opaque colonies attached more often than isogenic variants from transparent colonies. Day-to-day variation in rate of attachment was observed. Scanning electron microscopy studies showed the gonococcal attachment to be specific for microvilli of the host cells. It is concluded that more N. gonorrhoeae from opaque colonies, as compared with isogenic variants from transparent colonies, attach to eukaryotic cells grown in tissue culture

  5. Defensins: antifungal lessons from eukaryotes

    Directory of Open Access Journals (Sweden)

    Patrícia M. Silva

    2014-03-01

    Full Text Available Over the last years, antimicrobial peptides (AMPs have been the focus of intense research towards the finding of a viable alternative to current antifungal drugs. Defensins are one of the major families of AMPs and the most represented among all eukaryotic groups, providing an important first line of host defense against pathogenic microorganisms. Several of these cysteine-stabilized peptides present a relevant effect against fungi. Defensins are the AMPs with the broader distribution across all eukaryotic kingdoms, namely, Fungi, Plantæ and Animalia, and were recently shown to have an ancestor in a bacterial organism. As a part of the host defense, defensins act as an important vehicle of information between innate and adaptive immune system and have a role in immunomodulation. This multidimensionality represents a powerful host shield, hard for microorganisms to overcome using single approach resistance strategies. Pathogenic fungi resistance to conventional antimycotic drugs is becoming a major problem. Defensins, as other AMPs, have shown to be an effective alternative to the current antimycotic therapies, demonstrating potential as novel therapeutic agents or drug leads. In this review, we summarize the current knowledge on some eukaryotic defensins with antifungal action. An overview of the main targets in the fungal cell and the mechanism of action of these AMPs (namely, the selectivity for some fungal membrane components are presented. Additionally, recent works on antifungal defensins structure, activity and citotoxicity are also reviewed.

  6. Archaeal “Dark Matter” and the Origin of Eukaryotes

    Science.gov (United States)

    Williams, Tom A.; Embley, T. Martin

    2014-01-01

    Current hypotheses about the history of cellular life are mainly based on analyses of cultivated organisms, but these represent only a small fraction of extant biodiversity. The sequencing of new environmental lineages therefore provides an opportunity to test, revise, or reject existing ideas about the tree of life and the origin of eukaryotes. According to the textbook three domains hypothesis, the eukaryotes emerge as the sister group to a monophyletic Archaea. However, recent analyses incorporating better phylogenetic models and an improved sampling of the archaeal domain have generally supported the competing eocyte hypothesis, in which core genes of eukaryotic cells originated from within the Archaea, with important implications for eukaryogenesis. Given this trend, it was surprising that a recent analysis incorporating new genomes from uncultivated Archaea recovered a strongly supported three domains tree. Here, we show that this result was due in part to the use of a poorly fitting phylogenetic model and also to the inclusion by an automated pipeline of genes of putative bacterial origin rather than nucleocytosolic versions for some of the eukaryotes analyzed. When these issues were resolved, analyses including the new archaeal lineages placed core eukaryotic genes within the Archaea. These results are consistent with a number of recent studies in which improved archaeal sampling and better phylogenetic models agree in supporting the eocyte tree over the three domains hypothesis. PMID:24532674

  7. Genetic variation in the inwardly rectifying K channel subunits KCNJ3 (GIRK1) and KCNJ5 (GIRK4) in patients with sinus node dysfunction

    DEFF Research Database (Denmark)

    Holmegard, Haya N; Theilade, Juliane; Benn, Marianne

    2010-01-01

    BACKGROUND: Sinus node dysfunction (SND) is a heterogeneous disorder of unknown etiology characterized by a variety of supraventricular arrhythmias with symptoms of syncope, palpitations, and dizziness. The mechanism underlying the abnormal rhythm is incompletely understood. OBJECTIVE: Because va...

  8. How natural a kind is "eukaryote?".

    Science.gov (United States)

    Doolittle, W Ford

    2014-06-02

    Systematics balances uneasily between realism and nominalism, uncommitted as to whether biological taxa are discoveries or inventions. If the former, they might be taken as natural kinds. I briefly review some philosophers' concepts of natural kinds and then argue that several of these apply well enough to "eukaryote." Although there are some sticky issues around genomic chimerism and when eukaryotes first appeared, if we allow for degrees in the naturalness of kinds, existing eukaryotes rank highly, higher than prokaryotes. Most biologists feel this intuitively: All I attempt to do here is provide some conceptual justification. Copyright © 2014 Cold Spring Harbor Laboratory Press; all rights reserved.

  9. Arabinogalactan proteins have deep roots in eukaryotes

    DEFF Research Database (Denmark)

    Hervé, Cécile; Siméon, Amandine; Jam, Murielle

    2016-01-01

    Arabinogalactan proteins (AGPs) are highly glycosylated, hydroxyproline-rich proteins found at the cell surface of plants, where they play key roles in developmental processes. Brown algae are marine, multicellular, photosynthetic eukaryotes. They belong to the phylum Stramenopiles, which...

  10. Transfer of DNA from Bacteria to Eukaryotes

    Directory of Open Access Journals (Sweden)

    Benoît Lacroix

    2016-07-01

    Full Text Available Historically, the members of the Agrobacterium genus have been considered the only bacterial species naturally able to transfer and integrate DNA into the genomes of their eukaryotic hosts. Yet, increasing evidence suggests that this ability to genetically transform eukaryotic host cells might be more widespread in the bacterial world. Indeed, analyses of accumulating genomic data reveal cases of horizontal gene transfer from bacteria to eukaryotes and suggest that it represents a significant force in adaptive evolution of eukaryotic species. Specifically, recent reports indicate that bacteria other than Agrobacterium, such as Bartonella henselae (a zoonotic pathogen, Rhizobium etli (a plant-symbiotic bacterium related to Agrobacterium, or even Escherichia coli, have the ability to genetically transform their host cells under laboratory conditions. This DNA transfer relies on type IV secretion systems (T4SSs, the molecular machines that transport macromolecules during conjugative plasmid transfer and also during transport of proteins and/or DNA to the eukaryotic recipient cells. In this review article, we explore the extent of possible transfer of genetic information from bacteria to eukaryotic cells as well as the evolutionary implications and potential applications of this transfer.

  11. Horizontal transfer of a eukaryotic plastid-targeted protein gene to cyanobacteria

    Directory of Open Access Journals (Sweden)

    Keeling Patrick J

    2007-06-01

    Full Text Available Abstract Background Horizontal or lateral transfer of genetic material between distantly related prokaryotes has been shown to play a major role in the evolution of bacterial and archaeal genomes, but exchange of genes between prokaryotes and eukaryotes is not as well understood. In particular, gene flow from eukaryotes to prokaryotes is rarely documented with strong support, which is unusual since prokaryotic genomes appear to readily accept foreign genes. Results Here, we show that abundant marine cyanobacteria in the related genera Synechococcus and Prochlorococcus acquired a key Calvin cycle/glycolytic enzyme from a eukaryote. Two non-homologous forms of fructose bisphosphate aldolase (FBA are characteristic of eukaryotes and prokaryotes respectively. However, a eukaryotic gene has been inserted immediately upstream of the ancestral prokaryotic gene in several strains (ecotypes of Synechococcus and Prochlorococcus. In one lineage this new gene has replaced the ancestral gene altogether. The eukaryotic gene is most closely related to the plastid-targeted FBA from red algae. This eukaryotic-type FBA once replaced the plastid/cyanobacterial type in photosynthetic eukaryotes, hinting at a possible functional advantage in Calvin cycle reactions. The strains that now possess this eukaryotic FBA are scattered across the tree of Synechococcus and Prochlorococcus, perhaps because the gene has been transferred multiple times among cyanobacteria, or more likely because it has been selectively retained only in certain lineages. Conclusion A gene for plastid-targeted FBA has been transferred from red algae to cyanobacteria, where it has inserted itself beside its non-homologous, functional analogue. Its current distribution in Prochlorococcus and Synechococcus is punctate, suggesting a complex history since its introduction to this group.

  12. Atypical mitochondrial inheritance patterns in eukaryotes.

    Science.gov (United States)

    Breton, Sophie; Stewart, Donald T

    2015-10-01

    Mitochondrial DNA (mtDNA) is predominantly maternally inherited in eukaryotes. Diverse molecular mechanisms underlying the phenomenon of strict maternal inheritance (SMI) of mtDNA have been described, but the evolutionary forces responsible for its predominance in eukaryotes remain to be elucidated. Exceptions to SMI have been reported in diverse eukaryotic taxa, leading to the prediction that several distinct molecular mechanisms controlling mtDNA transmission are present among the eukaryotes. We propose that these mechanisms will be better understood by studying the deviations from the predominating pattern of SMI. This minireview summarizes studies on eukaryote species with unusual or rare mitochondrial inheritance patterns, i.e., other than the predominant SMI pattern, such as maternal inheritance of stable heteroplasmy, paternal leakage of mtDNA, biparental and strictly paternal inheritance, and doubly uniparental inheritance of mtDNA. The potential genes and mechanisms involved in controlling mitochondrial inheritance in these organisms are discussed. The linkage between mitochondrial inheritance and sex determination is also discussed, given that the atypical systems of mtDNA inheritance examined in this minireview are frequently found in organisms with uncommon sexual systems such as gynodioecy, monoecy, or andromonoecy. The potential of deviations from SMI for facilitating a better understanding of a number of fundamental questions in biology, such as the evolution of mtDNA inheritance, the coevolution of nuclear and mitochondrial genomes, and, perhaps, the role of mitochondria in sex determination, is considerable.

  13. Comparative genomics and evolution of eukaryotic phospholipidbiosynthesis

    Energy Technology Data Exchange (ETDEWEB)

    Lykidis, Athanasios

    2006-12-01

    Phospholipid biosynthetic enzymes produce diverse molecular structures and are often present in multiple forms encoded by different genes. This work utilizes comparative genomics and phylogenetics for exploring the distribution, structure and evolution of phospholipid biosynthetic genes and pathways in 26 eukaryotic genomes. Although the basic structure of the pathways was formed early in eukaryotic evolution, the emerging picture indicates that individual enzyme families followed unique evolutionary courses. For example, choline and ethanolamine kinases and cytidylyltransferases emerged in ancestral eukaryotes, whereas, multiple forms of the corresponding phosphatidyltransferases evolved mainly in a lineage specific manner. Furthermore, several unicellular eukaryotes maintain bacterial-type enzymes and reactions for the synthesis of phosphatidylglycerol and cardiolipin. Also, base-exchange phosphatidylserine synthases are widespread and ancestral enzymes. The multiplicity of phospholipid biosynthetic enzymes has been largely generated by gene expansion in a lineage specific manner. Thus, these observations suggest that phospholipid biosynthesis has been an actively evolving system. Finally, comparative genomic analysis indicates the existence of novel phosphatidyltransferases and provides a candidate for the uncharacterized eukaryotic phosphatidylglycerol phosphate phosphatase.

  14. Repair of DNA DSB in higher eukaryotes

    International Nuclear Information System (INIS)

    Wang, H.; Perrault, A.R.; Takeda, Y.; Iliakis, G.

    2003-01-01

    Cells of higher eukaryotes process within minutes double strand breaks (DSBs) in their genome using a NHEJ apparatus that engages DNA-PKcs, Ku, DNA ligase IV, XRCC4, and other as of yet unidentified factors. Although chemical inhibition, or mutation, in any of these factors delays processing, cells ultimately remove the majority of DNA DSBs using an alternative pathway operating with slower kinetics. This alternative pathway is active in mutants deficient in genes of the RAD52 epistasis group. We proposed, therefore, that it reflects an alternative form of NHEJ that operates as a backup (B-NHEJ) to the DNA-PK- dependent (D-NHEJ) pathway, rather than homology directed repair of DSBs. We studied the role of Ku and DNA-PKcs in the coordination of these pathways using as a model end joining of restriction endonuclease linearized plasmid DNA in whole cell extracts. Efficient error-free endjoining observed in such in-vitro reactions is strongly inhibited by anti-Ku antibodies. The inhibition requires DNA-PKcs, despite that fact that Ku efficiently binds DNA ends in the presence of antibodies, or in the absence of DNA-PKcs. Strong inhibition of DNA endjoining is also mediated by wortmannin, an inhibitor of DNA-PKcs, in the presence but not in the absence of Ku, and this inhibition can be rescued by pre-incubating the reaction with double stranded oligonucleotides. The results are compatible with a role of Ku in directing endjoining to a DNA-PK dependent pathway, mediated by efficient end binding and productive interactions with DNA-PKcs. On the other hand, efficient end joining is observed in extracts of cells lacking DNA-PKcs, as well as in Ku-depleted extracts sugggesting the operation of alternative pathways. Extracts depleted of Ku and DNA-PKcs rejoin blunt ends, as well as homologous ends with 3' or 5' protruding single strands with similar efficiency, but addition of Ku suppresses joining of blunt ends and homologous ends with 3' overhangs. We propose that the

  15. Communities of microbial eukaryotes in the mammalian gut within the context of environmental eukaryotic diversity

    Energy Technology Data Exchange (ETDEWEB)

    Parfrey, Laura Wegener; Walters, William A.; Lauber, Christian L.; Clemente, Jose C.; Berg-Lyons, Donna; Teiling, Clotilde; Kodira, Chinnappa; Mohiuddin, Mohammed; Brunelle, Julie; Driscoll, Mark; Fierer, Noah; Gilbert, Jack A.; Knight, Rob

    2014-06-19

    Eukaryotic microbes (protists) residing in the vertebrate gut influence host health and disease, but their diversity and distribution in healthy hosts is poorly understood. Protists found in the gut are typically considered parasites, but many are commensal and some are beneficial. Further, the hygiene hypothesis predicts that association with our co-evolved microbial symbionts may be important to overall health. It is therefore imperative that we understand the normal diversity of our eukaryotic gut microbiota to test for such effects and avoid eliminating commensal organisms. We assembled a dataset of healthy individuals from two populations, one with traditional, agrarian lifestyles and a second with modern, westernized lifestyles, and characterized the human eukaryotic microbiota via high-throughput sequencing. To place the human gut microbiota within a broader context our dataset also includes gut samples from diverse mammals and samples from other aquatic and terrestrial environments. We curated the SILVA ribosomal database to reflect current knowledge of eukaryotic taxonomy and employ it as a phylogenetic framework to compare eukaryotic diversity across environment. We show that adults from the non-western population harbor a diverse community of protists, and diversity in the human gut is comparable to that in other mammals. However, the eukaryotic microbiota of the western population appears depauperate. The distribution of symbionts found in mammals reflects both host phylogeny and diet. Eukaryotic microbiota in the gut are less diverse and more patchily distributed than bacteria. More broadly, we show that eukaryotic communities in the gut are less diverse than in aquatic and terrestrial habitats, and few taxa are shared across habitat types, and diversity patterns of eukaryotes are correlated with those observed for bacteria. These results outline the distribution and diversity of microbial eukaryotic communities in the mammalian gut and across

  16. Eukaryotes first: how could that be?

    Science.gov (United States)

    Mariscal, Carlos; Doolittle, W Ford

    2015-09-26

    In the half century since the formulation of the prokaryote : eukaryote dichotomy, many authors have proposed that the former evolved from something resembling the latter, in defiance of common (and possibly common sense) views. In such 'eukaryotes first' (EF) scenarios, the last universal common ancestor is imagined to have possessed significantly many of the complex characteristics of contemporary eukaryotes, as relics of an earlier 'progenotic' period or RNA world. Bacteria and Archaea thus must have lost these complex features secondarily, through 'streamlining'. If the canonical three-domain tree in which Archaea and Eukarya are sisters is accepted, EF entails that Bacteria and Archaea are convergently prokaryotic. We ask what this means and how it might be tested. © 2015 The Author(s).

  17. Reproduction, symbiosis, and the eukaryotic cell

    Science.gov (United States)

    Godfrey-Smith, Peter

    2015-01-01

    This paper develops a conceptual framework for addressing questions about reproduction, individuality, and the units of selection in symbiotic associations, with special attention to the origin of the eukaryotic cell. Three kinds of reproduction are distinguished, and a possible evolutionary sequence giving rise to a mitochondrion-containing eukaryotic cell from an endosymbiotic partnership is analyzed as a series of transitions between each of the three forms of reproduction. The sequence of changes seen in this “egalitarian” evolutionary transition is compared with those that apply in “fraternal” transitions, such as the evolution of multicellularity in animals. PMID:26286983

  18. The origin of the eukaryotic cell

    Science.gov (United States)

    Hartman, H.

    1984-01-01

    The endosymbiotic hypothesis for the origin of the eukaryotic cell has been applied to the origin of the mitochondria and chloroplasts. However as has been pointed out by Mereschowsky in 1905, it should also be applied to the nucleus as well. If the nucleus, mitochondria and chloroplasts are endosymbionts, then it is likely that the organism that did the engulfing was not a DNA-based organism. In fact, it is useful to postulate that this organism was a primitive RNA-based organism. This hypothesis would explain the preponderance of RNA viruses found in eukaryotic cells. The centriole and basal body do not have a double membrane or DNA. Like all MTOCs (microtubule organising centres), they have a structural or morphic RNA implicated in their formation. This would argue for their origin in the early RNA-based organism rather than in an endosymbiotic event involving bacteria. Finally, the eukaryotic cell uses RNA in ways quite unlike bacteria, thus pointing to a greater emphasis of RNA in both control and structure in the cell. The origin of the eukaryotic cell may tell us why it rather than its prokaryotic relative evolved into the metazoans who are reading this paper.

  19. Eukaryotic acquisition of a bacterial operon

    Science.gov (United States)

    The yeast Saccharomyces cerevisiae is one of the champions of basic biomedical research due to its compact eukaryotic genome and ease of experimental manipulation. Despite these immense strengths, its impact on understanding the genetic basis of natural phenotypic variation has been limited by strai...

  20. Anaerobic energy metabolism in unicellular photosynthetic eukaryotes.

    Science.gov (United States)

    Atteia, Ariane; van Lis, Robert; Tielens, Aloysius G M; Martin, William F

    2013-02-01

    Anaerobic metabolic pathways allow unicellular organisms to tolerate or colonize anoxic environments. Over the past ten years, genome sequencing projects have brought a new light on the extent of anaerobic metabolism in eukaryotes. A surprising development has been that free-living unicellular algae capable of photoautotrophic lifestyle are, in terms of their enzymatic repertoire, among the best equipped eukaryotes known when it comes to anaerobic energy metabolism. Some of these algae are marine organisms, common in the oceans, others are more typically soil inhabitants. All these species are important from the ecological (O(2)/CO(2) budget), biotechnological, and evolutionary perspectives. In the unicellular algae surveyed here, mixed-acid type fermentations are widespread while anaerobic respiration, which is more typical of eukaryotic heterotrophs, appears to be rare. The presence of a core anaerobic metabolism among the algae provides insights into its evolutionary origin, which traces to the eukaryote common ancestor. The predicted fermentative enzymes often exhibit an amino acid extension at the N-terminus, suggesting that these proteins might be compartmentalized in the cell, likely in the chloroplast or the mitochondrion. The green algae Chlamydomonas reinhardtii and Chlorella NC64 have the most extended set of fermentative enzymes reported so far. Among the eukaryotes with secondary plastids, the diatom Thalassiosira pseudonana has the most pronounced anaerobic capabilities as yet. From the standpoints of genomic, transcriptomic, and biochemical studies, anaerobic energy metabolism in C. reinhardtii remains the best characterized among photosynthetic protists. This article is part of a Special Issue entitled: The evolutionary aspects of bioenergetic systems. Copyright © 2012 Elsevier B.V. All rights reserved.

  1. Symbiosis and the origin of eukaryotic motility

    Science.gov (United States)

    Margulis, L.; Hinkle, G.

    1991-01-01

    Ongoing work to test the hypothesis of the origin of eukaryotic cell organelles by microbial symbioses is discussed. Because of the widespread acceptance of the serial endosymbiotic theory (SET) of the origin of plastids and mitochondria, the idea of the symbiotic origin of the centrioles and axonemes for spirochete bacteria motility symbiosis was tested. Intracellular microtubular systems are purported to derive from symbiotic associations between ancestral eukaryotic cells and motile bacteria. Four lines of approach to this problem are being pursued: (1) cloning the gene of a tubulin-like protein discovered in Spirocheata bajacaliforniesis; (2) seeking axoneme proteins in spirochets by antibody cross-reaction; (3) attempting to cultivate larger, free-living spirochetes; and (4) studying in detail spirochetes (e.g., Cristispira) symbiotic with marine animals. Other aspects of the investigation are presented.

  2. Towards New Antifolates Targeting Eukaryotic Opportunistic Infections

    Energy Technology Data Exchange (ETDEWEB)

    Liu, J.; Bolstad, D; Bolstad, E; Wright, D; Anderson, A

    2009-01-01

    Trimethoprim, an antifolate commonly prescribed in combination with sulfamethoxazole, potently inhibits several prokaryotic species of dihydrofolate reductase (DHFR). However, several eukaryotic pathogenic organisms are resistant to trimethoprim, preventing its effective use as a therapeutic for those infections. We have been building a program to reengineer trimethoprim to more potently and selectively inhibit eukaryotic species of DHFR as a viable strategy for new drug discovery targeting several opportunistic pathogens. We have developed a series of compounds that exhibit potent and selective inhibition of DHFR from the parasitic protozoa Cryptosporidium and Toxoplasma as well as the fungus Candida glabrata. A comparison of the structures of DHFR from the fungal species Candida glabrata and Pneumocystis suggests that the compounds may also potently inhibit Pneumocystis DHFR.

  3. Inorganic phosphate uptake in unicellular eukaryotes.

    Science.gov (United States)

    Dick, Claudia F; Dos-Santos, André L A; Meyer-Fernandes, José R

    2014-07-01

    Inorganic phosphate (Pi) is an essential nutrient for all organisms. The route of Pi utilization begins with Pi transport across the plasma membrane. Here, we analyzed the gene sequences and compared the biochemical profiles, including kinetic and modulator parameters, of Pi transporters in unicellular eukaryotes. The objective of this review is to evaluate the recent findings regarding Pi uptake mechanisms in microorganisms, such as the fungi Neurospora crassa and Saccharomyces cerevisiae and the parasite protozoans Trypanosoma cruzi, Trypanosoma rangeli, Leishmania infantum and Plasmodium falciparum. Pi uptake is the key step of Pi homeostasis and in the subsequent signaling event in eukaryotic microorganisms. Biochemical and structural studies are important for clarifying mechanisms of Pi homeostasis, as well as Pi sensor and downstream pathways, and raise possibilities for future studies in this field. Copyright © 2014 Elsevier B.V. All rights reserved.

  4. In silico ionomics segregates parasitic from free-living eukaryotes.

    Science.gov (United States)

    Greganova, Eva; Steinmann, Michael; Mäser, Pascal; Fankhauser, Niklaus

    2013-01-01

    Ion transporters are fundamental to life. Due to their ancient origin and conservation in sequence, ion transporters are also particularly well suited for comparative genomics of distantly related species. Here, we perform genome-wide ion transporter profiling as a basis for comparative genomics of eukaryotes. From a given predicted proteome, we identify all bona fide ion channels, ion porters, and ion pumps. Concentrating on unicellular eukaryotes (n = 37), we demonstrate that clustering of species according to their repertoire of ion transporters segregates obligate endoparasites (n = 23) on the one hand, from free-living species and facultative parasites (n = 14) on the other hand. This surprising finding indicates strong convergent evolution of the parasites regarding the acquisition and homeostasis of inorganic ions. Random forest classification identifies transporters of ammonia, plus transporters of iron and other transition metals, as the most informative for distinguishing the obligate parasites. Thus, in silico ionomics further underscores the importance of iron in infection biology and suggests access to host sources of nitrogen and transition metals to be selective forces in the evolution of parasitism. This finding is in agreement with the phenomenon of iron withholding as a primordial antimicrobial strategy of infected mammals.

  5. Characterization of an eukaryotic peptide deformylase from Plasmodium falciparum.

    Science.gov (United States)

    Bracchi-Ricard, V; Nguyen, K T; Zhou, Y; Rajagopalan, P T; Chakrabarti, D; Pei, D

    2001-12-15

    Ribosomal protein synthesis in eubacteria and eukaryotic organelles initiates with an N-formylmethionyl-tRNA(i), resulting in N-terminal formylation of all nascent polypeptides. Peptide deformylase (PDF) catalyzes the subsequent removal of the N-terminal formyl group from the majority of bacterial proteins. Until recently, PDF has been thought as an enzyme unique to the bacterial kingdom. Searches of the genomic DNA databases identified several genes that encode proteins of high sequence homology to bacterial PDF from eukaryotic organisms. The cDNA encoding Plasmodium falciparum PDF (PfPDF) has been cloned and overexpressed in Escherichia coli. The recombinant protein is catalytically active in deformylating N-formylated peptides, shares many of the properties of bacterial PDF, and is inhibited by specific PDF inhibitors. Western blot analysis indicated expression of mature PfPDF in trophozoite, schizont, and segmenter stages of intraerythrocytic development. These results provide strong evidence that a functional PDF is present in P. falciparum. In addition, PDF inhibitors inhibited the growth of P. falciparum in the intraerythrocytic culture. (c)2001 Elsevier Science.

  6. Enzymes from Higher Eukaryotes for Industrial Biocatalysis

    Directory of Open Access Journals (Sweden)

    Zhibin Liu

    2004-01-01

    Full Text Available The industrial production of fine chemicals, feed and food ingredients, pharmaceuticals, agrochemicals and their respective intermediates relies on an increasing application of biocatalysis, i.e. on enzyme or whole-cell catalyzed conversions of molecules. Simple procedures for discovery, cloning and over-expression as well as fast growth favour fungi, yeasts and especially bacteria as sources of biocatalysts. Higher eukaryotes also harbour an almost unlimited number of potential biocatalysts, although to date the limited supply of enzymes, the high heterogeneity of enzyme preparations and the hazard of infectious contaminants keep some interesting candidates out of reach for industrial bioprocesses. In the past only a few animal and plant enzymes from agricultural waste materials were employed in food processing. The use of bacterial expression strains or non-conventional yeasts for the heterologous production of efficient eukaryotic enzymes can overcome the bottleneck in enzyme supply and provide sufficient amounts of homogenous enzyme preparations for reliable and economically feasible applications at large scale. Ideal enzymatic processes represent an environmentally friendly, »near-to-completion« conversion of (mostly non-natural substrates to pure products. Recent developments demonstrate the commercial feasibility of large-scale biocatalytic processes employing enzymes from higher eukaryotes (e.g. plants, animals and also their usefulness in some small-scale industrial applications.

  7. Arsenic and Antimony Transporters in Eukaryotes

    Directory of Open Access Journals (Sweden)

    Ewa Maciaszczyk-Dziubinska

    2012-03-01

    Full Text Available Arsenic and antimony are toxic metalloids, naturally present in the environment and all organisms have developed pathways for their detoxification. The most effective metalloid tolerance systems in eukaryotes include downregulation of metalloid uptake, efflux out of the cell, and complexation with phytochelatin or glutathione followed by sequestration into the vacuole. Understanding of arsenic and antimony transport system is of high importance due to the increasing usage of arsenic-based drugs in the treatment of certain types of cancer and diseases caused by protozoan parasites as well as for the development of bio- and phytoremediation strategies for metalloid polluted areas. However, in contrast to prokaryotes, the knowledge about specific transporters of arsenic and antimony and the mechanisms of metalloid transport in eukaryotes has been very limited for a long time. Here, we review the recent advances in understanding of arsenic and antimony transport pathways in eukaryotes, including a dual role of aquaglyceroporins in uptake and efflux of metalloids, elucidation of arsenic transport mechanism by the yeast Acr3 transporter and its role in arsenic hyperaccumulation in ferns, identification of vacuolar transporters of arsenic-phytochelatin complexes in plants and forms of arsenic substrates recognized by mammalian ABC transporters.

  8. Arsenic and Antimony Transporters in Eukaryotes

    Science.gov (United States)

    Maciaszczyk-Dziubinska, Ewa; Wawrzycka, Donata; Wysocki, Robert

    2012-01-01

    Arsenic and antimony are toxic metalloids, naturally present in the environment and all organisms have developed pathways for their detoxification. The most effective metalloid tolerance systems in eukaryotes include downregulation of metalloid uptake, efflux out of the cell, and complexation with phytochelatin or glutathione followed by sequestration into the vacuole. Understanding of arsenic and antimony transport system is of high importance due to the increasing usage of arsenic-based drugs in the treatment of certain types of cancer and diseases caused by protozoan parasites as well as for the development of bio- and phytoremediation strategies for metalloid polluted areas. However, in contrast to prokaryotes, the knowledge about specific transporters of arsenic and antimony and the mechanisms of metalloid transport in eukaryotes has been very limited for a long time. Here, we review the recent advances in understanding of arsenic and antimony transport pathways in eukaryotes, including a dual role of aquaglyceroporins in uptake and efflux of metalloids, elucidation of arsenic transport mechanism by the yeast Acr3 transporter and its role in arsenic hyperaccumulation in ferns, identification of vacuolar transporters of arsenic-phytochelatin complexes in plants and forms of arsenic substrates recognized by mammalian ABC transporters. PMID:22489166

  9. Diversity patterns of microbial eukaryotes mirror those of bacteria in Antarctic cryoconite holes.

    Science.gov (United States)

    Sommers, Pacifica; Darcy, John L; Gendron, Eli M S; Stanish, Lee F; Bagshaw, Elizabeth A; Porazinska, Dorota L; Schmidt, Steven K

    2018-01-01

    Ice-lidded cryoconite holes on glaciers in the Taylor Valley, Antarctica, provide a unique system of natural mesocosms for studying community structure and assembly. We used high-throughput DNA sequencing to characterize both microbial eukaryotic communities and bacterial communities within cryoconite holes across three glaciers to study similarities in their spatial patterns. We expected that the alpha (phylogenetic diversity) and beta (pairwise community dissimilarity) diversity patterns of eukaryotes in cryoconite holes would be related to those of bacteria, and that they would be related to the biogeochemical gradient within the Taylor Valley. We found that eukaryotic alpha and beta diversity were strongly related to those of bacteria across scales ranging from 140 m to 41 km apart. Alpha diversity of both was significantly related to position in the valley and surface area of the cryoconite hole, with pH also significantly correlated with the eukaryotic diversity. Beta diversity for both bacteria and eukaryotes was significantly related to position in the valley, with bacterial beta diversity also related to nitrate. These results are consistent with transport of sediments onto glaciers occurring primarily at local scales relative to the size of the valley, thus creating feedbacks in local chemistry and diversity. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  10. Genetic exchange in eukaryotes through horizontal transfer: connected by the mobilome.

    Science.gov (United States)

    Wallau, Gabriel Luz; Vieira, Cristina; Loreto, Élgion Lúcio Silva

    2018-01-01

    All living species contain genetic information that was once shared by their common ancestor. DNA is being inherited through generations by vertical transmission (VT) from parents to offspring and from ancestor to descendant species. This process was considered the sole pathway by which biological entities exchange inheritable information. However, Horizontal Transfer (HT), the exchange of genetic information by other means than parents to offspring, was discovered in prokaryotes along with strong evidence showing that it is a very important process by which prokaryotes acquire new genes. For some time now, it has been a scientific consensus that HT events were rare and non-relevant for evolution of eukaryotic species, but there is growing evidence supporting that HT is an important and frequent phenomenon in eukaryotes as well. Here, we will discuss the latest findings regarding HT among eukaryotes, mainly HT of transposons (HTT), establishing HTT once and for all as an important phenomenon that should be taken into consideration to fully understand eukaryotes genome evolution. In addition, we will discuss the latest development methods to detect such events in a broader scale and highlight the new approaches which should be pursued by researchers to fill the knowledge gaps regarding HTT among eukaryotes.

  11. Patterns of prokaryotic lateral gene transfers affecting parasitic microbial eukaryotes

    DEFF Research Database (Denmark)

    Alsmark, Cecilia; Foster, Peter G; Sicheritz-Pontén, Thomas

    2013-01-01

    BACKGROUND: The influence of lateral gene transfer on gene origins and biology in eukaryotes is poorly understood compared with those of prokaryotes. A number of independent investigations focusing on specific genes, individual genomes, or specific functional categories from various eukaryotes have...... approach to systematically investigate lateral gene transfer affecting the proteomes of thirteen, mainly parasitic, microbial eukaryotes, representing four of the six eukaryotic super-groups. All of the genomes investigated have been significantly affected by prokaryote-to-eukaryote lateral gene transfers...... indicated that lateral gene transfer does indeed affect eukaryotic genomes. However, the lack of common methodology and criteria in these studies makes it difficult to assess the general importance and influence of lateral gene transfer on eukaryotic genome evolution. RESULTS: We used a phylogenomic...

  12. <strong>Mini-project>

    DEFF Research Database (Denmark)

    Katajainen, Jyrki

    2008-01-01

    In this project the goal is to develop the safe * family of containers for the CPH STL. The containers to be developed should be safer and more reliable than any of the existing implementations. A special focus should be put on strong exception safety since none of the existing prototypes available...

  13. Strong interactions

    International Nuclear Information System (INIS)

    Froissart, Marcel

    1976-01-01

    Strong interactions are introduced by their more obvious aspect: nuclear forces. In hadron family, the nucleon octet, OMEGA - decuplet, and quark triply are successively considered. Pion wave having been put at the origin of nuclear forces, low energy phenomena are described, the force being explained as an exchange of structure corresponding to a Regge trajectory in a variable rotating state instead of the exchange of a well defined particle. At high energies the concepts of pomeron, parton and stratons are introduced, pionization and fragmentation are briefly differentiated [fr

  14. Prokaryotes versus Eukaryotes: Who is hosting whom?

    Directory of Open Access Journals (Sweden)

    Guillermo eTellez

    2014-10-01

    Full Text Available Microorganisms represent the largest component of biodiversity in our world. For millions of years, prokaryotic microorganisms have functioned as a major selective force shaping eukaryotic evolution. Microbes that live inside and on animals outnumber the animals’ actual somatic and germ cells by an estimated 10-fold. Collectively, the intestinal microbiome represents a ‘forgotten organ’, functioning as an organ inside another that can execute many physiological responsibilities. The nature of primitive eukaryotes was drastically changed due to the association with symbiotic prokaryotes facilitating mutual coevolution of host and microbe. Phytophagous insects have long been used to test theories of evolutionary diversification; moreover, the diversification of a number of phytophagous insect lineages has been linked to mutualisms with microbes. From termites and honey bees to ruminants and mammals, depending on novel biochemistries provided by the prokaryotic microbiome, the association helps to metabolize several nutrients that the host cannot digest and converting these into useful end products (such as short chain fatty acids, a process which has huge impact on the biology and homeostasis of metazoans. More importantly, in a direct and/or indirect way, the intestinal microbiota influences the assembly of gut-associated lymphoid tissue, helps to educate immune system, affects the integrity of the intestinal mucosal barrier, modulates proliferation and differentiation of its epithelial lineages, regulates angiogenesis, and modifies the activity of enteric as well as the central nervous system,. Despite these important effects, the mechanisms by which the gut microbial community influences the host’s biology remains almost entirely unknown. Our aim here is to encourage empirical inquiry into the relationship between mutualism and evolutionary diversification between prokaryotes and eukaryotes which encourage us to postulate: Who is

  15. Mitochondrial uncoupling proteins in unicellular eukaryotes.

    Science.gov (United States)

    Jarmuszkiewicz, Wieslawa; Woyda-Ploszczyca, Andrzej; Antos-Krzeminska, Nina; Sluse, Francis E

    2010-01-01

    Uncoupling proteins (UCPs) are members of the mitochondrial anion carrier protein family that are present in the mitochondrial inner membrane and mediate free fatty acid (FFA)-activated, purine nucleotide (PN)-inhibited proton conductance. Since 1999, the presence of UCPs has been demonstrated in some non-photosynthesising unicellular eukaryotes, including amoeboid and parasite protists, as well as in non-fermentative yeast and filamentous fungi. In the mitochondria of these organisms, UCP activity is revealed upon FFA-induced, PN-inhibited stimulation of resting respiration and a decrease in membrane potential, which are accompanied by a decrease in membranous ubiquinone (Q) reduction level. UCPs in unicellular eukaryotes are able to divert energy from oxidative phosphorylation and thus compete for a proton electrochemical gradient with ATP synthase. Our recent work indicates that membranous Q is a metabolic sensor that might utilise its redox state to release the PN inhibition of UCP-mediated mitochondrial uncoupling under conditions of phosphorylation and resting respiration. The action of reduced Q (QH2) could allow higher or complete activation of UCP. As this regulatory feature was demonstrated for microorganism UCPs (A. castellanii UCP), plant and mammalian UCP1 analogues, and UCP1 in brown adipose tissue, the process could involve all UCPs. Here, we discuss the functional connection and physiological role of UCP and alternative oxidase, two main energy-dissipating systems in the plant-type mitochondrial respiratory chain of unicellular eukaryotes, including the control of cellular energy balance as well as preventive action against the production of reactive oxygen species. Copyright © 2009 Elsevier B.V. All rights reserved.

  16. Consistent mutational paths predict eukaryotic thermostability

    Directory of Open Access Journals (Sweden)

    van Noort Vera

    2013-01-01

    Full Text Available Abstract Background Proteomes of thermophilic prokaryotes have been instrumental in structural biology and successfully exploited in biotechnology, however many proteins required for eukaryotic cell function are absent from bacteria or archaea. With Chaetomium thermophilum, Thielavia terrestris and Thielavia heterothallica three genome sequences of thermophilic eukaryotes have been published. Results Studying the genomes and proteomes of these thermophilic fungi, we found common strategies of thermal adaptation across the different kingdoms of Life, including amino acid biases and a reduced genome size. A phylogenetics-guided comparison of thermophilic proteomes with those of other, mesophilic Sordariomycetes revealed consistent amino acid substitutions associated to thermophily that were also present in an independent lineage of thermophilic fungi. The most consistent pattern is the substitution of lysine by arginine, which we could find in almost all lineages but has not been extensively used in protein stability engineering. By exploiting mutational paths towards the thermophiles, we could predict particular amino acid residues in individual proteins that contribute to thermostability and validated some of them experimentally. By determining the three-dimensional structure of an exemplar protein from C. thermophilum (Arx1, we could also characterise the molecular consequences of some of these mutations. Conclusions The comparative analysis of these three genomes not only enhances our understanding of the evolution of thermophily, but also provides new ways to engineer protein stability.

  17. RNA Export through the NPC in Eukaryotes.

    Science.gov (United States)

    Okamura, Masumi; Inose, Haruko; Masuda, Seiji

    2015-03-20

    In eukaryotic cells, RNAs are transcribed in the nucleus and exported to the cytoplasm through the nuclear pore complex. The RNA molecules that are exported from the nucleus into the cytoplasm include messenger RNAs (mRNAs), ribosomal RNAs (rRNAs), transfer RNAs (tRNAs), small nuclear RNAs (snRNAs), micro RNAs (miRNAs), and viral mRNAs. Each RNA is transported by a specific nuclear export receptor. It is believed that most of the mRNAs are exported by Nxf1 (Mex67 in yeast), whereas rRNAs, snRNAs, and a certain subset of mRNAs are exported in a Crm1/Xpo1-dependent manner. tRNAs and miRNAs are exported by Xpot and Xpo5. However, multiple export receptors are involved in the export of some RNAs, such as 60S ribosomal subunit. In addition to these export receptors, some adapter proteins are required to export RNAs. The RNA export system of eukaryotic cells is also used by several types of RNA virus that depend on the machineries of the host cell in the nucleus for replication of their genome, therefore this review describes the RNA export system of two representative viruses. We also discuss the NPC anchoring-dependent mRNA export factors that directly recruit specific genes to the NPC.

  18. The Genome of Naegleria gruberi Illuminates Early Eukaryotic Versatility

    Energy Technology Data Exchange (ETDEWEB)

    Fritz-Laylin, Lillian K.; Prochnik, Simon E.; Ginger, Michael L.; Dacks, Joel; Carpenter, Meredith L.; Field, Mark C.; Kuo, Alan; Paredez, Alex; Chapman, Jarrod; Pham, Jonathan; Shu, Shengqiang; Neupane, Rochak; Cipriano, Michael; Mancuso, Joel; Tu, Hank; Salamov, Asaf; Lindquist, Erika; Shapiro, Harris; Lucas, Susan; Grigoriev, Igor V.; Cande, W. Zacheus; Fulton, Chandler; Rokhsar, Daniel S.; Dawson, Scott C.

    2010-03-01

    Genome sequences of diverse free-living protists are essential for understanding eukaryotic evolution and molecular and cell biology. The free-living amoeboflagellate Naegleria gruberi belongs to a varied and ubiquitous protist clade (Heterolobosea) that diverged from other eukaryotic lineages over a billion years ago. Analysis of the 15,727 protein-coding genes encoded by Naegleria's 41 Mb nuclear genome indicates a capacity for both aerobic respiration and anaerobic metabolism with concomitant hydrogen production, with fundamental implications for the evolution of organelle metabolism. The Naegleria genome facilitates substantially broader phylogenomic comparisons of free-living eukaryotes than previously possible, allowing us to identify thousands of genes likely present in the pan-eukaryotic ancestor, with 40% likely eukaryotic inventions. Moreover, we construct a comprehensive catalog of amoeboid-motility genes. The Naegleria genome, analyzed in the context of other protists, reveals a remarkably complex ancestral eukaryote with a rich repertoire of cytoskeletal, sexual, signaling, and metabolic modules.

  19. Ultrastructural diversity between centrioles of eukaryotes.

    Science.gov (United States)

    Gupta, Akshari; Kitagawa, Daiju

    2018-02-16

    Several decades of centriole research have revealed the beautiful symmetry present in these microtubule-based organelles, which are required to form centrosomes, cilia, and flagella in many eukaryotes. Centriole architecture is largely conserved across most organisms, however, individual centriolar features such as the central cartwheel or microtubule walls exhibit considerable variability when examined with finer resolution. Here, we review the ultrastructural characteristics of centrioles in commonly studied organisms, highlighting the subtle and not-so-subtle differences between specific structural components of these centrioles. Additionally, we survey some non-canonical centriole structures that have been discovered in various species, from the coaxial bicentrioles of protists and lower land plants to the giant irregular centrioles of the fungus gnat Sciara. Finally, we speculate on the functional significance of these differences between centrioles, and the contribution of individual structural elements such as the cartwheel or microtubules towards the stability of centrioles.Centriole structure, cartwheel, triplet microtubules, SAS-6, centrosome.

  20. Protein splicing and its evolution in eukaryotes

    Directory of Open Access Journals (Sweden)

    Starokadomskyy P. L.

    2010-02-01

    Full Text Available Inteins, or protein introns, are parts of protein sequences that are post-translationally excised, their flanking regions (exteins being spliced together. This process was called protein splicing. Originally inteins were found in prokaryotic or unicellular eukaryotic organisms. But the general principles of post-translation protein rearrangement are evolving yielding different post-translation modification of proteins in multicellular organisms. For clarity, these non-intein mediated events call either protein rearrangements or protein editing. The most intriguing example of protein editing is proteasome-mediated splicing of antigens in vertebrates that may play important role in antigen presentation. Other examples of protein rearrangements are maturation of Hg-proteins (critical receptors in embryogenesis as well as maturation of several metabolic enzymes. Despite a lack of experimental data we try to analyze some intriguing examples of protein splicing evolution.

  1. Redox characteristics of the eukaryotic cytosol

    DEFF Research Database (Denmark)

    López-Mirabal, H Reynaldo; Winther, Jakob R

    2007-01-01

    The eukaryotic cytoplasm has long been regarded as a cellular compartment in which the reduced state of protein cysteines is largely favored. Under normal conditions, the cytosolic low-molecular weight redox buffer, comprising primarily of glutathione, is highly reducing and reactive oxygen species...... (ROS) and glutathionylated proteins are maintained at very low levels. In the present review, recent progress in the understanding of the cytosolic thiol-disulfide redox metabolism and novel analytical approaches to studying cytosolic redox properties are discussed. We will focus on the yeast model...... organism, Saccharomyces cerevisiae, where the combination of genetic and biochemical approaches has brought us furthest in understanding the mechanisms underlying cellular redox regulation. It has been shown in yeast that, in addition to the enzyme glutathione reductase, other mechanisms may exist...

  2. Bacterial proteins pinpoint a single eukaryotic root

    Czech Academy of Sciences Publication Activity Database

    Derelle, R.; Torruella, G.; Klimeš, V.; Brinkmann, H.; Kim, E.; Vlček, Čestmír; Lang, B.F.; Eliáš, M.

    2015-01-01

    Roč. 112, č. 7 (2015), E693-E699 ISSN 0027-8424 R&D Projects: GA ČR GA13-24983S Grant - others:GA MŠk(CZ) ED2.1.00/03.0100; Howard Hughes Medical Institute International Early Career Scientist Program(US) 55007424; Spanish Ministry of Economy and Competitiveness, European Molecular Biology Organization Young Investigator Program(ES) BFU2012-31329; Spanish Ministry of Economy and Competitiveness, "Centro de Excelencia Severo Ochoa" - European Regional Development Fund(ES) Sev-2012-0208, BES-2013-064004 Institutional support: RVO:68378050 Keywords : eukaryote phylogeny * phylogenomics * Opimoda * Diphoda * LECA Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 9.423, year: 2015

  3. Counterintuitive effect of fall mixed layer deepening on eukaryotic new production in the Sargasso Sea

    Science.gov (United States)

    Fawcett, S. E.; Lomas, M. W.; Ward, B. B.; Sigman, D. M.

    2012-12-01

    The Sargasso Sea is characterized by a short period of deep vertical mixing in the late winter and early spring, followed by strong thermal stratification during the summer. Stratification persists into the fall, impeding the upward flux of nitrate from depth so that recycled forms of nitrogen (N) such as ammonium are thought to support most primary production. We collected particles from surface waters during March, July, October, and December, used flow cytometry to separate the prokaryotic and eukaryotic phytoplankton, and analyzed their respective 15N/14N. In all months, the 15N/14N of the prokaryotic genera, Prochlorococcus and Synechococcus, was low, indicative of reliance on recycled N throughout the year. In July, the 15N/14N of eukaryotic phytoplankton was variable but consistently higher than that of the prokaryotes, reflecting eukaryotic consumption of subsurface nitrate. Two eukaryotic profiles from October and December were similar to those from July. In three other fall profiles, the eukaryotes had a 15N/14N similar to that of the prokaryotes, suggesting a switch toward greater reliance on recycled N. This change in the dominant N source supporting eukaryotic production appears to be driven by the density structure of the upper water column. The very shallow low-density surface "mixed layer" (≤20 m) that develops in early-to-mid summer does not contribute to stratification at the base of the euphotic zone, and subsurface nitrate can mix up into the lower euphotic zone, facilitating continued production. The deepening of the mixed layer into the fall, typically taken as an indication of weaker overall stratification, actually strengthens the isolation of the euphotic zone as a whole, reducing the upward supply of nitrate to the photosynthetically active layer. The same counterintuitive dynamic explains the latitudinal patterns in a set of three October depth profiles. Two northern stations (32°N and 27°N) were characterized by a thick, low

  4. The COG database: an updated version includes eukaryotes

    Directory of Open Access Journals (Sweden)

    Sverdlov Alexander V

    2003-09-01

    Full Text Available Abstract Background The availability of multiple, essentially complete genome sequences of prokaryotes and eukaryotes spurred both the demand and the opportunity for the construction of an evolutionary classification of genes from these genomes. Such a classification system based on orthologous relationships between genes appears to be a natural framework for comparative genomics and should facilitate both functional annotation of genomes and large-scale evolutionary studies. Results We describe here a major update of the previously developed system for delineation of Clusters of Orthologous Groups of proteins (COGs from the sequenced genomes of prokaryotes and unicellular eukaryotes and the construction of clusters of predicted orthologs for 7 eukaryotic genomes, which we named KOGs after eukaryotic orthologous groups. The COG collection currently consists of 138,458 proteins, which form 4873 COGs and comprise 75% of the 185,505 (predicted proteins encoded in 66 genomes of unicellular organisms. The eukaryotic orthologous groups (KOGs include proteins from 7 eukaryotic genomes: three animals (the nematode Caenorhabditis elegans, the fruit fly Drosophila melanogaster and Homo sapiens, one plant, Arabidopsis thaliana, two fungi (Saccharomyces cerevisiae and Schizosaccharomyces pombe, and the intracellular microsporidian parasite Encephalitozoon cuniculi. The current KOG set consists of 4852 clusters of orthologs, which include 59,838 proteins, or ~54% of the analyzed eukaryotic 110,655 gene products. Compared to the coverage of the prokaryotic genomes with COGs, a considerably smaller fraction of eukaryotic genes could be included into the KOGs; addition of new eukaryotic genomes is expected to result in substantial increase in the coverage of eukaryotic genomes with KOGs. Examination of the phyletic patterns of KOGs reveals a conserved core represented in all analyzed species and consisting of ~20% of the KOG set. This conserved portion of the

  5. Three distinct modes of intron dynamics in the evolution of eukaryotes.

    Science.gov (United States)

    Carmel, Liran; Wolf, Yuri I; Rogozin, Igor B; Koonin, Eugene V

    2007-07-01

    Several contrasting scenarios have been proposed for the origin and evolution of spliceosomal introns, a hallmark of eukaryotic genes. A comprehensive probabilistic model to obtain a definitive reconstruction of intron evolution was developed and applied to 391 sets of conserved genes from 19 eukaryotic species. It is inferred that a relatively high intron density was reached early, i.e., the last common ancestor of eukaryotes contained >2.15 introns/kilobase, and the last common ancestor of multicellular life forms harbored approximately 3.4 introns/kilobase, a greater intron density than in most of the extant fungi and in some animals. The rates of intron gain and intron loss appear to have been dropping during the last approximately 1.3 billion years, with the decline in the gain rate being much steeper. Eukaryotic lineages exhibit three distinct modes of evolution of the intron-exon structure. The primary, balanced mode, apparently, operates in all lineages. In this mode, intron gain and loss are strongly and positively correlated, in contrast to previous reports on inverse correlation between these processes. The second mode involves an elevated rate of intron loss and is prevalent in several lineages, such as fungi and insects. The third mode, characterized by elevated rate of intron gain, is seen only in deep branches of the tree, indicating that bursts of intron invasion occurred at key points in eukaryotic evolution, such as the origin of animals. Intron dynamics could depend on multiple mechanisms, and in the balanced mode, gain and loss of introns might share common mechanistic features.

  6. Genomic impact of eukaryotic transposable elements.

    Science.gov (United States)

    Arkhipova, Irina R; Batzer, Mark A; Brosius, Juergen; Feschotte, Cédric; Moran, John V; Schmitz, Jürgen; Jurka, Jerzy

    2012-11-21

    The third international conference on the genomic impact of eukaryotic transposable elements (TEs) was held 24 to 28 February 2012 at the Asilomar Conference Center, Pacific Grove, CA, USA. Sponsored in part by the National Institutes of Health grant 5 P41 LM006252, the goal of the conference was to bring together researchers from around the world who study the impact and mechanisms of TEs using multiple computational and experimental approaches. The meeting drew close to 170 attendees and included invited floor presentations on the biology of TEs and their genomic impact, as well as numerous talks contributed by young scientists. The workshop talks were devoted to computational analysis of TEs with additional time for discussion of unresolved issues. Also, there was ample opportunity for poster presentations and informal evening discussions. The success of the meeting reflects the important role of Repbase in comparative genomic studies, and emphasizes the need for close interactions between experimental and computational biologists in the years to come.

  7. Genomic impact of eukaryotic transposable elements

    Directory of Open Access Journals (Sweden)

    Arkhipova Irina R

    2012-11-01

    Full Text Available Abstract The third international conference on the genomic impact of eukaryotic transposable elements (TEs was held 24 to 28 February 2012 at the Asilomar Conference Center, Pacific Grove, CA, USA. Sponsored in part by the National Institutes of Health grant 5 P41 LM006252, the goal of the conference was to bring together researchers from around the world who study the impact and mechanisms of TEs using multiple computational and experimental approaches. The meeting drew close to 170 attendees and included invited floor presentations on the biology of TEs and their genomic impact, as well as numerous talks contributed by young scientists. The workshop talks were devoted to computational analysis of TEs with additional time for discussion of unresolved issues. Also, there was ample opportunity for poster presentations and informal evening discussions. The success of the meeting reflects the important role of Repbase in comparative genomic studies, and emphasizes the need for close interactions between experimental and computational biologists in the years to come.

  8. Do lipids shape the eukaryotic cell cycle?

    Science.gov (United States)

    Furse, Samuel; Shearman, Gemma C

    2018-01-01

    Successful passage through the cell cycle presents a number of structural challenges to the cell. Inceptive studies carried out in the last five years have produced clear evidence of modulations in the lipid profile (sometimes referred to as the lipidome) of eukaryotes as a function of the cell cycle. This mounting body of evidence indicates that lipids play key roles in the structural transformations seen across the cycle. The accumulation of this evidence coincides with a revolution in our understanding of how lipid composition regulates a plethora of biological processes ranging from protein activity through to cellular signalling and membrane compartmentalisation. In this review, we discuss evidence from biological, chemical and physical studies of the lipid fraction across the cell cycle that demonstrate that lipids are well-developed cellular components at the heart of the biological machinery responsible for managing progress through the cell cycle. Furthermore, we discuss the mechanisms by which this careful control is exercised. Crown Copyright © 2017. Published by Elsevier B.V. All rights reserved.

  9. Origins and evolution of viruses of eukaryotes: The ultimate modularity

    International Nuclear Information System (INIS)

    Koonin, Eugene V.; Dolja, Valerian V.; Krupovic, Mart

    2015-01-01

    Viruses and other selfish genetic elements are dominant entities in the biosphere, with respect to both physical abundance and genetic diversity. Various selfish elements parasitize on all cellular life forms. The relative abundances of different classes of viruses are dramatically different between prokaryotes and eukaryotes. In prokaryotes, the great majority of viruses possess double-stranded (ds) DNA genomes, with a substantial minority of single-stranded (ss) DNA viruses and only limited presence of RNA viruses. In contrast, in eukaryotes, RNA viruses account for the majority of the virome diversity although ssDNA and dsDNA viruses are common as well. Phylogenomic analysis yields tangible clues for the origins of major classes of eukaryotic viruses and in particular their likely roots in prokaryotes. Specifically, the ancestral genome of positive-strand RNA viruses of eukaryotes might have been assembled de novo from genes derived from prokaryotic retroelements and bacteria although a primordial origin of this class of viruses cannot be ruled out. Different groups of double-stranded RNA viruses derive either from dsRNA bacteriophages or from positive-strand RNA viruses. The eukaryotic ssDNA viruses apparently evolved via a fusion of genes from prokaryotic rolling circle-replicating plasmids and positive-strand RNA viruses. Different families of eukaryotic dsDNA viruses appear to have originated from specific groups of bacteriophages on at least two independent occasions. Polintons, the largest known eukaryotic transposons, predicted to also form virus particles, most likely, were the evolutionary intermediates between bacterial tectiviruses and several groups of eukaryotic dsDNA viruses including the proposed order “Megavirales” that unites diverse families of large and giant viruses. Strikingly, evolution of all classes of eukaryotic viruses appears to have involved fusion between structural and replicative gene modules derived from different sources

  10. Origins and evolution of viruses of eukaryotes: The ultimate modularity

    Energy Technology Data Exchange (ETDEWEB)

    Koonin, Eugene V., E-mail: koonin@ncbi.nlm.nih.gov [National Center for Biotechnology Information, National Library of Medicine, National Institutes of Health, Bethesda, MD 20894 (United States); Dolja, Valerian V., E-mail: doljav@science.oregonstate.edu [Department of Botany and Plant Pathology, Oregon State University, Corvallis, OR 97331 (United States); Krupovic, Mart, E-mail: krupovic@pasteur.fr [Institut Pasteur, Unité Biologie Moléculaire du Gène chez les Extrêmophiles, Department of Microbiology, Paris 75015 (France)

    2015-05-15

    Viruses and other selfish genetic elements are dominant entities in the biosphere, with respect to both physical abundance and genetic diversity. Various selfish elements parasitize on all cellular life forms. The relative abundances of different classes of viruses are dramatically different between prokaryotes and eukaryotes. In prokaryotes, the great majority of viruses possess double-stranded (ds) DNA genomes, with a substantial minority of single-stranded (ss) DNA viruses and only limited presence of RNA viruses. In contrast, in eukaryotes, RNA viruses account for the majority of the virome diversity although ssDNA and dsDNA viruses are common as well. Phylogenomic analysis yields tangible clues for the origins of major classes of eukaryotic viruses and in particular their likely roots in prokaryotes. Specifically, the ancestral genome of positive-strand RNA viruses of eukaryotes might have been assembled de novo from genes derived from prokaryotic retroelements and bacteria although a primordial origin of this class of viruses cannot be ruled out. Different groups of double-stranded RNA viruses derive either from dsRNA bacteriophages or from positive-strand RNA viruses. The eukaryotic ssDNA viruses apparently evolved via a fusion of genes from prokaryotic rolling circle-replicating plasmids and positive-strand RNA viruses. Different families of eukaryotic dsDNA viruses appear to have originated from specific groups of bacteriophages on at least two independent occasions. Polintons, the largest known eukaryotic transposons, predicted to also form virus particles, most likely, were the evolutionary intermediates between bacterial tectiviruses and several groups of eukaryotic dsDNA viruses including the proposed order “Megavirales” that unites diverse families of large and giant viruses. Strikingly, evolution of all classes of eukaryotic viruses appears to have involved fusion between structural and replicative gene modules derived from different sources

  11. Specificity and evolvability in eukaryotic protein interaction networks.

    Directory of Open Access Journals (Sweden)

    Pedro Beltrao

    2007-02-01

    Full Text Available Progress in uncovering the protein interaction networks of several species has led to questions of what underlying principles might govern their organization. Few studies have tried to determine the impact of protein interaction network evolution on the observed physiological differences between species. Using comparative genomics and structural information, we show here that eukaryotic species have rewired their interactomes at a fast rate of approximately 10(-5 interactions changed per protein pair, per million years of divergence. For Homo sapiens this corresponds to 10(3 interactions changed per million years. Additionally we find that the specificity of binding strongly determines the interaction turnover and that different biological processes show significantly different link dynamics. In particular, human proteins involved in immune response, transport, and establishment of localization show signs of positive selection for change of interactions. Our analysis suggests that a small degree of molecular divergence can give rise to important changes at the network level. We propose that the power law distribution observed in protein interaction networks could be partly explained by the cell's requirement for different degrees of protein binding specificity.

  12. Discrepancy variation of dinucleotide microsatellite repeats in eukaryotic genomes

    Directory of Open Access Journals (Sweden)

    HUAN GAO

    2009-01-01

    Full Text Available To address whether there are differences of variation among repeat motif types and among taxonomic groups, we present here an analysis of variation and correlation of dinucleotide microsatellite repeats in eukaryotic genomes. Ten taxonomic groups were compared, those being primates, mammalia (excluding primates and rodentia, rodentia, birds, fish, amphibians and reptiles, insects, molluscs, plants and fungi, respectively. The data used in the analysis is from the literature published in the Journal of Molecular Ecology Notes. Analysis of variation reveals that there are no significant differences between AC and AG repeat motif types. Moreover, the number of alleles correlates positively with the copy number in both AG and AC repeats. Similar conclusions can be obtained from each taxonomic group. These results strongly suggest that the increase of SSR variation is almost linear with the increase of the copy number of each repeat motif. As well, the results suggest that the variability of SSR in the genomes of low-ranking species seem to be more than that of high-ranking species, excluding primates and fungi.

  13. Genome-reconstruction for eukaryotes from complex natural microbial communities.

    Science.gov (United States)

    West, Patrick T; Probst, Alexander J; Grigoriev, Igor V; Thomas, Brian C; Banfield, Jillian F

    2018-04-01

    Microbial eukaryotes are integral components of natural microbial communities, and their inclusion is critical for many ecosystem studies, yet the majority of published metagenome analyses ignore eukaryotes. In order to include eukaryotes in environmental studies, we propose a method to recover eukaryotic genomes from complex metagenomic samples. A key step for genome recovery is separation of eukaryotic and prokaryotic fragments. We developed a k -mer-based strategy, EukRep, for eukaryotic sequence identification and applied it to environmental samples to show that it enables genome recovery, genome completeness evaluation, and prediction of metabolic potential. We used this approach to test the effect of addition of organic carbon on a geyser-associated microbial community and detected a substantial change of the community metabolism, with selection against almost all candidate phyla bacteria and archaea and for eukaryotes. Near complete genomes were reconstructed for three fungi placed within the Eurotiomycetes and an arthropod. While carbon fixation and sulfur oxidation were important functions in the geyser community prior to carbon addition, the organic carbon-impacted community showed enrichment for secreted proteases, secreted lipases, cellulose targeting CAZymes, and methanol oxidation. We demonstrate the broader utility of EukRep by reconstructing and evaluating relatively high-quality fungal, protist, and rotifer genomes from complex environmental samples. This approach opens the way for cultivation-independent analyses of whole microbial communities. © 2018 West et al.; Published by Cold Spring Harbor Laboratory Press.

  14. Potential of industrial biotechnology with cyanobacteria and eukaryotic microalgae.

    Science.gov (United States)

    Wijffels, René H; Kruse, Olaf; Hellingwerf, Klaas J

    2013-06-01

    Both cyanobacteria and eukaryotic microalgae are promising organisms for sustainable production of bulk products such as food, feed, materials, chemicals and fuels. In this review we will summarize the potential and current biotechnological developments. Cyanobacteria are promising host organisms for the production of small molecules that can be secreted such as ethanol, butanol, fatty acids and other organic acids. Eukaryotic microalgae are interesting for products for which cellular storage is important such as proteins, lipids, starch and alkanes. For the development of new and promising lines of production, strains of both cyanobacteria and eukaryotic microalgae have to be improved. Transformation systems have been much better developed in cyanobacteria. However, several products would be preferably produced with eukaryotic microalgae. In the case of cyanobacteria a synthetic-systems biology approach has a great potential to exploit cyanobacteria as cell factories. For eukaryotic microalgae transformation systems need to be further developed. A promising strategy is transformation of heterologous (prokaryotic and eukaryotic) genes in established eukaryotic hosts such as Chlamydomonas reinhardtii. Experimental outdoor pilots under containment for the production of genetically modified cyanobacteria and microalgae are in progress. For full scale production risks of release of genetically modified organisms need to be assessed. Copyright © 2013. Published by Elsevier Ltd.

  15. Sequence analysis of RNase MRP RNA reveals its origination from eukaryotic RNase P RNA

    Science.gov (United States)

    Zhu, Yanglong; Stribinskis, Vilius; Ramos, Kenneth S.; Li, Yong

    2006-01-01

    RNase MRP is a eukaryote-specific endoribonuclease that generates RNA primers for mitochondrial DNA replication and processes precursor rRNA. RNase P is a ubiquitous endoribonuclease that cleaves precursor tRNA transcripts to produce their mature 5′ termini. We found extensive sequence homology of catalytic domains and specificity domains between their RNA subunits in many organisms. In Candida glabrata, the internal loop of helix P3 is 100% conserved between MRP and P RNAs. The helix P8 of MRP RNA from microsporidia Encephalitozoon cuniculi is identical to that of P RNA. Sequence homology can be widely spread over the whole molecule of MRP RNA and P RNA, such as those from Dictyostelium discoideum. These conserved nucleotides between the MRP and P RNAs strongly support the hypothesis that the MRP RNA is derived from the P RNA molecule in early eukaryote evolution. PMID:16540690

  16. Origins of robustness in translational control via eukaryotic translation initiation factor (eIF) 2.

    Science.gov (United States)

    Khan, Mohammad Farhan; Spurgeon, Sarah; von der Haar, Tobias

    2018-05-14

    Phosphorylation of eukaryotic translation initiation factor 2 (eIF2) is one of the best studied and most widely used means for regulating protein synthesis activity in eukaryotic cells. This pathway regulates protein synthesis in response to stresses, viral infections, and nutrient depletion, among others. We present analyses of an ordinary differential equation-based model of this pathway, which aim to identify its principal robustness-conferring features. Our analyses indicate that robustness is a distributed property, rather than arising from the properties of any one individual pathway species. However, robustness-conferring properties are unevenly distributed between the different species, and we identify a guanine nucleotide dissociation inhibitor (GDI) complex as a species that likely contributes strongly to the robustness of the pathway. Our analyses make further predictions on the dynamic response to different types of kinases that impinge on eIF2. Copyright © 2018 Elsevier Ltd. All rights reserved.

  17. Engineering a palette of eukaryotic chromoproteins for bacterial synthetic biology.

    Science.gov (United States)

    Liljeruhm, Josefine; Funk, Saskia K; Tietscher, Sandra; Edlund, Anders D; Jamal, Sabri; Wistrand-Yuen, Pikkei; Dyrhage, Karl; Gynnå, Arvid; Ivermark, Katarina; Lövgren, Jessica; Törnblom, Viktor; Virtanen, Anders; Lundin, Erik R; Wistrand-Yuen, Erik; Forster, Anthony C

    2018-01-01

    Coral reefs are colored by eukaryotic chromoproteins (CPs) that are homologous to green fluorescent protein. CPs differ from fluorescent proteins (FPs) by intensely absorbing visible light to give strong colors in ambient light. This endows CPs with certain advantages over FPs, such as instrument-free detection uncomplicated by ultra-violet light damage or background fluorescence, efficient Förster resonance energy transfer (FRET) quenching, and photoacoustic imaging. Thus, CPs have found utility as genetic markers and in teaching, and are attractive for potential cell biosensor applications in the field. Most near-term applications of CPs require expression in a different domain of life: bacteria. However, it is unclear which of the eukaryotic CP genes might be suitable and how best to assay them. Here, taking advantage of codon optimization programs in 12 cases, we engineered 14 CP sequences (meffRed, eforRed, asPink, spisPink, scOrange, fwYellow, amilGFP, amajLime, cjBlue, meffBlue, aeBlue, amilCP, tsPurple and gfasPurple) into a palette of Escherichia coli BioBrick plasmids. BioBricks comply with synthetic biology's most widely used, simplified, cloning standard. Differences in color intensities, maturation times and fitness costs of expression were compared under the same conditions, and visible readout of gene expression was quantitated. A surprisingly large variation in cellular fitness costs was found, resulting in loss of color in some overnight liquid cultures of certain high-copy-plasmid-borne CPs, and cautioning the use of multiple CPs as markers in competition assays. We solved these two problems by integrating pairs of these genes into the chromosome and by engineering versions of the same CP with very different colors. Availability of 14 engineered CP genes compared in E. coli , together with chromosomal mutants suitable for competition assays, should simplify and expand CP study and applications. There was no single plasmid-borne CP that combined

  18. Functions and structures of eukaryotic recombination proteins

    International Nuclear Information System (INIS)

    Ogawa, Tomoko

    1994-01-01

    We have found that Rad51 and RecA Proteins form strikingly similar structures together with dsDNA and ATP. Their right handed helical nucleoprotein filaments extend the B-form DNA double helixes to 1.5 times in length and wind the helix. The similarity and uniqueness of their structures must reflect functional homologies between these proteins. Therefore, it is highly probable that similar recombination proteins are present in various organisms of different evolutional states. We have succeeded to clone RAD51 genes from human, mouse, chicken and fission yeast genes, and found that the homologues are widely distributed in eukaryotes. The HsRad51 and MmRad51 or ChRad51 proteins consist of 339 amino acids differing only by 4 or 12 amino acids, respectively, and highly homologous to both yeast proteins, but less so to Dmcl. All of these proteins are homologous to the region from residues 33 to 240 of RecA which was named ''homologous core. The homologous core is likely to be responsible for functions common for all of them, such as the formation of helical nucleoprotein filament that is considered to be involved in homologous pairing in the recombination reaction. The mouse gene is transcribed at a high level in thymus, spleen, testis, and ovary, at lower level in brain and at a further lower level in some other tissues. It is transcribed efficiently in recombination active tissues. A clear functional difference of Rad51 homologues from RecA was suggested by the failure of heterologous genes to complement the deficiency of Scrad51 mutants. This failure seems to reflect the absence of a compatible partner, such as ScRad52 protein in the case of ScRad51 protein, between different species. Thus, these discoveries play a role of the starting point to understand the fundamental gene targeting in mammalian cells and in gene therapy. (J.P.N.)

  19. HIV-1 Replication and the Cellular Eukaryotic Translation Apparatus

    Directory of Open Access Journals (Sweden)

    Santiago Guerrero

    2015-01-01

    Full Text Available Eukaryotic translation is a complex process composed of three main steps: initiation, elongation, and termination. During infections by RNA- and DNA-viruses, the eukaryotic translation machinery is used to assure optimal viral protein synthesis. Human immunodeficiency virus type I (HIV-1 uses several non-canonical pathways to translate its own proteins, such as leaky scanning, frameshifting, shunt, and cap-independent mechanisms. Moreover, HIV-1 modulates the host translation machinery by targeting key translation factors and overcomes different cellular obstacles that affect protein translation. In this review, we describe how HIV-1 proteins target several components of the eukaryotic translation machinery, which consequently improves viral translation and replication.

  20. AUG is the only initiation codon in eukaryotes

    Energy Technology Data Exchange (ETDEWEB)

    Sherman, F; McKnight, G; Stewart, J W

    1980-01-01

    An analysis of mutants of the yeast Saccharomyces cerevisiae indicates that AUG is the sole codon capable of initiating translation of iso-1-cytochrome c. This result with yeast and the sequence results of numerous eukaryotic genes indicate that AUG is the only initiation codon in eukaryotes; in contrast, results with Escherichia colia and bacteriophages indicate that both AUG and GUG are initiation codons in prokaryotes. The difference can be explained by the lack of the t/sup 6/ A hypermodified nucleoside (N-(9-(..beta..-D-ribofuranosyl)purin-6-ylcarbamoyl)threonine) in prokaryotic initiator tRNA and its presence in eukaryotic initiator tRNA.

  1. David and Goliath: chemical perturbation of eukaryotes by bacteria.

    Science.gov (United States)

    Ho, Louis K; Nodwell, Justin R

    2016-03-01

    Environmental microbes produce biologically active small molecules that have been mined extensively as antibiotics and a smaller number of drugs that act on eukaryotic cells. It is known that there are additional bioactives to be discovered from this source. While the discovery of new antibiotics is challenged by the frequent discovery of known compounds, we contend that the eukaryote-active compounds may be less saturated. Indeed, despite there being far fewer eukaryotic-active natural products these molecules interact with a far richer diversity of molecular and cellular targets.

  2. Potential of industrial biotechnology with cyanobacteria and eukaryotic microalgae.

    NARCIS (Netherlands)

    Wijffels, R.H.; Kruse, O.; Hellingwerf, K.J.

    2013-01-01

    Both cyanobacteria and eukaryotic microalgae are promising organisms for sustainable production of bulk products such as food, feed, materials, chemicals and fuels. In this review we will summarize the potential and current biotechnological developments. Cyanobacteria are promising host organisms

  3. Conservation and Variability of Meiosis Across the Eukaryotes.

    Science.gov (United States)

    Loidl, Josef

    2016-11-23

    Comparisons among a variety of eukaryotes have revealed considerable variability in the structures and processes involved in their meiosis. Nevertheless, conventional forms of meiosis occur in all major groups of eukaryotes, including early-branching protists. This finding confirms that meiosis originated in the common ancestor of all eukaryotes and suggests that primordial meiosis may have had many characteristics in common with conventional extant meiosis. However, it is possible that the synaptonemal complex and the delicate crossover control related to its presence were later acquisitions. Later still, modifications to meiotic processes occurred within different groups of eukaryotes. Better knowledge on the spectrum of derived and uncommon forms of meiosis will improve our understanding of many still mysterious aspects of the meiotic process and help to explain the evolutionary basis of functional adaptations to the meiotic program.

  4. Nitrate storage and dissimilatory nitrate reduction by eukaryotic microbes

    DEFF Research Database (Denmark)

    Kamp, Anja; Høgslund, Signe; Risgaard-Petersen, Nils

    2015-01-01

    The microbial nitrogen cycle is one of the most complex and environmentally important element cycles on Earth and has long been thought to be mediated exclusively by prokaryotic microbes. Rather recently, it was discovered that certain eukaryotic microbes are able to store nitrate intracellularly......, suggesting that eukaryotes may rival prokaryotes in terms of dissimilatory nitrate reduction. Finally, this review article sketches some evolutionary perspectives of eukaryotic nitrate metabolism and identifies open questions that need to be addressed in future investigations....... and use it for dissimilatory nitrate reduction in the absence of oxygen. The paradigm shift that this entailed is ecologically significant because the eukaryotes in question comprise global players like diatoms, foraminifers, and fungi. This review article provides an unprecedented overview of nitrate...

  5. Structure and Mechanism of a Eukaryotic FMN Adenylyltransferase

    OpenAIRE

    Huerta, Carlos; Borek, Dominika; Machius, Mischa; Grishin, Nick V.; Zhang, Hong

    2009-01-01

    Flavin mononucleotide adenylyltransferase (FMNAT) catalyzes the formation of the essential flavocoenzyme FAD and plays an important role in flavocoenzyme homeostasis regulation. By sequence comparison, bacterial and eukaryotic FMNAT enzymes belong to two different protein superfamilies and apparently utilize different set of active site residues to accomplish the same chemistry. Here we report the first structural characterization of a eukaryotic FMNAT from a pathogenic yeast Candida glabrata...

  6. Massive expansion of the calpain gene family in unicellular eukaryotes

    Directory of Open Access Journals (Sweden)

    Zhao Sen

    2012-09-01

    Full Text Available Abstract Background Calpains are Ca2+-dependent cysteine proteases that participate in a range of crucial cellular processes. Dysfunction of these enzymes may cause, for instance, life-threatening diseases in humans, the loss of sex determination in nematodes and embryo lethality in plants. Although the calpain family is well characterized in animal and plant model organisms, there is a great lack of knowledge about these genes in unicellular eukaryote species (i.e. protists. Here, we study the distribution and evolution of calpain genes in a wide range of eukaryote genomes from major branches in the tree of life. Results Our investigations reveal 24 types of protein domains that are combined with the calpain-specific catalytic domain CysPc. In total we identify 41 different calpain domain architectures, 28 of these domain combinations have not been previously described. Based on our phylogenetic inferences, we propose that at least four calpain variants were established in the early evolution of eukaryotes, most likely before the radiation of all the major supergroups of eukaryotes. Many domains associated with eukaryotic calpain genes can be found among eubacteria or archaebacteria but never in combination with the CysPc domain. Conclusions The analyses presented here show that ancient modules present in prokaryotes, and a few de novo eukaryote domains, have been assembled into many novel domain combinations along the evolutionary history of eukaryotes. Some of the new calpain genes show a narrow distribution in a few branches in the tree of life, likely representing lineage-specific innovations. Hence, the functionally important classical calpain genes found among humans and vertebrates make up only a tiny fraction of the calpain family. In fact, a massive expansion of the calpain family occurred by domain shuffling among unicellular eukaryotes and contributed to a wealth of functionally different genes.

  7. Massive expansion of the calpain gene family in unicellular eukaryotes.

    Science.gov (United States)

    Zhao, Sen; Liang, Zhe; Demko, Viktor; Wilson, Robert; Johansen, Wenche; Olsen, Odd-Arne; Shalchian-Tabrizi, Kamran

    2012-09-29

    Calpains are Ca2+-dependent cysteine proteases that participate in a range of crucial cellular processes. Dysfunction of these enzymes may cause, for instance, life-threatening diseases in humans, the loss of sex determination in nematodes and embryo lethality in plants. Although the calpain family is well characterized in animal and plant model organisms, there is a great lack of knowledge about these genes in unicellular eukaryote species (i.e. protists). Here, we study the distribution and evolution of calpain genes in a wide range of eukaryote genomes from major branches in the tree of life. Our investigations reveal 24 types of protein domains that are combined with the calpain-specific catalytic domain CysPc. In total we identify 41 different calpain domain architectures, 28 of these domain combinations have not been previously described. Based on our phylogenetic inferences, we propose that at least four calpain variants were established in the early evolution of eukaryotes, most likely before the radiation of all the major supergroups of eukaryotes. Many domains associated with eukaryotic calpain genes can be found among eubacteria or archaebacteria but never in combination with the CysPc domain. The analyses presented here show that ancient modules present in prokaryotes, and a few de novo eukaryote domains, have been assembled into many novel domain combinations along the evolutionary history of eukaryotes. Some of the new calpain genes show a narrow distribution in a few branches in the tree of life, likely representing lineage-specific innovations. Hence, the functionally important classical calpain genes found among humans and vertebrates make up only a tiny fraction of the calpain family. In fact, a massive expansion of the calpain family occurred by domain shuffling among unicellular eukaryotes and contributed to a wealth of functionally different genes.

  8. On the Diversification of the Translation Apparatus across Eukaryotes

    Directory of Open Access Journals (Sweden)

    Greco Hernández

    2012-01-01

    Full Text Available Diversity is one of the most remarkable features of living organisms. Current assessments of eukaryote biodiversity reaches 1.5 million species, but the true figure could be several times that number. Diversity is ingrained in all stages and echelons of life, namely, the occupancy of ecological niches, behavioral patterns, body plans and organismal complexity, as well as metabolic needs and genetics. In this review, we will discuss that diversity also exists in a key biochemical process, translation, across eukaryotes. Translation is a fundamental process for all forms of life, and the basic components and mechanisms of translation in eukaryotes have been largely established upon the study of traditional, so-called model organisms. By using modern genome-wide, high-throughput technologies, recent studies of many nonmodel eukaryotes have unveiled a surprising diversity in the configuration of the translation apparatus across eukaryotes, showing that this apparatus is far from being evolutionarily static. For some of the components of this machinery, functional differences between different species have also been found. The recent research reviewed in this article highlights the molecular and functional diversification the translational machinery has undergone during eukaryotic evolution. A better understanding of all aspects of organismal diversity is key to a more profound knowledge of life.

  9. Single Cell Genomics and Transcriptomics for Unicellular Eukaryotes

    Energy Technology Data Exchange (ETDEWEB)

    Ciobanu, Doina; Clum, Alicia; Singh, Vasanth; Salamov, Asaf; Han, James; Copeland, Alex; Grigoriev, Igor; James, Timothy; Singer, Steven; Woyke, Tanja; Malmstrom, Rex; Cheng, Jan-Fang

    2014-03-14

    Despite their small size, unicellular eukaryotes have complex genomes with a high degree of plasticity that allow them to adapt quickly to environmental changes. Unicellular eukaryotes live with prokaryotes and higher eukaryotes, frequently in symbiotic or parasitic niches. To this day their contribution to the dynamics of the environmental communities remains to be understood. Unfortunately, the vast majority of eukaryotic microorganisms are either uncultured or unculturable, making genome sequencing impossible using traditional approaches. We have developed an approach to isolate unicellular eukaryotes of interest from environmental samples, and to sequence and analyze their genomes and transcriptomes. We have tested our methods with six species: an uncharacterized protist from cellulose-enriched compost identified as Platyophrya, a close relative of P. vorax; the fungus Metschnikowia bicuspidate, a parasite of water flea Daphnia; the mycoparasitic fungi Piptocephalis cylindrospora, a parasite of Cokeromyces and Mucor; Caulochytrium protosteloides, a parasite of Sordaria; Rozella allomycis, a parasite of the water mold Allomyces; and the microalgae Chlamydomonas reinhardtii. Here, we present the four components of our approach: pre-sequencing methods, sequence analysis for single cell genome assembly, sequence analysis of single cell transcriptomes, and genome annotation. This technology has the potential to uncover the complexity of single cell eukaryotes and their role in the environmental samples.

  10. An Evolutionary Framework for Understanding the Origin of Eukaryotes

    Directory of Open Access Journals (Sweden)

    Neil W. Blackstone

    2016-04-01

    Full Text Available Two major obstacles hinder the application of evolutionary theory to the origin of eukaryotes. The first is more apparent than real—the endosymbiosis that led to the mitochondrion is often described as “non-Darwinian” because it deviates from the incremental evolution championed by the modern synthesis. Nevertheless, endosymbiosis can be accommodated by a multi-level generalization of evolutionary theory, which Darwin himself pioneered. The second obstacle is more serious—all of the major features of eukaryotes were likely present in the last eukaryotic common ancestor thus rendering comparative methods ineffective. In addition to a multi-level theory, the development of rigorous, sequence-based phylogenetic and comparative methods represents the greatest achievement of modern evolutionary theory. Nevertheless, the rapid evolution of major features in the eukaryotic stem group requires the consideration of an alternative framework. Such a framework, based on the contingent nature of these evolutionary events, is developed and illustrated with three examples: the putative intron proliferation leading to the nucleus and the cell cycle; conflict and cooperation in the origin of eukaryotic bioenergetics; and the inter-relationship between aerobic metabolism, sterol synthesis, membranes, and sex. The modern synthesis thus provides sufficient scope to develop an evolutionary framework to understand the origin of eukaryotes.

  11. Compositional patterns in the genomes of unicellular eukaryotes.

    Science.gov (United States)

    Costantini, Maria; Alvarez-Valin, Fernando; Costantini, Susan; Cammarano, Rosalia; Bernardi, Giorgio

    2013-11-05

    The genomes of multicellular eukaryotes are compartmentalized in mosaics of isochores, large and fairly homogeneous stretches of DNA that belong to a small number of families characterized by different average GC levels, by different gene concentration (that increase with GC), different chromatin structures, different replication timing in the cell cycle, and other different properties. A question raised by these basic results concerns how far back in evolution the compartmentalized organization of the eukaryotic genomes arose. In the present work we approached this problem by studying the compositional organization of the genomes from the unicellular eukaryotes for which full sequences are available, the sample used being representative. The average GC levels of the genomes from unicellular eukaryotes cover an extremely wide range (19%-60% GC) and the compositional patterns of individual genomes are extremely different but all genomes tested show a compositional compartmentalization. The average GC range of the genomes of unicellular eukaryotes is very broad (as broad as that of prokaryotes) and individual compositional patterns cover a very broad range from very narrow to very complex. Both features are not surprising for organisms that are very far from each other both in terms of phylogenetic distances and of environmental life conditions. Most importantly, all genomes tested, a representative sample of all supergroups of unicellular eukaryotes, are compositionally compartmentalized, a major difference with prokaryotes.

  12. Eukaryotic systematics: a user's guide for cell biologists and parasitologists.

    Science.gov (United States)

    Walker, Giselle; Dorrell, Richard G; Schlacht, Alexander; Dacks, Joel B

    2011-11-01

    Single-celled parasites like Entamoeba, Trypanosoma, Phytophthora and Plasmodium wreak untold havoc on human habitat and health. Understanding the position of the various protistan pathogens in the larger context of eukaryotic diversity informs our study of how these parasites operate on a cellular level, as well as how they have evolved. Here, we review the literature that has brought our understanding of eukaryotic relationships from an idea of parasites as primitive cells to a crystallized view of diversity that encompasses 6 major divisions, or supergroups, of eukaryotes. We provide an updated taxonomic scheme (for 2011), based on extensive genomic, ultrastructural and phylogenetic evidence, with three differing levels of taxonomic detail for ease of referencing and accessibility (see supplementary material at Cambridge Journals On-line). Two of the most pressing issues in cellular evolution, the root of the eukaryotic tree and the evolution of photosynthesis in complex algae, are also discussed along with ideas about what the new generation of genome sequencing technologies may contribute to the field of eukaryotic systematics. We hope that, armed with this user's guide, cell biologists and parasitologists will be encouraged about taking an increasingly evolutionary point of view in the battle against parasites representing real dangers to our livelihoods and lives.

  13. Evolution of DNA replication protein complexes in eukaryotes and Archaea.

    Directory of Open Access Journals (Sweden)

    Nicholas Chia

    Full Text Available BACKGROUND: The replication of DNA in Archaea and eukaryotes requires several ancillary complexes, including proliferating cell nuclear antigen (PCNA, replication factor C (RFC, and the minichromosome maintenance (MCM complex. Bacterial DNA replication utilizes comparable proteins, but these are distantly related phylogenetically to their archaeal and eukaryotic counterparts at best. METHODOLOGY/PRINCIPAL FINDINGS: While the structures of each of the complexes do not differ significantly between the archaeal and eukaryotic versions thereof, the evolutionary dynamic in the two cases does. The number of subunits in each complex is constant across all taxa. However, they vary subtly with regard to composition. In some taxa the subunits are all identical in sequence, while in others some are homologous rather than identical. In the case of eukaryotes, there is no phylogenetic variation in the makeup of each complex-all appear to derive from a common eukaryotic ancestor. This is not the case in Archaea, where the relationship between the subunits within each complex varies taxon-to-taxon. We have performed a detailed phylogenetic analysis of these relationships in order to better understand the gene duplications and divergences that gave rise to the homologous subunits in Archaea. CONCLUSION/SIGNIFICANCE: This domain level difference in evolution suggests that different forces have driven the evolution of DNA replication proteins in each of these two domains. In addition, the phylogenies of all three gene families support the distinctiveness of the proposed archaeal phylum Thaumarchaeota.

  14. Interaction of tRNA with Eukaryotic Ribosome

    Directory of Open Access Journals (Sweden)

    Dmitri Graifer

    2015-03-01

    Full Text Available This paper is a review of currently available data concerning interactions of tRNAs with the eukaryotic ribosome at various stages of translation. These data include the results obtained by means of cryo-electron microscopy and X-ray crystallography applied to various model ribosomal complexes, site-directed cross-linking with the use of tRNA derivatives bearing chemically or photochemically reactive groups in the CCA-terminal fragment and chemical probing of 28S rRNA in the region of the peptidyl transferase center. Similarities and differences in the interactions of tRNAs with prokaryotic and eukaryotic ribosomes are discussed with concomitant consideration of the extent of resemblance between molecular mechanisms of translation in eukaryotes and bacteria.

  15. Arabidopsis transcription factors: genome-wide comparative analysis among eukaryotes.

    Science.gov (United States)

    Riechmann, J L; Heard, J; Martin, G; Reuber, L; Jiang, C; Keddie, J; Adam, L; Pineda, O; Ratcliffe, O J; Samaha, R R; Creelman, R; Pilgrim, M; Broun, P; Zhang, J Z; Ghandehari, D; Sherman, B K; Yu, G

    2000-12-15

    The completion of the Arabidopsis thaliana genome sequence allows a comparative analysis of transcriptional regulators across the three eukaryotic kingdoms. Arabidopsis dedicates over 5% of its genome to code for more than 1500 transcription factors, about 45% of which are from families specific to plants. Arabidopsis transcription factors that belong to families common to all eukaryotes do not share significant similarity with those of the other kingdoms beyond the conserved DNA binding domains, many of which have been arranged in combinations specific to each lineage. The genome-wide comparison reveals the evolutionary generation of diversity in the regulation of transcription.

  16. Eukaryotic ribosome display with in situ DNA recovery.

    Science.gov (United States)

    He, Mingyue; Edwards, Bryan M; Kastelic, Damjana; Taussig, Michael J

    2012-01-01

    Ribosome display is a cell-free display technology for in vitro selection and optimisation of proteins from large diversified libraries. It operates through the formation of stable protein-ribosome-mRNA (PRM) complexes and selection of ligand-binding proteins, followed by DNA recovery from the selected genetic information. Both prokaryotic and eukaryotic ribosome display systems have been developed. In this chapter, we describe the eukaryotic rabbit reticulocyte method in which a distinct in situ single-primer RT-PCR procedure is used to recover DNA from the selected PRM complexes without the need for prior disruption of the ribosome.

  17. Characterization of prokaryotic and eukaryotic promoters usinghidden Markov models

    DEFF Research Database (Denmark)

    Pedersen, Anders Gorm; Baldi, Pierre; Brunak, Søren

    1996-01-01

    In this paper we utilize hidden Markov models (HMMs) and information theory to analyze prokaryotic and eukaryotic promoters. We perform this analysis with special emphasis on the fact that promoters are divided into a number of different classes, depending on which polymerase-associated factors...... that bind to them. We find that HMMs trained on such subclasses of Escherichia coli promoters (specifically, the so-called sigma-70 and sigma-54 classes) give an excellent classification of unknown promoters with respect to sigma-class. HMMs trained on eukaryotic sequences from human genes also model nicely...

  18. Characterization of prokaryotic and eukaryotic promoters using hidden Markov models

    DEFF Research Database (Denmark)

    Pedersen, Anders Gorm; Baldi, P.; Chauvin, Y.

    1996-01-01

    In this paper we utilize hidden Markov models (HMMs) and information theory to analyze prokaryotic and eukaryotic promoters. We perform this analysis with special emphasis on the fact that promoters are divided into a number of different classes, depending on which polymerase-associated factors...... that bind to them. We find that HMMs trained on such subclasses of Escherichia coli promoters (specifically, the so-called sigma 70 and sigma 54 classes) give an excellent classification of unknown promoters with respect to sigma-class. HMMs trained on eukaryotic sequences from human genes also model nicely...

  19. Distinct gene number-genome size relationships for eukaryotes and non-eukaryotes: gene content estimation for dinoflagellate genomes.

    Directory of Open Access Journals (Sweden)

    Yubo Hou

    Full Text Available The ability to predict gene content is highly desirable for characterization of not-yet sequenced genomes like those of dinoflagellates. Using data from completely sequenced and annotated genomes from phylogenetically diverse lineages, we investigated the relationship between gene content and genome size using regression analyses. Distinct relationships between log(10-transformed protein-coding gene number (Y' versus log(10-transformed genome size (X', genome size in kbp were found for eukaryotes and non-eukaryotes. Eukaryotes best fit a logarithmic model, Y' = ln(-46.200+22.678X', whereas non-eukaryotes a linear model, Y' = 0.045+0.977X', both with high significance (p0.91. Total gene number shows similar trends in both groups to their respective protein coding regressions. The distinct correlations reflect lower and decreasing gene-coding percentages as genome size increases in eukaryotes (82%-1% compared to higher and relatively stable percentages in prokaryotes and viruses (97%-47%. The eukaryotic regression models project that the smallest dinoflagellate genome (3x10(6 kbp contains 38,188 protein-coding (40,086 total genes and the largest (245x10(6 kbp 87,688 protein-coding (92,013 total genes, corresponding to 1.8% and 0.05% gene-coding percentages. These estimates do not likely represent extraordinarily high functional diversity of the encoded proteome but rather highly redundant genomes as evidenced by high gene copy numbers documented for various dinoflagellate species.

  20. Metabarcoding of benthic eukaryote communities predicts the ecological condition of estuaries

    International Nuclear Information System (INIS)

    Chariton, Anthony A.; Stephenson, Sarah; Morgan, Matthew J.; Steven, Andrew D.L.; Colloff, Matthew J.; Court, Leon N.; Hardy, Christopher M.

    2015-01-01

    DNA-derived measurements of biological composition have the potential to produce data covering all of life, and provide a tantalizing proposition for researchers and managers. We used metabarcoding to compare benthic eukaryote composition from five estuaries of varying condition. In contrast to traditional studies, we found biotic richness was greatest in the most disturbed estuary, with this being due to the large volume of extraneous material (i.e. run-off from aquaculture, agriculture and other catchment activities) being deposited in the system. In addition, we found strong correlations between composition and a number of environmental variables, including nutrients, pH and turbidity. A wide range of taxa responded to these environmental gradients, providing new insights into their sensitivities to natural and anthropogenic stressors. Metabarcoding has the capacity to bolster current monitoring techniques, enabling the decisions regarding ecological condition to be based on a more holistic view of biodiversity. - Highlights: • We used metabarcoding to examine the benthic eukaryote composition of five estuaries. • Biotic richness (based on MOTUs) was greater in the most impacted estuary. • Similarities among estuaries reflected their environmental condition. • Composition was strongly correlated with nutrients, turbidity and pH. • Metabarcoding can provide fast, comprehensive and ecologically informative data. - Using metabarcoding we were able discriminate benthos from five estuaries, and identify those taxa which responded negatively and positivity to the key environmental stressors

  1. An algorithm for detecting eukaryotic sequences in metagenomic ...

    Indian Academy of Sciences (India)

    species but also from accidental contamination from the genome of eukaryotic host cells. The latter scenario generally occurs in the case of host-associated metagenomes, e.g. microbes living in human gut. In such cases, one needs to identify and remove contaminating host DNA sequences, since the latter sequences will ...

  2. Potential of industrial biotechnology with cyanobacteria and eukaryotic microalgae

    NARCIS (Netherlands)

    Wijffels, R.H.; Kruse, O.; Hellingwerf, K.J.

    2013-01-01

    Both cyanobacteria and eukaryotic microalgae are promising organisms for sustainable production of bulk products such as food, feed, materials, chemicals and fuels. In this review we will summarize the potential and current biotechnological developments.Cyanobacteria are promising host organisms for

  3. A Synthetic Biology Framework for Programming Eukaryotic Transcription Functions

    Science.gov (United States)

    Khalil, Ahmad S.; Lu, Timothy K.; Bashor, Caleb J.; Ramirez, Cherie L.; Pyenson, Nora C.; Joung, J. Keith; Collins, James J.

    2013-01-01

    SUMMARY Eukaryotic transcription factors (TFs) perform complex and combinatorial functions within transcriptional networks. Here, we present a synthetic framework for systematically constructing eukaryotic transcription functions using artificial zinc fingers, modular DNA-binding domains found within many eukaryotic TFs. Utilizing this platform, we construct a library of orthogonal synthetic transcription factors (sTFs) and use these to wire synthetic transcriptional circuits in yeast. We engineer complex functions, such as tunable output strength and transcriptional cooperativity, by rationally adjusting a decomposed set of key component properties, e.g., DNA specificity, affinity, promoter design, protein-protein interactions. We show that subtle perturbations to these properties can transform an individual sTF between distinct roles (activator, cooperative factor, inhibitory factor) within a transcriptional complex, thus drastically altering the signal processing behavior of multi-input systems. This platform provides new genetic components for synthetic biology and enables bottom-up approaches to understanding the design principles of eukaryotic transcriptional complexes and networks. PMID:22863014

  4. Geminin: a major DNA replication safeguard in higher eukaryotes

    DEFF Research Database (Denmark)

    Melixetian, Marina; Helin, Kristian

    2004-01-01

    Eukaryotes have evolved multiple mechanisms to restrict DNA replication to once per cell cycle. These mechanisms prevent relicensing of origins of replication after initiation of DNA replication in S phase until the end of mitosis. Most of our knowledge of mechanisms controlling prereplication...

  5. Eu-Detect: An algorithm for detecting eukaryotic sequences in ...

    Indian Academy of Sciences (India)

    Supplementary figure 1. Plots depicting the classification accuracy of Eu-Detect with various combinations of. 'cumulative sequence count' (40K, 50K, 60K, 70K, 80K) and 'coverage threshold' (20%, 30%, 40%, 50%, 60%, 70%,. 80%). While blue bars represent Eu-Detect's average classification accuracy with eukaryotic ...

  6. Uncoupling of Sister Replisomes during Eukaryotic DNA Replication

    NARCIS (Netherlands)

    Yardimci, Hasan; Loveland, Anna B.; Habuchi, Satoshi; van Oijen, Antoine M.; Walter, Johannes C.

    2010-01-01

    The duplication of eukaryotic genomes involves the replication of DNA from multiple origins of replication. In S phase, two sister replisomes assemble at each active origin, and they replicate DNA in opposite directions. Little is known about the functional relationship between sister replisomes.

  7. Uniting sex and eukaryote origins in an emerging oxygenic world.

    Science.gov (United States)

    Gross, Jeferson; Bhattacharya, Debashish

    2010-08-23

    Theories about eukaryote origins (eukaryogenesis) need to provide unified explanations for the emergence of diverse complex features that define this lineage. Models that propose a prokaryote-to-eukaryote transition are gridlocked between the opposing "phagocytosis first" and "mitochondria as seed" paradigms, neither of which fully explain the origins of eukaryote cell complexity. Sex (outcrossing with meiosis) is an example of an elaborate trait not yet satisfactorily addressed in theories about eukaryogenesis. The ancestral nature of meiosis and its dependence on eukaryote cell biology suggest that the emergence of sex and eukaryogenesis were simultaneous and synergic and may be explained by a common selective pressure. We propose that a local rise in oxygen levels, due to cyanobacterial photosynthesis in ancient Archean microenvironments, was highly toxic to the surrounding biota. This selective pressure drove the transformation of an archaeal (archaebacterial) lineage into the first eukaryotes. Key is that oxygen might have acted in synergy with environmental stresses such as ultraviolet (UV) radiation and/or desiccation that resulted in the accumulation of reactive oxygen species (ROS). The emergence of eukaryote features such as the endomembrane system and acquisition of the mitochondrion are posited as strategies to cope with a metabolic crisis in the cell plasma membrane and the accumulation of ROS, respectively. Selective pressure for efficient repair of ROS/UV-damaged DNA drove the evolution of sex, which required cell-cell fusions, cytoskeleton-mediated chromosome movement, and emergence of the nuclear envelope. Our model implies that evolution of sex and eukaryogenesis were inseparable processes. Several types of data can be used to test our hypothesis. These include paleontological predictions, simulation of ancient oxygenic microenvironments, and cell biological experiments with Archaea exposed to ROS and UV stresses. Studies of archaeal conjugation

  8. Patterns of intron gain and conservation in eukaryotic genes

    Directory of Open Access Journals (Sweden)

    Wolf Yuri I

    2007-10-01

    Full Text Available Abstract Background: The presence of introns in protein-coding genes is a universal feature of eukaryotic genome organization, and the genes of multicellular eukaryotes, typically, contain multiple introns, a substantial fraction of which share position in distant taxa, such as plants and animals. Depending on the methods and data sets used, researchers have reached opposite conclusions on the causes of the high fraction of shared introns in orthologous genes from distant eukaryotes. Some studies conclude that shared intron positions reflect, almost entirely, a remarkable evolutionary conservation, whereas others attribute it to parallel gain of introns. To resolve these contradictions, it is crucial to analyze the evolution of introns by using a model that minimally relies on arbitrary assumptions. Results: We developed a probabilistic model of evolution that allows for variability of intron gain and loss rates over branches of the phylogenetic tree, individual genes, and individual sites. Applying this model to an extended set of conserved eukaryotic genes, we find that parallel gain, on average, accounts for only ~8% of the shared intron positions. However, the distribution of parallel gains over the phylogenetic tree of eukaryotes is highly non-uniform. There are, practically, no parallel gains in closely related lineages, whereas for distant lineages, such as animals and plants, parallel gains appear to contribute up to 20% of the shared intron positions. In accord with these findings, we estimated that ancestral introns have a high probability to be retained in extant genomes, and conversely, that a substantial fraction of extant introns have retained their positions since the early stages of eukaryotic evolution. In addition, the density of sites that are available for intron insertion is estimated to be, approximately, one in seven basepairs. Conclusion: We obtained robust estimates of the contribution of parallel gain to the observed

  9. UniProt search blastx result: AK288448 [KOME

    Lifescience Database Archive (English)

    Full Text Available AK288448 J090034N14 P48545|IRK5_MOUSE G protein-activated inward rectifier potassiu...m channel 4 (GIRK4) (Potassium channel, inwardly rectifying subfamily J member 5) (Inward rectifier K(+) cha...nnel Kir3.4) (Heart KATP channel) (KATP-1) (Cardiac inward rectifier) (CIR) - Mus musculus (Mouse) 0 ...

  10. Genome-wide analysis of eukaryote thaumatin-like proteins (TLPs with an emphasis on poplar

    Directory of Open Access Journals (Sweden)

    Duplessis Sébastien

    2011-02-01

    Full Text Available Abstract Background Plant inducible immunity includes the accumulation of a set of defense proteins during infection called pathogenesis-related (PR proteins, which are grouped into families termed PR-1 to PR-17. The PR-5 family is composed of thaumatin-like proteins (TLPs, which are responsive to biotic and abiotic stress and are widely studied in plants. TLPs were also recently discovered in fungi and animals. In the poplar genome, TLPs are over-represented compared with annual species and their transcripts strongly accumulate during stress conditions. Results Our analysis of the poplar TLP family suggests that the expansion of this gene family was followed by diversification, as differences in expression patterns and predicted properties correlate with phylogeny. In particular, we identified a clade of poplar TLPs that cluster to a single 350 kb locus of chromosome I and that are up-regulated by poplar leaf rust infection. A wider phylogenetic analysis of eukaryote TLPs - including plant, animal and fungi sequences - shows that TLP gene content and diversity increased markedly during land plant evolution. Mapping the reported functions of characterized TLPs to the eukaryote phylogenetic tree showed that antifungal or glycan-lytic properties are widespread across eukaryote phylogeny, suggesting that these properties are shared by most TLPs and are likely associated with the presence of a conserved acidic cleft in their 3D structure. Also, we established an exhaustive catalog of TLPs with atypical architectures such as small-TLPs, TLP-kinases and small-TLP-kinases, which have potentially developed alternative functions (such as putative receptor kinases for pathogen sensing and signaling. Conclusion Our study, based on the most recent plant genome sequences, provides evidence for TLP gene family diversification during land plant evolution. We have shown that the diverse functions described for TLPs are not restricted to specific clades but seem

  11. Anionic lipids and the maintenance of membrane electrostatics in eukaryotes.

    Science.gov (United States)

    Platre, Matthieu Pierre; Jaillais, Yvon

    2017-02-01

    A wide range of signaling processes occurs at the cell surface through the reversible association of proteins from the cytosol to the plasma membrane. Some low abundant lipids are enriched at the membrane of specific compartments and thereby contribute to the identity of cell organelles by acting as biochemical landmarks. Lipids also influence membrane biophysical properties, which emerge as an important feature in specifying cellular territories. Such parameters are crucial for signal transduction and include lipid packing, membrane curvature and electrostatics. In particular, membrane electrostatics specifies the identity of the plasma membrane inner leaflet. Membrane surface charges are carried by anionic phospholipids, however the exact nature of the lipid(s) that powers the plasma membrane electrostatic field varies among eukaryotes and has been hotly debated during the last decade. Herein, we discuss the role of anionic lipids in setting up plasma membrane electrostatics and we compare similarities and differences that were found in different eukaryotic cells.

  12. Regulated eukaryotic DNA replication origin firing with purified proteins.

    Science.gov (United States)

    Yeeles, Joseph T P; Deegan, Tom D; Janska, Agnieszka; Early, Anne; Diffley, John F X

    2015-03-26

    Eukaryotic cells initiate DNA replication from multiple origins, which must be tightly regulated to promote precise genome duplication in every cell cycle. To accomplish this, initiation is partitioned into two temporally discrete steps: a double hexameric minichromosome maintenance (MCM) complex is first loaded at replication origins during G1 phase, and then converted to the active CMG (Cdc45-MCM-GINS) helicase during S phase. Here we describe the reconstitution of budding yeast DNA replication initiation with 16 purified replication factors, made from 42 polypeptides. Origin-dependent initiation recapitulates regulation seen in vivo. Cyclin-dependent kinase (CDK) inhibits MCM loading by phosphorylating the origin recognition complex (ORC) and promotes CMG formation by phosphorylating Sld2 and Sld3. Dbf4-dependent kinase (DDK) promotes replication by phosphorylating MCM, and can act either before or after CDK. These experiments define the minimum complement of proteins, protein kinase substrates and co-factors required for regulated eukaryotic DNA replication.

  13. Complex multicellular functions at a unicellular eukaryote level: Learning, memory, and immunity.

    Science.gov (United States)

    Csaba, György

    2017-06-01

    According to experimental data, eukaryote unicellulars are able to learn, have immunity and memory. Learning is carried out in a very primitive form, and the memory is not neural but an epigenetic one. However, this epigenetic memory, which is well justified by the presence and manifestation of hormonal imprinting, is strong and permanent in the life of cell and also in its progenies. This memory is epigenetically executed by the alteration and fixation of methylation pattern of genes without changes in base sequences. The immunity of unicellulars is based on self/non-self discrimination, which leads to the destruction of non-self invaders and utilization of them as nourishment (by phagocytosis). The tools of learning, memory, and immunity of unicellulars are uniformly found in plasma membrane receptors, which formed under the effect of dynamic receptor pattern generation, suggested by Koch et al., and this is the basis of hormonal imprinting, by which the encounter between a chemical substance and the cell is specifically memorized. The receptors and imprinting are also used in the later steps of evolution up to mammals (including man) in each mentioned functions. This means that learning, memory, and immunity can be deduced to a unicellular eukaryote level.

  14. Polyglutamine repeats are associated to specific sequence biases that are conserved among eukaryotes.

    Directory of Open Access Journals (Sweden)

    Matteo Ramazzotti

    Full Text Available Nine human neurodegenerative diseases, including Huntington's disease and several spinocerebellar ataxia, are associated to the aggregation of proteins comprising an extended tract of consecutive glutamine residues (polyQs once it exceeds a certain length threshold. This event is believed to be the consequence of the expansion of polyCAG codons during the replication process. This is in apparent contradiction with the fact that many polyQs-containing proteins remain soluble and are encoded by invariant genes in a number of eukaryotes. The latter suggests that polyQs expansion and/or aggregation might be counter-selected through a genetic and/or protein context. To identify this context, we designed a software that scrutinize entire proteomes in search for imperfect polyQs. The nature of residues flanking the polyQs and that of residues other than Gln within polyQs (insertions were assessed. We discovered strong amino acid residue biases robustly associated to polyQs in the 15 eukaryotic proteomes we examined, with an over-representation of Pro, Leu and His and an under-representation of Asp, Cys and Gly amino acid residues. These biases are conserved amongst unrelated proteins and are independent of specific functional classes. Our findings suggest that specific residues have been co-selected with polyQs during evolution. We discuss the possible selective pressures responsible of the observed biases.

  15. The MCM Helicase Motor of the Eukaryotic Replisome.

    Science.gov (United States)

    Abid Ali, Ferdos; Costa, Alessandro

    2016-05-08

    The MCM motor of the CMG helicase powers ahead of the eukaryotic replication machinery to unwind DNA, in a process that requires ATP hydrolysis. The reconstitution of DNA replication in vitro has established the succession of events that lead to replication origin activation by the MCM and recent studies have started to elucidate the structural basis of duplex DNA unwinding. Despite the exciting progress, how the MCM translocates on DNA remains a matter of debate. Copyright © 2016. Published by Elsevier Ltd.

  16. Biological Influence of Deuterium on Procariotic and Eukaryotic Cells

    OpenAIRE

    Oleg Mosin; Ignat Ignatov

    2014-01-01

    Biologic influence of deuterium (D) on cells of various taxonomic groups of prokaryotic and eukaryotic microorganisms realizing methylotrophic, chemoheterotrophic, photo-organotrophic, and photosynthetic ways of assimilation of carbon substrates are investigated at growth on media with heavy water (D2О). The method of step by step adaptation technique of cells to D2О was developed, consisting in plating of cells on 2 % agarose nutrient media containing increasing gradient of concentration of ...

  17. An Evolutionary Framework for Understanding the Origin of Eukaryotes

    OpenAIRE

    Neil W. Blackstone

    2016-01-01

    Two major obstacles hinder the application of evolutionary theory to the origin of eukaryotes. The first is more apparent than real?the endosymbiosis that led to the mitochondrion is often described as ?non-Darwinian? because it deviates from the incremental evolution championed by the modern synthesis. Nevertheless, endosymbiosis can be accommodated by a multi-level generalization of evolutionary theory, which Darwin himself pioneered. The second obstacle is more serious?all of the major fea...

  18. Gram-Negative Bacterial Sensors for Eukaryotic Signal Molecules

    Directory of Open Access Journals (Sweden)

    Olivier Lesouhaitier

    2009-09-01

    Full Text Available Ample evidence exists showing that eukaryotic signal molecules synthesized and released by the host can activate the virulence of opportunistic pathogens. The sensitivity of prokaryotes to host signal molecules requires the presence of bacterial sensors. These prokaryotic sensors, or receptors, have a double function: stereospecific recognition in a complex environment and transduction of the message in order to initiate bacterial physiological modifications. As messengers are generally unable to freely cross the bacterial membrane, they require either the presence of sensors anchored in the membrane or transporters allowing direct recognition inside the bacterial cytoplasm. Since the discovery of quorum sensing, it was established that the production of virulence factors by bacteria is tightly growth-phase regulated. It is now obvious that expression of bacterial virulence is also controlled by detection of the eukaryotic messengers released in the micro-environment as endocrine or neuro-endocrine modulators. In the presence of host physiological stress many eukaryotic factors are released and detected by Gram-negative bacteria which in return rapidly adapt their physiology. For instance, Pseudomonas aeruginosa can bind elements of the host immune system such as interferon-γ and dynorphin and then through quorum sensing circuitry enhance its virulence. Escherichia coli sensitivity to the neurohormones of the catecholamines family appears relayed by a recently identified bacterial adrenergic receptor. In the present review, we will describe the mechanisms by which various eukaryotic signal molecules produced by host may activate Gram-negative bacteria virulence. Particular attention will be paid to Pseudomonas, a genus whose representative species, P. aeruginosa, is a common opportunistic pathogen. The discussion will be particularly focused on the pivotal role played by these new types of pathogen sensors from the sensing to the transduction

  19. Replication and Transcription of Eukaryotic DNA in Esherichia coli

    Science.gov (United States)

    Morrow, John F.; Cohen, Stanley N.; Chang, Annie C. Y.; Boyer, Herbert W.; Goodman, Howard M.; Helling, Robert B.

    1974-01-01

    Fragments of amplified Xenopus laevis DNA, coding for 18S and 28S ribosomal RNA and generated by EcoRI restriction endonuclease, have been linked in vitro to the bacterial plasmid pSC101; and the recombinant molecular species have been introduced into E. coli by transformation. These recombinant plasmids, containing both eukaryotic and prokaryotic DNA, replicate stably in E. coli. RNA isolated from E. coli minicells harboring the plasmids hybridizes to amplified X. laevis rDNA. Images PMID:4600264

  20. Extreme Diversity of Diplonemid Eukaryotes in the Ocean

    Czech Academy of Sciences Publication Activity Database

    Flegontova, Olga; Flegontov, Pavel; Malviya, S.; Audic, S.; Wincker, P.; de Vargas, C.; Bowler, C.; Lukeš, Julius; Horák, Aleš

    2016-01-01

    Roč. 26, č. 22 (2016), s. 3060-3065 ISSN 0960-9822 R&D Projects: GA ČR GPP506/12/P931; GA ČR(CZ) GA14-23986S Institutional support: RVO:60077344 Keywords : virus-sized particles * microbial eukaryotes * sea-floor * phytoplankton * communities * euglenozoa * dispersal * ecosystem Subject RIV: EG - Zoology Impact factor: 8.851, year: 2016

  1. diArk – a resource for eukaryotic genome research

    Directory of Open Access Journals (Sweden)

    Kollmar Martin

    2007-04-01

    Full Text Available Abstract Background The number of completed eukaryotic genome sequences and cDNA projects has increased exponentially in the past few years although most of them have not been published yet. In addition, many microarray analyses yielded thousands of sequenced EST and cDNA clones. For the researcher interested in single gene analyses (from a phylogenetic, a structural biology or other perspective it is therefore important to have up-to-date knowledge about the various resources providing primary data. Description The database is built around 3 central tables: species, sequencing projects and publications. The species table contains commonly and alternatively used scientific names, common names and the complete taxonomic information. For projects the sequence type and links to species project web-sites and species homepages are stored. All publications are linked to projects. The web-interface provides comprehensive search modules with detailed options and three different views of the selected data. We have especially focused on developing an elaborate taxonomic tree search tool that allows the user to instantaneously identify e.g. the closest relative to the organism of interest. Conclusion We have developed a database, called diArk, to store, organize, and present the most relevant information about completed genome projects and EST/cDNA data from eukaryotes. Currently, diArk provides information about 415 eukaryotes, 823 sequencing projects, and 248 publications.

  2. Biotransformation of arsenic by a Yellowstone thermoacidophilic eukaryotic alga.

    Science.gov (United States)

    Qin, Jie; Lehr, Corinne R; Yuan, Chungang; Le, X Chris; McDermott, Timothy R; Rosen, Barry P

    2009-03-31

    Arsenic is the most common toxic substance in the environment, ranking first on the Superfund list of hazardous substances. It is introduced primarily from geochemical sources and is acted on biologically, creating an arsenic biogeocycle. Geothermal environments are known for their elevated arsenic content and thus provide an excellent setting in which to study microbial redox transformations of arsenic. To date, most studies of microbial communities in geothermal environments have focused on Bacteria and Archaea, with little attention to eukaryotic microorganisms. Here, we show the potential of an extremophilic eukaryotic alga of the order Cyanidiales to influence arsenic cycling at elevated temperatures. Cyanidioschyzon sp. isolate 5508 oxidized arsenite [As(III)] to arsenate [As(V)], reduced As(V) to As(III), and methylated As(III) to form trimethylarsine oxide (TMAO) and dimethylarsenate [DMAs(V)]. Two arsenic methyltransferase genes, CmarsM7 and CmarsM8, were cloned from this organism and demonstrated to confer resistance to As(III) in an arsenite hypersensitive strain of Escherichia coli. The 2 recombinant CmArsMs were purified and shown to transform As(III) into monomethylarsenite, DMAs(V), TMAO, and trimethylarsine gas, with a T(opt) of 60-70 degrees C. These studies illustrate the importance of eukaryotic microorganisms to the biogeochemical cycling of arsenic in geothermal systems, offer a molecular explanation for how these algae tolerate arsenic in their environment, and provide the characterization of algal methyltransferases.

  3. Ubiquitination dynamics in the early-branching eukaryote Giardia intestinalis

    Science.gov (United States)

    Niño, Carlos A; Chaparro, Jenny; Soffientini, Paolo; Polo, Simona; Wasserman, Moises

    2013-01-01

    Ubiquitination is a highly dynamic and versatile posttranslational modification that regulates protein function, stability, and interactions. To investigate the roles of ubiquitination in a primitive eukaryotic lineage, we utilized the early-branching eukaryote Giardia intestinalis. Using a combination of biochemical, immunofluorescence-based, and proteomics approaches, we assessed the ubiquitination status during the process of differentiation in Giardia. We observed that different types of ubiquitin modifications present specific cellular and temporal distribution throughout the Giardia life cycle from trophozoites to cyst maturation. Ubiquitin signal was detected in the wall of mature cysts, and enzymes implicated in cyst wall biogenesis were identified as substrates for ubiquitination. Interestingly, inhibition of proteasome activity did not affect trophozoite replication and differentiation, while it caused a decrease in cyst viability, arguing for proteasome involvement in cyst wall maturation. Using a proteomics approach, we identified around 200 high-confidence ubiquitinated candidates that vary their ubiquitination status during differentiation. Our results indicate that ubiquitination is critical for several cellular processes in this primitive eukaryote. PMID:23613346

  4. Gene Transfer in Eukaryotic Cells Using Activated Dendrimers

    Science.gov (United States)

    Dennig, Jörg

    Gene transfer into eukaryotic cells plays an important role in cell biology. Over the last 30 years a number of transfection methods have been developed to mediate gene transfer into eukaryotic cells. Classical methods include co-precipitation of DNA with calcium phosphate, charge-dependent precipitation of DNA with DEAE-dextran, electroporation of nucleic acids, and formation of transfection complexes between DNA and cationic liposomes. Gene transfer technologies based on activated PAMAM-dendrimers provide another class of transfection reagents. PAMAM-dendrimers are highly branched, spherical molecules. Activation of newly synthesized dendrimers involves hydrolytic removal of some of the branches, and results in a molecule with a higher degree of flexibility. Activated dendrimers assemble DNA into compact structures via charge interactions. Activated dendrimer - DNA complexes bind to the cell membrane of eukaryotic cells, and are transported into the cell by non-specific endocytosis. A structural model of the activated dendrimer - DNA complex and a potential mechanism for its uptake into cells will be discussed.

  5. Enzymes involved in organellar DNA replication in photosynthetic eukaryotes.

    Science.gov (United States)

    Moriyama, Takashi; Sato, Naoki

    2014-01-01

    Plastids and mitochondria possess their own genomes. Although the replication mechanisms of these organellar genomes remain unclear in photosynthetic eukaryotes, several organelle-localized enzymes related to genome replication, including DNA polymerase, DNA primase, DNA helicase, DNA topoisomerase, single-stranded DNA maintenance protein, DNA ligase, primer removal enzyme, and several DNA recombination-related enzymes, have been identified. In the reference Eudicot plant Arabidopsis thaliana, the replication-related enzymes of plastids and mitochondria are similar because many of them are dual targeted to both organelles, whereas in the red alga Cyanidioschyzon merolae, plastids and mitochondria contain different replication machinery components. The enzymes involved in organellar genome replication in green plants and red algae were derived from different origins, including proteobacterial, cyanobacterial, and eukaryotic lineages. In the present review, we summarize the available data for enzymes related to organellar genome replication in green plants and red algae. In addition, based on the type and distribution of replication enzymes in photosynthetic eukaryotes, we discuss the transitional history of replication enzymes in the organelles of plants.

  6. Mechanisms and regulation of DNA replication initiation in eukaryotes.

    Science.gov (United States)

    Parker, Matthew W; Botchan, Michael R; Berger, James M

    2017-04-01

    Cellular DNA replication is initiated through the action of multiprotein complexes that recognize replication start sites in the chromosome (termed origins) and facilitate duplex DNA melting within these regions. In a typical cell cycle, initiation occurs only once per origin and each round of replication is tightly coupled to cell division. To avoid aberrant origin firing and re-replication, eukaryotes tightly regulate two events in the initiation process: loading of the replicative helicase, MCM2-7, onto chromatin by the origin recognition complex (ORC), and subsequent activation of the helicase by its incorporation into a complex known as the CMG. Recent work has begun to reveal the details of an orchestrated and sequential exchange of initiation factors on DNA that give rise to a replication-competent complex, the replisome. Here, we review the molecular mechanisms that underpin eukaryotic DNA replication initiation - from selecting replication start sites to replicative helicase loading and activation - and describe how these events are often distinctly regulated across different eukaryotic model organisms.

  7. Origin of phagotrophic eukaryotes as social cheaters in microbial biofilms

    Directory of Open Access Journals (Sweden)

    Jékely Gáspár

    2007-01-01

    Full Text Available Abstract Background The origin of eukaryotic cells was one of the most dramatic evolutionary transitions in the history of life. It is generally assumed that eukaryotes evolved later then prokaryotes by the transformation or fusion of prokaryotic lineages. However, as yet there is no consensus regarding the nature of the prokaryotic group(s ancestral to eukaryotes. Regardless of this, a hardly debatable fundamental novel characteristic of the last eukaryotic common ancestor was the ability to exploit prokaryotic biomass by the ingestion of entire cells, i.e. phagocytosis. The recent advances in our understanding of the social life of prokaryotes may help to explain the origin of this form of total exploitation. Presentation of the hypothesis Here I propose that eukaryotic cells originated in a social environment, a differentiated microbial mat or biofilm that was maintained by the cooperative action of its members. Cooperation was costly (e.g. the production of developmental signals or an extracellular matrix but yielded benefits that increased the overall fitness of the social group. I propose that eukaryotes originated as selfish cheaters that enjoyed the benefits of social aggregation but did not contribute to it themselves. The cheaters later evolved into predators that lysed other cells and eventually became professional phagotrophs. During several cycles of social aggregation and dispersal the number of cheaters was contained by a chicken game situation, i.e. reproductive success of cheaters was high when they were in low abundance but was reduced when they were over-represented. Radical changes in cell structure, including the loss of the rigid prokaryotic cell wall and the development of endomembranes, allowed the protoeukaryotes to avoid cheater control and to exploit nutrients more efficiently. Cellular changes were buffered by both the social benefits and the protective physico-chemical milieu of the interior of biofilms. Symbiosis

  8. Testing strong interaction theories

    International Nuclear Information System (INIS)

    Ellis, J.

    1979-01-01

    The author discusses possible tests of the current theories of the strong interaction, in particular, quantum chromodynamics. High energy e + e - interactions should provide an excellent means of studying the strong force. (W.D.L.)

  9. Molecular analyses reveal high levels of eukaryotic richness associated with enigmatic deep-sea protists (Komokiacea)

    DEFF Research Database (Denmark)

    Lecroq, Beatrice; Gooday, Andrew John; Cedhagen, Tomas

    2009-01-01

    Komokiaceans are testate agglutinated protists, extremely diverse and abundant in the deep sea. About 40 species are described and share the same main morpholog- ical feature: a test consisting of narrow branching tubules forming a complex system. In some species, the interstices between the tubu......Komokiaceans are testate agglutinated protists, extremely diverse and abundant in the deep sea. About 40 species are described and share the same main morpholog- ical feature: a test consisting of narrow branching tubules forming a complex system. In some species, the interstices between...... suggest strongly that komokiaceans, and probably many other large testate protists, provide a habitat structure for a large spectrum of eukaryotes, significantly contributing to maintaining the biodiversity of micro- and meiofaunal communities in the deep sea....

  10. Origin and evolution of the self-organizing cytoskeleton in the network of eukaryotic organelles.

    Science.gov (United States)

    Jékely, Gáspár

    2014-09-02

    The eukaryotic cytoskeleton evolved from prokaryotic cytomotive filaments. Prokaryotic filament systems show bewildering structural and dynamic complexity and, in many aspects, prefigure the self-organizing properties of the eukaryotic cytoskeleton. Here, the dynamic properties of the prokaryotic and eukaryotic cytoskeleton are compared, and how these relate to function and evolution of organellar networks is discussed. The evolution of new aspects of filament dynamics in eukaryotes, including severing and branching, and the advent of molecular motors converted the eukaryotic cytoskeleton into a self-organizing "active gel," the dynamics of which can only be described with computational models. Advances in modeling and comparative genomics hold promise of a better understanding of the evolution of the self-organizing cytoskeleton in early eukaryotes, and its role in the evolution of novel eukaryotic functions, such as amoeboid motility, mitosis, and ciliary swimming. Copyright © 2014 Cold Spring Harbor Laboratory Press; all rights reserved.

  11. Insights into the Initiation of Eukaryotic DNA Replication.

    Science.gov (United States)

    Bruck, Irina; Perez-Arnaiz, Patricia; Colbert, Max K; Kaplan, Daniel L

    2015-01-01

    The initiation of DNA replication is a highly regulated event in eukaryotic cells to ensure that the entire genome is copied once and only once during S phase. The primary target of cellular regulation of eukaryotic DNA replication initiation is the assembly and activation of the replication fork helicase, the 11-subunit assembly that unwinds DNA at a replication fork. The replication fork helicase, called CMG for Cdc45-Mcm2-7, and GINS, assembles in S phase from the constituent Cdc45, Mcm2-7, and GINS proteins. The assembly and activation of the CMG replication fork helicase during S phase is governed by 2 S-phase specific kinases, CDK and DDK. CDK stimulates the interaction between Sld2, Sld3, and Dpb11, 3 initiation factors that are each required for the initiation of DNA replication. DDK, on the other hand, phosphorylates the Mcm2, Mcm4, and Mcm6 subunits of the Mcm2-7 complex. Sld3 recruits Cdc45 to Mcm2-7 in a manner that depends on DDK, and recent work suggests that Sld3 binds directly to Mcm2-7 and also to single-stranded DNA. Furthermore, recent work demonstrates that Sld3 and its human homolog Treslin substantially stimulate DDK phosphorylation of Mcm2. These data suggest that the initiation factor Sld3/Treslin coordinates the assembly and activation of the eukaryotic replication fork helicase by recruiting Cdc45 to Mcm2-7, stimulating DDK phosphorylation of Mcm2, and binding directly to single-stranded DNA as the origin is melted.

  12. [Structure and evolution of the eukaryotic FANCJ-like proteins].

    Science.gov (United States)

    Wuhe, Jike; Zefeng, Wu; Sanhong, Fan; Xuguang, Xi

    2015-02-01

    The FANCJ-like protein family is a class of ATP-dependent helicases that can catalytically unwind duplex DNA along the 5'-3' direction. It is involved in the processes of DNA damage repair, homologous recombination and G-quadruplex DNA unwinding, and plays a critical role in maintaining genome integrity. In this study, we systemically analyzed FNACJ-like proteins from 47 eukaryotic species and discussed their sequences diversity, origin and evolution, motif organization patterns and spatial structure differences. Four members of FNACJ-like proteins, including XPD, CHL1, RTEL1 and FANCJ, were found in eukaryotes, but some of them were seriously deficient in most fungi and some insects. For example, the Zygomycota fungi lost RTEL1, Basidiomycota and Ascomycota fungi lost RTEL1 and FANCJ, and Diptera insect lost FANCJ. FANCJ-like proteins contain canonical motor domains HD1 and HD2, and the HD1 domain further integrates with three unique domains Fe-S, Arch and Extra-D. Fe-S and Arch domains are relatively conservative in all members of the family, but the Extra-D domain is lost in XPD and differs from one another in rest members. There are 7, 10 and 2 specific motifs found from the three unique domains respectively, while 5 and 12 specific motifs are found from HD1 and HD2 domains except the conserved motifs reported previously. By analyzing the arrangement pattern of these specific motifs, we found that RTEL1 and FANCJ are more closer and share two specific motifs Vb2 and Vc in HD2 domain, which are likely related with their G-quadruplex DNA unwinding activity. The evidence of evolution showed that FACNJ-like proteins were originated from a helicase, which has a HD1 domain inserted by extra Fe-S domain and Arch domain. By three continuous gene duplication events and followed specialization, eukaryotes finally possessed the current four members of FANCJ-like proteins.

  13. Challenges in Whole-Genome Annotation of Pyrosequenced Eukaryotic Genomes

    Energy Technology Data Exchange (ETDEWEB)

    Kuo, Alan; Grigoriev, Igor

    2009-04-17

    Pyrosequencing technologies such as 454/Roche and Solexa/Illumina vastly lower the cost of nucleotide sequencing compared to the traditional Sanger method, and thus promise to greatly expand the number of sequenced eukaryotic genomes. However, the new technologies also bring new challenges such as shorter reads and new kinds and higher rates of sequencing errors, which complicate genome assembly and gene prediction. At JGI we are deploying 454 technology for the sequencing and assembly of ever-larger eukaryotic genomes. Here we describe our first whole-genome annotation of a purely 454-sequenced fungal genome that is larger than a yeast (>30 Mbp). The pezizomycotine (filamentous ascomycote) Aspergillus carbonarius belongs to the Aspergillus section Nigri species complex, members of which are significant as platforms for bioenergy and bioindustrial technology, as members of soil microbial communities and players in the global carbon cycle, and as agricultural toxigens. Application of a modified version of the standard JGI Annotation Pipeline has so far predicted ~;;10k genes. ~;;12percent of these preliminary annotations suffer a potential frameshift error, which is somewhat higher than the ~;;9percent rate in the Sanger-sequenced and conventionally assembled and annotated genome of fellow Aspergillus section Nigri member A. niger. Also,>90percent of A. niger genes have potential homologs in the A. carbonarius preliminary annotation. Weconclude, and with further annotation and comparative analysis expect to confirm, that 454 sequencing strategies provide a promising substrate for annotation of modestly sized eukaryotic genomes. We will also present results of annotation of a number of other pyrosequenced fungal genomes of bioenergy interest.

  14. Recognition of extremophilic archaeal viruses by eukaryotic cells

    DEFF Research Database (Denmark)

    Uldahl, Kristine Buch; Wu, Linping; Hall, Arnaldur

    2016-01-01

    followed viral uptake, intracellular trafficking and cell viability in human endothelial cells of brain (hCMEC/D3 cells) and umbilical vein (HUVEC) origin. Whereas SMV1 is efficiently internalized into both types of human cells, SSV2 differentiates between HUVECs and hCMEC/D3 cells, thus opening a path......Viruses from the third domain of life, Archaea, exhibit unusual features including extreme stability that allow their survival in harsh environments. In addition, these species have never been reported to integrate into human or any other eukaryotic genomes, and could thus serve for exploration...

  15. Eukaryotic snoRNAs: a paradigm for gene expression flexibility.

    Science.gov (United States)

    Dieci, Giorgio; Preti, Milena; Montanini, Barbara

    2009-08-01

    Small nucleolar RNAs (snoRNAs) are one of the most ancient and numerous families of non-protein-coding RNAs (ncRNAs). The main function of snoRNAs - to guide site-specific rRNA modification - is the same in Archaea and all eukaryotic lineages. In contrast, as revealed by recent genomic and RNomic studies, their genomic organization and expression strategies are the most varied. Seemingly snoRNA coding units have adopted, in the course of evolution, all the possible ways of being transcribed, thus providing a unique paradigm of gene expression flexibility. By focusing on representative fungal, plant and animal genomes, we review here all the documented types of snoRNA gene organization and expression, and we provide a comprehensive account of snoRNA expressional freedom by precisely estimating the frequency, in each genome, of each type of genomic organization. We finally discuss the relevance of snoRNA genomic studies for our general understanding of ncRNA family evolution and expression in eukaryotes.

  16. Universal Temporal Profile of Replication Origin Activation in Eukaryotes

    Science.gov (United States)

    Goldar, Arach

    2011-03-01

    The complete and faithful transmission of eukaryotic genome to daughter cells involves the timely duplication of mother cell's DNA. DNA replication starts at multiple chromosomal positions called replication origin. From each activated replication origin two replication forks progress in opposite direction and duplicate the mother cell's DNA. While it is widely accepted that in eukaryotic organisms replication origins are activated in a stochastic manner, little is known on the sources of the observed stochasticity. It is often associated to the population variability to enter S phase. We extract from a growing Saccharomyces cerevisiae population the average rate of origin activation in a single cell by combining single molecule measurements and a numerical deconvolution technique. We show that the temporal profile of the rate of origin activation in a single cell is similar to the one extracted from a replicating cell population. Taking into account this observation we exclude the population variability as the origin of observed stochasticity in origin activation. We confirm that the rate of origin activation increases in the early stage of S phase and decreases at the latter stage. The population average activation rate extracted from single molecule analysis is in prefect accordance with the activation rate extracted from published micro-array data, confirming therefore the homogeneity and genome scale invariance of dynamic of replication process. All these observations point toward a possible role of replication fork to control the rate of origin activation.

  17. Marine biofilm bacteria evade eukaryotic predation by targeted chemical defense.

    Directory of Open Access Journals (Sweden)

    Carsten Matz

    Full Text Available Many plants and animals are defended from predation or herbivory by inhibitory secondary metabolites, which in the marine environment are very common among sessile organisms. Among bacteria, where there is the greatest metabolic potential, little is known about chemical defenses against bacterivorous consumers. An emerging hypothesis is that sessile bacterial communities organized as biofilms serve as bacterial refuge from predation. By testing growth and survival of two common bacterivorous nanoflagellates, we find evidence that chemically mediated resistance against protozoan predators is common among biofilm populations in a diverse set of marine bacteria. Using bioassay-guided chemical and genetic analysis, we identified one of the most effective antiprotozoal compounds as violacein, an alkaloid that we demonstrate is produced predominately within biofilm cells. Nanomolar concentrations of violacein inhibit protozoan feeding by inducing a conserved eukaryotic cell death program. Such biofilm-specific chemical defenses could contribute to the successful persistence of biofilm bacteria in various environments and provide the ecological and evolutionary context for a number of eukaryote-targeting bacterial metabolites.

  18. Introns Protect Eukaryotic Genomes from Transcription-Associated Genetic Instability.

    Science.gov (United States)

    Bonnet, Amandine; Grosso, Ana R; Elkaoutari, Abdessamad; Coleno, Emeline; Presle, Adrien; Sridhara, Sreerama C; Janbon, Guilhem; Géli, Vincent; de Almeida, Sérgio F; Palancade, Benoit

    2017-08-17

    Transcription is a source of genetic instability that can notably result from the formation of genotoxic DNA:RNA hybrids, or R-loops, between the nascent mRNA and its template. Here we report an unexpected function for introns in counteracting R-loop accumulation in eukaryotic genomes. Deletion of endogenous introns increases R-loop formation, while insertion of an intron into an intronless gene suppresses R-loop accumulation and its deleterious impact on transcription and recombination in yeast. Recruitment of the spliceosome onto the mRNA, but not splicing per se, is shown to be critical to attenuate R-loop formation and transcription-associated genetic instability. Genome-wide analyses in a number of distant species differing in their intron content, including human, further revealed that intron-containing genes and the intron-richest genomes are best protected against R-loop accumulation and subsequent genetic instability. Our results thereby provide a possible rationale for the conservation of introns throughout the eukaryotic lineage. Copyright © 2017 Elsevier Inc. All rights reserved.

  19. Abortion: Strong's counterexamples fail

    DEFF Research Database (Denmark)

    Di Nucci, Ezio

    2009-01-01

    This paper shows that the counterexamples proposed by Strong in 2008 in the Journal of Medical Ethics to Marquis's argument against abortion fail. Strong's basic idea is that there are cases--for example, terminally ill patients--where killing an adult human being is prima facie seriously morally...

  20. Strong Langmuir turbulence

    International Nuclear Information System (INIS)

    Goldman, M.V.

    1984-01-01

    After a brief discussion of beam-excited Langmuir turbulence in the solar wind, we explain the criteria for wave-particle, three-wave and strong turbulence interactions. We then present the results of a numerical integration of the Zakharov equations, which describe the strong turbulence saturation of a weak (low-density) high energy, bump-on-tail beam instability. (author)

  1. Guidelines for the use and interpretation of assays for monitoring autophagy in higher eukaryotes

    Science.gov (United States)

    Klionsky, Daniel J.; Abeliovich, Hagai; Agostinis, Patrizia; Agrawal, Devendra K.; Aliev, Gjumrakch; Askew, David S.; Baba, Misuzu; Baehrecke, Eric H.; Bahr, Ben A.; Ballabio, Andrea; Bamber, Bruce A.; Bassham, Diane C.; Bergamini, Ettore; Bi, Xiaoning; Biard-Piechaczyk, Martine; Blum, Janice S.; Bredesen, Dale E.; Brodsky, Jeffrey L.; Brumell, John H.; Brunk, Ulf T.; Bursch, Wilfried; Camougrand, Nadine; Cebollero, Eduardo; Cecconi, Francesco; Chen, Yingyu; Chin, Lih-Shen; Choi, Augustine; Chu, Charleen T.; Chung, Jongkyeong; Clarke, Peter G.H.; Clark, Robert S.B.; Clarke, Steven G.; Clavé, Corinne; Cleveland, John L.; Codogno, Patrice; Colombo, María I.; Coto-Montes, Ana; Cregg, James M.; Cuervo, Ana Maria; Debnath, Jayanta; Demarchi, Francesca; Dennis, Patrick B.; Dennis, Phillip A.; Deretic, Vojo; Devenish, Rodney J.; Di Sano, Federica; Dice, J. Fred; DiFiglia, Marian; Dinesh-Kumar, Savithramma; Distelhorst, Clark W.; Djavaheri-Mergny, Mojgan; Dorsey, Frank C.; Dröge, Wulf; Dron, Michel; Dunn, William A.; Duszenko, Michael; Eissa, N. Tony; Elazar, Zvulun; Esclatine, Audrey; Eskelinen, Eeva-Liisa; Fésüs, László; Finley, Kim D.; Fuentes, José M.; Fueyo, Juan; Fujisaki, Kozo; Galliot, Brigitte; Gao, Fen-Biao; Gewirtz, David A.; Gibson, Spencer B.; Gohla, Antje; Goldberg, Alfred L.; Gonzalez, Ramon; González-Estévez, Cristina; Gorski, Sharon; Gottlieb, Roberta A.; Häussinger, Dieter; He, You-Wen; Heidenreich, Kim; Hill, Joseph A.; Høyer-Hansen, Maria; Hu, Xun; Huang, Wei-Pang; Iwasaki, Akiko; Jäättelä, Marja; Jackson, William T.; Jiang, Xuejun; Jin, Shengkan; Johansen, Terje; Jung, Jae U.; Kadowaki, Motoni; Kang, Chanhee; Kelekar, Ameeta; Kessel, David H.; Kiel, Jan A.K.W.; Kim, Hong Pyo; Kimchi, Adi; Kinsella, Timothy J.; Kiselyov, Kirill; Kitamoto, Katsuhiko; Knecht, Erwin; Komatsu, Masaaki; Kominami, Eiki; Kondo, Seiji; Kovács, Attila L.; Kroemer, Guido; Kuan, Chia-Yi; Kumar, Rakesh; Kundu, Mondira; Landry, Jacques; Laporte, Marianne; Le, Weidong; Lei, Huan-Yao; Lenardo, Michael J.; Levine, Beth; Lieberman, Andrew; Lim, Kah-Leong; Lin, Fu-Cheng; Liou, Willisa; Liu, Leroy F.; Lopez-Berestein, Gabriel; López-Otín, Carlos; Lu, Bo; Macleod, Kay F.; Malorni, Walter; Martinet, Wim; Matsuoka, Ken; Mautner, Josef; Meijer, Alfred J.; Meléndez, Alicia; Michels, Paul; Miotto, Giovanni; Mistiaen, Wilhelm P.; Mizushima, Noboru; Mograbi, Baharia; Monastyrska, Iryna; Moore, Michael N.; Moreira, Paula I.; Moriyasu, Yuji; Motyl, Tomasz; Münz, Christian; Murphy, Leon O.; Naqvi, Naweed I.; Neufeld, Thomas P.; Nishino, Ichizo; Nixon, Ralph A.; Noda, Takeshi; Nürnberg, Bernd; Ogawa, Michinaga; Oleinick, Nancy L.; Olsen, Laura J.; Ozpolat, Bulent; Paglin, Shoshana; Palmer, Glen E.; Papassideri, Issidora; Parkes, Miles; Perlmutter, David H.; Perry, George; Piacentini, Mauro; Pinkas-Kramarski, Ronit; Prescott, Mark; Proikas-Cezanne, Tassula; Raben, Nina; Rami, Abdelhaq; Reggiori, Fulvio; Rohrer, Bärbel; Rubinsztein, David C.; Ryan, Kevin M.; Sadoshima, Junichi; Sakagami, Hiroshi; Sakai, Yasuyoshi; Sandri, Marco; Sasakawa, Chihiro; Sass, Miklós; Schneider, Claudio; Seglen, Per O.; Seleverstov, Oleksandr; Settleman, Jeffrey; Shacka, John J.; Shapiro, Irving M.; Sibirny, Andrei; Silva-Zacarin, Elaine C.M.; Simon, Hans-Uwe; Simone, Cristiano; Simonsen, Anne; Smith, Mark A.; Spanel-Borowski, Katharina; Srinivas, Vickram; Steeves, Meredith; Stenmark, Harald; Stromhaug, Per E.; Subauste, Carlos S.; Sugimoto, Seiichiro; Sulzer, David; Suzuki, Toshihiko; Swanson, Michele S.; Tabas, Ira; Takeshita, Fumihiko; Talbot, Nicholas J.; Tallóczy, Zsolt; Tanaka, Keiji; Tanaka, Kozo; Tanida, Isei; Taylor, Graham S.; Taylor, J. Paul; Terman, Alexei; Tettamanti, Gianluca; Thompson, Craig B.; Thumm, Michael; Tolkovsky, Aviva M.; Tooze, Sharon A.; Truant, Ray; Tumanovska, Lesya V.; Uchiyama, Yasuo; Ueno, Takashi; Uzcátegui, Néstor L.; van der Klei, Ida; Vaquero, Eva C.; Vellai, Tibor; Vogel, Michael W.; Wang, Hong-Gang; Webster, Paul; Wiley, John W.; Xi, Zhijun; Xiao, Gutian; Yahalom, Joachim; Yang, Jin-Ming; Yap, George; Yin, Xiao-Ming; Yoshimori, Tamotsu; Yu, Li; Yue, Zhenyu; Yuzaki, Michisuke; Zabirnyk, Olga; Zheng, Xiaoxiang; Zhu, Xiongwei; Deter, Russell L.

    2009-01-01

    Research in autophagy continues to accelerate,1 and as a result many new scientists are entering the field. Accordingly, it is important to establish a standard set of criteria for monitoring macroautophagy in different organisms. Recent reviews have described the range of assays that have been used for this purpose.2,3 There are many useful and convenient methods that can be used to monitor macroautophagy in yeast, but relatively few in other model systems, and there is much confusion regarding acceptable methods to measure macroautophagy in higher eukaryotes. A key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers of autophagosomes versus those that measure flux through the autophagy pathway; thus, a block in macroautophagy that results in autophagosome accumulation needs to be differentiated from fully functional autophagy that includes delivery to, and degradation within, lysosomes (in most higher eukaryotes) or the vacuole (in plants and fungi). Here, we present a set of guidelines for the selection and interpretation of the methods that can be used by investigators who are attempting to examine macroautophagy and related processes, as well as by reviewers who need to provide realistic and reasonable critiques of papers that investigate these processes. This set of guidelines is not meant to be a formulaic set of rules, because the appropriate assays depend in part on the question being asked and the system being used. In addition, we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to verify an autophagic response. PMID:18188003

  2. Strong intrinsic motivation

    OpenAIRE

    Dessi, Roberta; Rustichini, Aldo

    2015-01-01

    A large literature in psychology, and more recently in economics, has argued that monetary rewards can reduce intrinsic motivation. We investigate whether the negative impact persists when intrinsic motivation is strong, and test this hypothesis experimentally focusing on the motivation to undertake interesting and challenging tasks, informative about individual ability. We find that this type of task can generate strong intrinsic motivation, that is impervious to the effect of monetary incen...

  3. Bitcoin Meets Strong Consistency

    OpenAIRE

    Decker, Christian; Seidel, Jochen; Wattenhofer, Roger

    2014-01-01

    The Bitcoin system only provides eventual consistency. For everyday life, the time to confirm a Bitcoin transaction is prohibitively slow. In this paper we propose a new system, built on the Bitcoin blockchain, which enables strong consistency. Our system, PeerCensus, acts as a certification authority, manages peer identities in a peer-to-peer network, and ultimately enhances Bitcoin and similar systems with strong consistency. Our extensive analysis shows that PeerCensus is in a secure state...

  4. Strong gravity and supersymmetry

    International Nuclear Information System (INIS)

    Chamseddine, Ali H.; Salam, A.; Strathdee, J.

    1977-11-01

    A supersymmetric theory is constructed for a strong f plus a weak g graviton, together with their accompanying massive gravitinos, by gaugin the gradel 0Sp(2,2,1)x 0Sp(2,2,1) structure. The mixing term between f and g fields, which makes the strong graviton massive, can be introduced through a spontaneous symmetry-breaking mechanism implemented in this note by constructing a non-linear realization of the symmetry group

  5. An Interactive Exercise To Learn Eukaryotic Cell Structure and Organelle Function.

    Science.gov (United States)

    Klionsky, Daniel J.; Tomashek, John J.

    1999-01-01

    Describes a cooperative, interactive problem-solving exercise for studying eukaryotic cell structure and function. Highlights the dynamic aspects of movement through the cell. Contains 15 references. (WRM)

  6. [MiRNA system in unicellular eukaryotes and its evolutionary implications].

    Science.gov (United States)

    Zhang, Yan-Qiong; Wen, Jian-Fan

    2010-02-01

    microRNAs (miRNAs) in higher multicellular eukaryotes have been extensively studied in recent years. Great progresses have also been achieved for miRNAs in unicellular eukaryotes. All these studies not only enrich our knowledge about the complex expression regulation system in diverse organisms, but also have evolutionary significance for understanding the origin of this system. In this review, Authors summarize the recent advance in the studies of miRNA in unicellular eukaryotes, including that on the most primitive unicellular eukaryote--Giardia. The origin and evolution of miRNA system is also discussed.

  7. Automatic generation of gene finders for eukaryotic species

    DEFF Research Database (Denmark)

    Terkelsen, Kasper Munch; Krogh, A.

    2006-01-01

    and quality of reliable gene annotation grows. Results We present a procedure, Agene, that automatically generates a species-specific gene predictor from a set of reliable mRNA sequences and a genome. We apply a Hidden Markov model (HMM) that implements explicit length distribution modelling for all gene......Background The number of sequenced eukaryotic genomes is rapidly increasing. This means that over time it will be hard to keep supplying customised gene finders for each genome. This calls for procedures to automatically generate species-specific gene finders and to re-train them as the quantity...... structure blocks using acyclic discrete phase type distributions. The state structure of the each HMM is generated dynamically from an array of sub-models to include only gene features represented in the training set. Conclusion Acyclic discrete phase type distributions are well suited to model sequence...

  8. Nucleosome mediated crosstalk between transcription factors at eukaryotic enhancers

    International Nuclear Information System (INIS)

    Teif, Vladimir B; Rippe, Karsten

    2011-01-01

    A recent study of transcription regulation in Drosophila embryonic development revealed a complex non-monotonic dependence of gene expression on the distance between binding sites of repressor and activator proteins at the corresponding enhancer cis-regulatory modules (Fakhouri et al 2010 Mol. Syst. Biol. 6 341). The repressor efficiency was high at small separations, low around 30 bp, reached a maximum at 50–60 bp, and decreased at larger distances to the activator binding sites. Here, we propose a straightforward explanation for the distance dependence of repressor activity by considering the effect of the presence of a nucleosome. Using a method that considers partial unwrapping of nucleosomal DNA from the histone octamer core, we calculated the dependence of activator binding on the repressor–activator distance and found a quantitative agreement with the distance dependence reported for the Drosophila enhancer element. In addition, the proposed model offers explanations for other distance-dependent effects at eukaryotic enhancers. (communication)

  9. The biology of eukaryotic promoter prediction - a review

    DEFF Research Database (Denmark)

    Pedersen, Anders Gorm; Baldi, Pierre; Chauvin, Yves

    1999-01-01

    between functional promoters has been estimated to be in the range of 30-40 kilobases. Although it is conceivable that some of these predicted promoters correspond to cryptic initiation sites that are used in vivo, it is likely that most are false positives. This suggests that it is important to carefully......Computational prediction of eukaryotic promoters from the nucleotide sequence is one of the most attractive problems in sequence analysis today, but it is also a very difficult one. Thus, current methods predict in the order of one promoter per kilobase in human DNA, while the average distance...... reconsider the biological data that forms the basis of current algorithms, and we here present a review of data that may be useful in this regard. The review covers the following topics: (1) basal transcription and core promoters, (2) activated transcription and transcription factor binding sites, (3) Cp...

  10. A general strategy to construct small molecule biosensors in eukaryotes.

    Science.gov (United States)

    Feng, Justin; Jester, Benjamin W; Tinberg, Christine E; Mandell, Daniel J; Antunes, Mauricio S; Chari, Raj; Morey, Kevin J; Rios, Xavier; Medford, June I; Church, George M; Fields, Stanley; Baker, David

    2015-12-29

    Biosensors for small molecules can be used in applications that range from metabolic engineering to orthogonal control of transcription. Here, we produce biosensors based on a ligand-binding domain (LBD) by using a method that, in principle, can be applied to any target molecule. The LBD is fused to either a fluorescent protein or a transcriptional activator and is destabilized by mutation such that the fusion accumulates only in cells containing the target ligand. We illustrate the power of this method by developing biosensors for digoxin and progesterone. Addition of ligand to yeast, mammalian, or plant cells expressing a biosensor activates transcription with a dynamic range of up to ~100-fold. We use the biosensors to improve the biotransformation of pregnenolone to progesterone in yeast and to regulate CRISPR activity in mammalian cells. This work provides a general methodology to develop biosensors for a broad range of molecules in eukaryotes.

  11. Origin and evolution of SINEs in eukaryotic genomes.

    Science.gov (United States)

    Kramerov, D A; Vassetzky, N S

    2011-12-01

    Short interspersed elements (SINEs) are one of the two most prolific mobile genomic elements in most of the higher eukaryotes. Although their biology is still not thoroughly understood, unusual life cycle of these simple elements amplified as genomic parasites makes their evolution unique in many ways. In contrast to most genetic elements including other transposons, SINEs emerged de novo many times in evolution from available molecules (for example, tRNA). The involvement of reverse transcription in their amplification cycle, huge number of genomic copies and modular structure allow variation mechanisms in SINEs uncommon or rare in other genetic elements (module exchange between SINE families, dimerization, and so on.). Overall, SINE evolution includes their emergence, progressive optimization and counteraction to the cell's defense against mobile genetic elements.

  12. Cytoplasmic Flow Enhances Organelle Dispersion in Eukaryotic Cells

    Science.gov (United States)

    Koslover, Elena; Mogre, Saurabh; Chan, Caleb; Theriot, Julie

    The cytoplasm of a living cell is an active environment through which intracellular components move and mix. We explore, using theoretical modeling coupled with microrheological measurements, the efficiency of particle dispersion via different modes of transport within this active environment. In particular, we focus on the role of cytoplasmic flow over different scales in contributing to organelle transport within two different cell types. In motile neutrophil cells, we show that bulk fluid flow associated with rapid cell deformation enhances particle transport to and from the cell periphery. In narrow fungal hyphae, localized flows due to hydrodynamic entrainment are shown to contribute to optimally efficient organelle dispersion. Our results highlight the importance of non-traditional modes of transport associated with flow of the cytoplasmic fluid in the distribution of organelles throughout eukaryotic cells.

  13. Ancient photosynthetic eukaryote biofilms in an Atacama Desert coastal cave

    Science.gov (United States)

    Azua-Bustos, A.; Gonzalez-Silva, C.; Mancilla, R.A.; Salas, L.; Palma, R.E.; Wynne, J.J.; McKay, C.P.; Vicuna, R.

    2009-01-01

    Caves offer a stable and protected environment from harsh and changing outside prevailing conditions. Hence, they represent an interesting habitat for studying life in extreme environments. Here, we report the presence of a member of the ancient eukaryote red algae Cyanidium group in a coastal cave of the hyperarid Atacama Desert. This microorganism was found to form a seemingly monospecific biofilm growing under extremely low photon flux levels. Our work suggests that this species, Cyanidium sp. Atacama, is a new member of a recently proposed novel monophyletic lineage of mesophilic "cave" Cyanidium sp., distinct from the remaining three other lineages which are all thermo-acidophilic. The cave described in this work may represent an evolutionary island for life in the midst of the Atacama Desert. ?? Springer Science + Business Media, LLC 2009.

  14. Short RNA guides cleavage by eukaryotic RNase III.

    Directory of Open Access Journals (Sweden)

    Bruno Lamontagne

    Full Text Available In eukaryotes, short RNAs guide a variety of enzymatic activities that range from RNA editing to translation repression. It is hypothesized that pre-existing proteins evolved to bind and use guide RNA during evolution. However, the capacity of modern proteins to adopt new RNA guides has never been demonstrated. Here we show that Rnt1p, the yeast orthologue of the bacterial dsRNA-specific RNase III, can bind short RNA transcripts and use them as guides for sequence-specific cleavage. Target cleavage occurred at a constant distance from the Rnt1p binding site, leaving the guide RNA intact for subsequent cleavage. Our results indicate that RNase III may trigger sequence-specific RNA degradation independent of the RNAi machinery, and they open the road for a new generation of precise RNA silencing tools that do not trigger a dsRNA-mediated immune response.

  15. Synthesis of eukaryotic lipid biomarkers in the bacterial domain

    Science.gov (United States)

    Welander, P. V.; Banta, A. B.; Lee, A. K.; Wei, J. H.

    2017-12-01

    Lipid biomarkers are organic molecules preserved in sediments and sedimentary rocks that can function as geological proxies for certain microbial taxa or for specific environmental conditions. These molecular fossils provide a link between organisms and their environments in both modern and ancient settings and have afforded significant insight into ancient climatic events, mass extinctions, and various evolutionary transitions throughout Earth's history. However, the proper interpretation of lipid biomarkers is dependent on a broad understanding of their diagenetic precursors in modern systems. This includes understanding the taphonomic transformations that these molecules undergo, their biosynthetic pathways, and the ecological conditions that affect their cellular production. In this study, we focus on one group of lipid biomarkers - the sterols. These are polycyclic isoprenoidal lipids that have a high preservation potential and play a critical role in the physiology of most eukaryotes. However, the synthesis and function of these lipids in the bacterial domain has not been fully explored. Here we utilize a combination of bioinformatics, microbial genetics, and biochemistry to demonstrate that bacterial sterol producers are more prevalent in environmental metagenomic samples than in the genomic databases of cultured organisms and to identify novel proteins required to synthesize and modify sterols in bacteria. These proteins represent a distinct pathway for sterol synthesis exclusive to bacteria and indicate that sterol synthesis in bacteria may have evolved independently of eukaryotic sterol biosynthesis. Taken together, these results demonstrate how studies in extant bacteria can provide insight into the biological sources and the biosynthetic pathways of specific lipid biomarkers and in turn may allow for more robust interpretation of biomarker signatures.

  16. Interactions between subunits of Saccharomyces cerevisiae RNase MRP support a conserved eukaryotic RNase P/MRP architecture.

    Science.gov (United States)

    Aspinall, Tanya V; Gordon, James M B; Bennett, Hayley J; Karahalios, Panagiotis; Bukowski, John-Paul; Walker, Scott C; Engelke, David R; Avis, Johanna M

    2007-01-01

    Ribonuclease MRP is an endonuclease, related to RNase P, which functions in eukaryotic pre-rRNA processing. In Saccharomyces cerevisiae, RNase MRP comprises an RNA subunit and ten proteins. To improve our understanding of subunit roles and enzyme architecture, we have examined protein-protein and protein-RNA interactions in vitro, complementing existing yeast two-hybrid data. In total, 31 direct protein-protein interactions were identified, each protein interacting with at least three others. Furthermore, seven proteins self-interact, four strongly, pointing to subunit multiplicity in the holoenzyme. Six protein subunits interact directly with MRP RNA and four with pre-rRNA. A comparative analysis with existing data for the yeast and human RNase P/MRP systems enables confident identification of Pop1p, Pop4p and Rpp1p as subunits that lie at the enzyme core, with probable addition of Pop5p and Pop3p. Rmp1p is confirmed as an integral subunit, presumably associating preferentially with RNase MRP, rather than RNase P, via interactions with Snm1p and MRP RNA. Snm1p and Rmp1p may act together to assist enzyme specificity, though roles in substrate binding are also indicated for Pop4p and Pop6p. The results provide further evidence of a conserved eukaryotic RNase P/MRP architecture and provide a strong basis for studies of enzyme assembly and subunit function.

  17. RNase MRP and the RNA processing cascade in the eukaryotic ancestor.

    Science.gov (United States)

    Woodhams, Michael D; Stadler, Peter F; Penny, David; Collins, Lesley J

    2007-02-08

    Within eukaryotes there is a complex cascade of RNA-based macromolecules that process other RNA molecules, especially mRNA, tRNA and rRNA. An example is RNase MRP processing ribosomal RNA (rRNA) in ribosome biogenesis. One hypothesis is that this complexity was present early in eukaryotic evolution; an alternative is that an initial simpler network later gained complexity by gene duplication in lineages that led to animals, fungi and plants. Recently there has been a rapid increase in support for the complexity-early theory because the vast majority of these RNA-processing reactions are found throughout eukaryotes, and thus were likely to be present in the last common ancestor of living eukaryotes, herein called the Eukaryotic Ancestor. We present an overview of the RNA processing cascade in the Eukaryotic Ancestor and investigate in particular, RNase MRP which was previously thought to have evolved later in eukaryotes due to its apparent limited distribution in fungi and animals and plants. Recent publications, as well as our own genomic searches, find previously unknown RNase MRP RNAs, indicating that RNase MRP has a wide distribution in eukaryotes. Combining secondary structure and promoter region analysis of RNAs for RNase MRP, along with analysis of the target substrate (rRNA), allows us to discuss this distribution in the light of eukaryotic evolution. We conclude that RNase MRP can now be placed in the RNA-processing cascade of the Eukaryotic Ancestor, highlighting the complexity of RNA-processing in early eukaryotes. Promoter analyses of MRP-RNA suggest that regulation of the critical processes of rRNA cleavage can vary, showing that even these key cellular processes (for which we expect high conservation) show some species-specific variability. We present our consensus MRP-RNA secondary structure as a useful model for further searches.

  18. Strongly interacting Fermi gases

    Directory of Open Access Journals (Sweden)

    Bakr W.

    2013-08-01

    Full Text Available Strongly interacting gases of ultracold fermions have become an amazingly rich test-bed for many-body theories of fermionic matter. Here we present our recent experiments on these systems. Firstly, we discuss high-precision measurements on the thermodynamics of a strongly interacting Fermi gas across the superfluid transition. The onset of superfluidity is directly observed in the compressibility, the chemical potential, the entropy, and the heat capacity. Our measurements provide benchmarks for current many-body theories on strongly interacting fermions. Secondly, we have studied the evolution of fermion pairing from three to two dimensions in these gases, relating to the physics of layered superconductors. In the presence of p-wave interactions, Fermi gases are predicted to display toplogical superfluidity carrying Majorana edge states. Two possible avenues in this direction are discussed, our creation and direct observation of spin-orbit coupling in Fermi gases and the creation of fermionic molecules of 23Na 40K that will feature strong dipolar interactions in their absolute ground state.

  19. A strong comeback

    International Nuclear Information System (INIS)

    Marier, D.

    1992-01-01

    This article presents the results of a financial rankings survey which show a strong economic activity in the independent energy industry. The topics of the article include advisor turnover, overseas banks, and the increase in public offerings. The article identifies the top project finance investors for new projects and restructurings and rankings for lenders

  20. What can we infer about the origin of sex in early eukaryotes?

    NARCIS (Netherlands)

    Speijer, Dave

    2016-01-01

    Current analysis shows that the last eukaryotic common ancestor (LECA) was capable of full meiotic sex. The original eukaryotic life cycle can probably be described as clonal, interrupted by episodic sex triggered by external or internal stressors. The cycle could have started in a highly flexible

  1. Sex is a ubiquitous, ancient, and inherent attribute of eukaryotic life

    NARCIS (Netherlands)

    Speijer, Dave; Lukeš, Julius; Eliáš, Marek

    2015-01-01

    Sexual reproduction and clonality in eukaryotes are mostly seen as exclusive, the latter being rather exceptional. This view might be biased by focusing almost exclusively on metazoans. We analyze and discuss reproduction in the context of extant eukaryotic diversity, paying special attention to

  2. Genome-wide Purification of Extrachromosomal Circular DNA from Eukaryotic Cells

    DEFF Research Database (Denmark)

    Møller, Henrik D.; Bojsen, Rasmus Kenneth; Tachibana, Chris

    2016-01-01

    Extrachromosomal circular DNAs (eccDNAs) are common genetic elements in Saccharomyces cerevisiae and are reported in other eukaryotes as well. EccDNAs contribute to genetic variation among somatic cells in multicellular organisms and to evolution of unicellular eukaryotes. Sensitive methods for d...

  3. Eelgrass Leaf Surface Microbiomes Are Locally Variable and Highly Correlated with Epibiotic Eukaryotes

    Directory of Open Access Journals (Sweden)

    Mia M. Bengtsson

    2017-07-01

    Full Text Available Eelgrass (Zostera marina is a marine foundation species essential for coastal ecosystem services around the northern hemisphere. Like all macroscopic organisms, it possesses a microbiome (here defined as an associated prokaryotic community which may play critical roles in modulating the interaction of eelgrass with its environment. For example, its leaf surface microbiome could inhibit or attract eukaryotic epibionts which may overgrow the eelgrass leading to reduced primary productivity and subsequent eelgrass meadow decline. We used amplicon sequencing of the 16S and 18S rRNA genes of prokaryotes and eukaryotes to assess the leaf surface microbiome (prokaryotes as well as eukaryotic epibionts in- and outside lagoons on the German Baltic Sea coast. Prokaryote microbiomes varied substantially both between sites inside lagoons and between open coastal and lagoon sites. Water depth, leaf area and biofilm chlorophyll a concentration explained a large amount of variation in both prokaryotic and eukaryotic community composition. The prokaryotic microbiome and eukaryotic epibiont communities were highly correlated, and network analysis revealed disproportionate co-occurrence between a limited number of eukaryotic taxa and several bacterial taxa. This suggests that eelgrass leaf surfaces are home to a mosaic of microbiomes of several epibiotic eukaryotes, in addition to the microbiome of the eelgrass itself. Our findings thereby underline that eukaryotic diversity should be taken into account in order to explain prokaryotic microbiome assembly and dynamics in aquatic environments.

  4. Use of prokaryotic transcriptional activators as metabolite biosensors in eukaryotic cells

    DEFF Research Database (Denmark)

    2018-01-01

    The present invention relates to the use of transcriptional activators from prokaryotic organisms for use in eukaryotic cells, such as yeast as sensors of intracellular and extracellular accumulation of a ligand or metabolite specifically activating this transcriptional activator in a eukaryot...

  5. Genome-wide Purification of Extrachromosomal Circular DNA from Eukaryotic Cells

    DEFF Research Database (Denmark)

    Møller, Henrik D.; Bojsen, Rasmus Kenneth; Tachibana, Chris

    2016-01-01

    Extrachromosomal circular DNAs (eccDNAs) are common genetic elements in Saccharomyces cerevisiae and are reported in other eukaryotes as well. EccDNAs contribute to genetic variation among somatic cells in multicellular organisms and to evolution of unicellular eukaryotes. Sensitive methods...

  6. On the Archaeal Origins of Eukaryotes and the Challenges of Inferring Phenotype from Genotype.

    Science.gov (United States)

    Dey, Gautam; Thattai, Mukund; Baum, Buzz

    2016-07-01

    If eukaryotes arose through a merger between archaea and bacteria, what did the first true eukaryotic cell look like? A major step toward an answer came with the discovery of Lokiarchaeum, an archaeon whose genome encodes small GTPases related to those used by eukaryotes to regulate membrane traffic. Although 'Loki' cells have yet to be seen, their existence has prompted the suggestion that the archaeal ancestor of eukaryotes engulfed the future mitochondrion by phagocytosis. We propose instead that the archaeal ancestor was a relatively simple cell, and that eukaryotic cellular organization arose as the result of a gradual transfer of bacterial genes and membranes driven by an ever-closer symbiotic partnership between a bacterium and an archaeon. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

  7. Strong Electroweak Symmetry Breaking

    CERN Document Server

    Grinstein, Benjamin

    2011-01-01

    Models of spontaneous breaking of electroweak symmetry by a strong interaction do not have fine tuning/hierarchy problem. They are conceptually elegant and use the only mechanism of spontaneous breaking of a gauge symmetry that is known to occur in nature. The simplest model, minimal technicolor with extended technicolor interactions, is appealing because one can calculate by scaling up from QCD. But it is ruled out on many counts: inappropriately low quark and lepton masses (or excessive FCNC), bad electroweak data fits, light scalar and vector states, etc. However, nature may not choose the minimal model and then we are stuck: except possibly through lattice simulations, we are unable to compute and test the models. In the LHC era it therefore makes sense to abandon specific models (of strong EW breaking) and concentrate on generic features that may indicate discovery. The Technicolor Straw Man is not a model but a parametrized search strategy inspired by a remarkable generic feature of walking technicolor,...

  8. Phylogenetic analysis of ferlin genes reveals ancient eukaryotic origins

    Directory of Open Access Journals (Sweden)

    Lek Monkol

    2010-07-01

    Full Text Available Abstract Background The ferlin gene family possesses a rare and identifying feature consisting of multiple tandem C2 domains and a C-terminal transmembrane domain. Much currently remains unknown about the fundamental function of this gene family, however, mutations in its two most well-characterised members, dysferlin and otoferlin, have been implicated in human disease. The availability of genome sequences from a wide range of species makes it possible to explore the evolution of the ferlin family, providing contextual insight into characteristic features that define the ferlin gene family in its present form in humans. Results Ferlin genes were detected from all species of representative phyla, with two ferlin subgroups partitioned within the ferlin phylogenetic tree based on the presence or absence of a DysF domain. Invertebrates generally possessed two ferlin genes (one with DysF and one without, with six ferlin genes in most vertebrates (three DysF, three non-DysF. Expansion of the ferlin gene family is evident between the divergence of lamprey (jawless vertebrates and shark (cartilaginous fish. Common to almost all ferlins is an N-terminal C2-FerI-C2 sandwich, a FerB motif, and two C-terminal C2 domains (C2E and C2F adjacent to the transmembrane domain. Preservation of these structural elements throughout eukaryotic evolution suggests a fundamental role of these motifs for ferlin function. In contrast, DysF, C2DE, and FerA are optional, giving rise to subtle differences in domain topologies of ferlin genes. Despite conservation of multiple C2 domains in all ferlins, the C-terminal C2 domains (C2E and C2F displayed higher sequence conservation and greater conservation of putative calcium binding residues across paralogs and orthologs. Interestingly, the two most studied non-mammalian ferlins (Fer-1 and Misfire in model organisms C. elegans and D. melanogaster, present as outgroups in the phylogenetic analysis, with results suggesting

  9. Plasmons in strong superconductors

    International Nuclear Information System (INIS)

    Baldo, M.; Ducoin, C.

    2011-01-01

    We present a study of the possible plasmon excitations that can occur in systems where strong superconductivity is present. In these systems the plasmon energy is comparable to or smaller than the pairing gap. As a prototype of these systems we consider the proton component of Neutron Star matter just below the crust when electron screening is not taken into account. For the realistic case we consider in detail the different aspects of the elementary excitations when the proton, electron components are considered within the Random-Phase Approximation generalized to the superfluid case, while the influence of the neutron component is considered only at qualitative level. Electron screening plays a major role in modifying the proton spectrum and spectral function. At the same time the electron plasmon is strongly modified and damped by the indirect coupling with the superfluid proton component, even at moderately low values of the gap. The excitation spectrum shows the interplay of the different components and their relevance for each excitation modes. The results are relevant for neutrino physics and thermodynamical processes in neutron stars. If electron screening is neglected, the spectral properties of the proton component show some resemblance with the physical situation in high-T c superconductors, and we briefly discuss similarities and differences in this connection. In a general prospect, the results of the study emphasize the role of Coulomb interaction in strong superconductors.

  10. The largest subunit of RNA polymerase II from the Glaucocystophyta: functional constraint and short-branch exclusion in deep eukaryotic phylogeny

    Directory of Open Access Journals (Sweden)

    Stiller John W

    2005-12-01

    Full Text Available Abstract Background Evolutionary analyses of the largest subunit of RNA polymerase II (RPB1 have yielded important and at times provocative results. One particularly troublesome outcome is the consistent inference of independent origins of red algae and green plants, at odds with the more widely accepted view of a monophyletic Plantae comprising all eukaryotes with primary plastids. If the hypothesis of a broader kingdom Plantae is correct, then RPB1 trees likely reflect a persistent phylogenetic artifact. To gain a better understanding of RNAP II evolution, and the presumed artifact relating to green plants and red algae, we isolated and analyzed RPB1 from representatives of Glaucocystophyta, the third eukaryotic group with primary plastids. Results Phylogenetic analyses incorporating glaucocystophytes do not recover a monophyletic Plantae; rather they result in additional conflicts with the most widely held views on eukaryotic relationships. In particular, glaucocystophytes are recovered as sister to several amoebozoans with strong support. A detailed investigation shows that this clade can be explained by what we call "short-branch exclusion," a phylogenetic artifact integrally associated with "long-branch attraction." Other systematic discrepancies observed in RPB1 trees can be explained as phylogenetic artifacts; however, these apparent artifacts also appear in regions of the tree that support widely held views of eukaryotic evolution. In fact, most of the RPB1 tree is consistent with artifacts of rate variation among sequences and co-variation due to functional constraints related to C-terminal domain based RNAP II transcription. Conclusion Our results reveal how subtle and easily overlooked biases can dominate the overall results of molecular phylogenetic analyses of ancient eukaryotic relationships. Sources of potential phylogenetic artifact should be investigated routinely, not just when obvious "long-branch attraction" is encountered.

  11. Relevance of intracellular polarity to accuracy of eukaryotic chemotaxis

    International Nuclear Information System (INIS)

    Hiraiwa, Tetsuya; Nishikawa, Masatoshi; Shibata, Tatsuo; Nagamatsu, Akihiro; Akuzawa, Naohiro

    2014-01-01

    Eukaryotic chemotaxis is usually mediated by intracellular signals that tend to localize at the front or back of the cell. Such intracellular polarities frequently require no extracellular guidance cues, indicating that spontaneous polarization occurs in the signal network. Spontaneous polarization activity is considered relevant to the persistent motions in random cell migrations and chemotaxis. In this study, we propose a theoretical model that connects spontaneous intracellular polarity and motile ability in a chemoattractant solution. We demonstrate that the intracellular polarity can enhance the accuracy of chemotaxis. Chemotactic accuracy should also depend on chemoattractant concentration through the concentration-dependent correlation time in the polarity direction. Both the polarity correlation time and the chemotactic accuracy depend on the degree of responsiveness to the chemical gradient. We show that optimally accurate chemotaxis occurs at an intermediate responsiveness of intracellular polarity. Experimentally, we find that the persistence time of randomly migrating Dictyostelium cells depends on the chemoattractant concentration, as predicted by our theory. At the optimum responsiveness, this ameboid cell can enhance its chemotactic accuracy tenfold. (paper)

  12. Graph theoretic analysis of protein interaction networks of eukaryotes

    Science.gov (United States)

    Goh, K.-I.; Kahng, B.; Kim, D.

    2005-11-01

    Owing to the recent progress in high-throughput experimental techniques, the datasets of large-scale protein interactions of prototypical multicellular species, the nematode worm Caenorhabditis elegans and the fruit fly Drosophila melanogaster, have been assayed. The datasets are obtained mainly by using the yeast hybrid method, which contains false-positive and false-negative simultaneously. Accordingly, while it is desirable to test such datasets through further wet experiments, here we invoke recent developed network theory to test such high-throughput datasets in a simple way. Based on the fact that the key biological processes indispensable to maintaining life are conserved across eukaryotic species, and the comparison of structural properties of the protein interaction networks (PINs) of the two species with those of the yeast PIN, we find that while the worm and yeast PIN datasets exhibit similar structural properties, the current fly dataset, though most comprehensively screened ever, does not reflect generic structural properties correctly as it is. The modularity is suppressed and the connectivity correlation is lacking. Addition of interologs to the current fly dataset increases the modularity and enhances the occurrence of triangular motifs as well. The connectivity correlation function of the fly, however, remains distinct under such interolog additions, for which we present a possible scenario through an in silico modeling.

  13. NMR comparison of prokaryotic and eukaryotic cytochromes c

    International Nuclear Information System (INIS)

    Chau, Meihing; Cai, Meng Li; Timkovich, R.

    1990-01-01

    1 H NMR spectroscopy has been used to examine ferrocytochrome c-551 from Pseudomonas aeruginosa (ATCC 19429) over the pH range 3.5-10.6 and the temperature range 4-60 degree C. Resonance assignments are proposed for main-chain and side-chain protons. Comparison of results for cytochrome c-551 to recently assigned spectra for horse cytochrome c and mutants of yeast iso-1 cytochrome reveals some unique resonances with unusual chemical shifts in all cytochromes that may serve as markers for the heme region. Results for cytochrome c-551 indicate that in the smaller prokaryotic cytochrome, all benzoid side chains are rapidly flipping on the NMR time scale. In contrast, in eukaryotic cytochromes there are some rings flipping slowly on the NMR time scale. The ferrocytochrome c-551 undergoes a transition linked to pH with a pK around 7. The pH behavior of assigned resonances provides evidence that the site of protonation is the inner or buried 17-propionic acid heme substituent (IUPAC-IUB porphyrin nomenclature). Conformational heterogeneity has been observed for segments near the inner heme propionate substituent

  14. Structural similarities between prokaryotic and eukaryotic 5S ribosomal RNAs

    International Nuclear Information System (INIS)

    Welfle, H.; Boehm, S.; Damaschun, G.; Fabian, H.; Gast, K.; Misselwitz, R.; Mueller, J.J.; Zirwer, D.; Filimonov, V.V.; Venyaminov, S.Yu.; Zalkova, T.N.

    1986-01-01

    5S RNAs from rat liver and E. coli have been studied by diffuse X-ray and dynamic light scattering and by infrared and Raman spectroscopy. Identical structures at a resolution of 1 nm can be deduced from the comparison of the experimental X-ray scattering curves and electron distance distribution functions and from the agreement of the shape parameters. A flat shape model with a compact central region and two protruding arms was derived. Double helical stems are eleven-fold helices with a mean base pair distance of 0.28 nm. The number of base pairs (26 GC, 9 AU for E. coli; 27 GC, 9 AU for rat liver) and the degree of base stacking are the same within the experimental error. A very high regularity in the ribophosphate backbone is indicated for both 5S RNAs. The observed structural similarity and the consensus secondary structure pattern derived from comparative sequence analyses suggest the conclusion that prokaryotic and eukaryotic 5S RNAs are in general very similar with respect to their fundamental structural features. (author)

  15. Biosurfactant gene clusters in eukaryotes: regulation and biotechnological potential.

    Science.gov (United States)

    Roelants, Sophie L K W; De Maeseneire, Sofie L; Ciesielska, Katarzyna; Van Bogaert, Inge N A; Soetaert, Wim

    2014-04-01

    Biosurfactants (BSs) are a class of secondary metabolites representing a wide variety of structures that can be produced from renewable feedstock by a wide variety of micro-organisms. They have (potential) applications in the medical world, personal care sector, mining processes, food industry, cosmetics, crop protection, pharmaceuticals, bio-remediation, household detergents, paper and pulp industry, textiles, paint industries, etc. Especially glycolipid BSs like sophorolipids (SLs), rhamnolipids (RLs), mannosylerythritol lipids (MELs) and cellobioselipids (CBLs) have been described to provide significant opportunities to (partially) replace chemical surfactants. The major two factors currently limiting the penetration of BSs into the market are firstly the limited structural variety and secondly the rather high production price linked with the productivity. One of the keys to resolve the above mentioned bottlenecks can be found in the genetic engineering of natural producers. This could not only result in more efficient (economical) recombinant producers, but also in a diversification of the spectrum of available BSs as such resolving both limiting factors at once. Unraveling the genetics behind the biosynthesis of these interesting biological compounds is indispensable for the tinkering, fine tuning and rearrangement of these biological pathways with the aim of obtaining higher yields and a more extensive structural variety. Therefore, this review focuses on recent developments in the investigation of the biosynthesis, genetics and regulation of some important members of the family of the eukaryotic glycolipid BSs (MELs, CBLs and SLs). Moreover, recent biotechnological achievements and the industrial potential of engineered strains are discussed.

  16. MCM Paradox: Abundance of Eukaryotic Replicative Helicases and Genomic Integrity.

    Science.gov (United States)

    Das, Mitali; Singh, Sunita; Pradhan, Satyajit; Narayan, Gopeshwar

    2014-01-01

    As a crucial component of DNA replication licensing system, minichromosome maintenance (MCM) 2-7 complex acts as the eukaryotic DNA replicative helicase. The six related MCM proteins form a heterohexamer and bind with ORC, CDC6, and Cdt1 to form the prereplication complex. Although the MCMs are well known as replicative helicases, their overabundance and distribution patterns on chromatin present a paradox called the "MCM paradox." Several approaches had been taken to solve the MCM paradox and describe the purpose of excess MCMs distributed beyond the replication origins. Alternative functions of these MCMs rather than a helicase had also been proposed. This review focuses on several models and concepts generated to solve the MCM paradox coinciding with their helicase function and provides insight into the concept that excess MCMs are meant for licensing dormant origins as a backup during replication stress. Finally, we extend our view towards the effect of alteration of MCM level. Though an excess MCM constituent is needed for normal cells to withstand stress, there must be a delineation of the threshold level in normal and malignant cells. This review also outlooks the future prospects to better understand the MCM biology.

  17. The prokaryote-eukaryote dichotomy: meanings and mythology.

    Science.gov (United States)

    Sapp, Jan

    2005-06-01

    Drawing on documents both published and archival, this paper explains how the prokaryote-eukaryote dichotomy of the 1960s was constructed, the purposes it served, and what it implied in terms of classification and phylogeny. In doing so, I first show how the concept was attributed to Edouard Chatton and the context in which he introduced the terms. Following, I examine the context in which the terms were reintroduced into biology in 1962 by Roger Stanier and C. B. van Niel. I study the discourse over the subsequent decade to understand how the organizational dichotomy took on the form of a natural classification as the kingdom Monera or superkingdom Procaryotae. Stanier and van Niel admitted that, in regard to constructing a natural classification of bacteria, structural characteristics were no more useful than physiological properties. They repeatedly denied that bacterial phylogenetics was possible. I thus examine the great historical irony that the "prokaryote," in both its organizational and phylogenetic senses, was defined (negatively) on the basis of structure. Finally, we see how phylogenetic research based on 16S rRNA led by Carl Woese and his collaborators confronted the prokaryote concept while moving microbiology to the center of evolutionary biology.

  18. The current state of eukaryotic DNA base damage and repair.

    Science.gov (United States)

    Bauer, Nicholas C; Corbett, Anita H; Doetsch, Paul W

    2015-12-02

    DNA damage is a natural hazard of life. The most common DNA lesions are base, sugar, and single-strand break damage resulting from oxidation, alkylation, deamination, and spontaneous hydrolysis. If left unrepaired, such lesions can become fixed in the genome as permanent mutations. Thus, evolution has led to the creation of several highly conserved, partially redundant pathways to repair or mitigate the effects of DNA base damage. The biochemical mechanisms of these pathways have been well characterized and the impact of this work was recently highlighted by the selection of Tomas Lindahl, Aziz Sancar and Paul Modrich as the recipients of the 2015 Nobel Prize in Chemistry for their seminal work in defining DNA repair pathways. However, how these repair pathways are regulated and interconnected is still being elucidated. This review focuses on the classical base excision repair and strand incision pathways in eukaryotes, considering both Saccharomyces cerevisiae and humans, and extends to some important questions and challenges facing the field of DNA base damage repair. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

  19. Susceptibilities to DNA Structural Transitions within Eukaryotic Genomes

    Science.gov (United States)

    Zhabinskaya, Dina; Benham, Craig; Madden, Sally

    2012-02-01

    We analyze the competitive transitions to alternate secondary DNA structures in a negatively supercoiled DNA molecule of kilobase length and specified base sequence. We use statistical mechanics to calculate the competition among all regions within the sequence that are susceptible to transitions to alternate structures. We use an approximate numerical method since the calculation of an exact partition function is numerically cumbersome for DNA molecules of lengths longer than hundreds of base pairs. This method yields accurate results in reasonable computational times. We implement algorithms that calculate the competition between transitions to denatured states and to Z-form DNA. We analyze these transitions near the transcription start sites (TSS) of a set of eukaryotic genes. We find an enhancement of Z-forming regions upstream of the TSS and a depletion of denatured regions around the start sites. We confirm that these finding are statistically significant by comparing our results to a set of randomized genes with preserved base composition at each position relative to the gene start sites. When we study the correlation of these transitions in orthologous mouse and human genes we find a clear evolutionary conservation of both types of transitions around the TSS.

  20. Isoprenoid biosynthesis in eukaryotic phototrophs: A spotlight on algae

    Energy Technology Data Exchange (ETDEWEB)

    Lohr M.; Schwender J.; Polle, J. E. W.

    2012-04-01

    Isoprenoids are one of the largest groups of natural compounds and have a variety of important functions in the primary metabolism of land plants and algae. In recent years, our understanding of the numerous facets of isoprenoid metabolism in land plants has been rapidly increasing, while knowledge on the metabolic network of isoprenoids in algae still lags behind. Here, current views on the biochemistry and genetics of the core isoprenoid metabolism in land plants and in the major algal phyla are compared and some of the most pressing open questions are highlighted. Based on the different evolutionary histories of the various groups of eukaryotic phototrophs, we discuss the distribution and regulation of the mevalonate (MVA) and the methylerythritol phosphate (MEP) pathways in land plants and algae and the potential consequences of the loss of the MVA pathway in groups such as the green algae. For the prenyltransferases, serving as gatekeepers to the various branches of terpenoid biosynthesis in land plants and algae, we explore the minimal inventory necessary for the formation of primary isoprenoids and present a preliminary analysis of their occurrence and phylogeny in algae with primary and secondary plastids. The review concludes with some perspectives on genetic engineering of the isoprenoid metabolism in algae.

  1. The Superoxide Reductase from the Early Diverging Eukaryote Giardia Intestinalis

    International Nuclear Information System (INIS)

    Cabelli, D.E.; Testa, F.; Mastronicola, D.; Bordi, E.; Pucillo, L.P.; Sarti, P.; Saraiva, L.M.; Giuffre, A.; Teixeira, M.

    2011-01-01

    Unlike superoxide dismutases (SODs), superoxidereductases (SORs) eliminate superoxide anion (O 2 # sm b ullet# - ) not through its dismutation, but via reduction to hydrogen peroxide (H 2 O 2 ) in the presence of an electron donor. The microaerobic protist Giardia intestinalis, responsible for a common intestinal disease in humans, though lacking SOD and other canonical reactive oxygen species-detoxifying systems, is among the very few eukaryotes encoding a SOR yet identified. In this study, the recombinant SOR from Giardia (SOR Gi ) was purified and characterized by pulse radiolysis and stopped-flow spectrophotometry. The protein, isolated in the reduced state, after oxidation by superoxide or hexachloroiridate(IV), yields a resting species (T final ) with Fe 3+ ligated to glutamate or hydroxide depending on pH (apparent pK a = 8.7). Although showing negligible SOD activity, reduced SOR Gi reacts with O 2 # sm b ullet# - with a pH-independent second-order rate constant k 1 = 1.0 x 10 9 M -1 s -1 and yields the ferric-(hydro)peroxo intermediate T 1 ; this in turn rapidly decays to the T final state with pH-dependent rates, without populating other detectable intermediates. Immunoblotting assays show that SOR Gi is expressed in the disease-causing trophozoite of Giardia. We propose that the superoxide-scavenging activity of SOR in Giardia may promote the survival of this air-sensitive parasite in the fairly aerobic proximal human small intestine during infection.

  2. Classification and Lineage Tracing of SH2 Domains Throughout Eukaryotes.

    Science.gov (United States)

    Liu, Bernard A

    2017-01-01

    Today there exists a rapidly expanding number of sequenced genomes. Cataloging protein interaction domains such as the Src Homology 2 (SH2) domain across these various genomes can be accomplished with ease due to existing algorithms and predictions models. An evolutionary analysis of SH2 domains provides a step towards understanding how SH2 proteins integrated with existing signaling networks to position phosphotyrosine signaling as a crucial driver of robust cellular communication networks in metazoans. However organizing and tracing SH2 domain across organisms and understanding their evolutionary trajectory remains a challenge. This chapter describes several methodologies towards analyzing the evolutionary trajectory of SH2 domains including a global SH2 domain classification system, which facilitates annotation of new SH2 sequences essential for tracing the lineage of SH2 domains throughout eukaryote evolution. This classification utilizes a combination of sequence homology, protein domain architecture and the boundary positions between introns and exons within the SH2 domain or genes encoding these domains. Discrete SH2 families can then be traced across various genomes to provide insight into its origins. Furthermore, additional methods for examining potential mechanisms for divergence of SH2 domains from structural changes to alterations in the protein domain content and genome duplication will be discussed. Therefore a better understanding of SH2 domain evolution may enhance our insight into the emergence of phosphotyrosine signaling and the expansion of protein interaction domains.

  3. Strongly intensive quantities

    International Nuclear Information System (INIS)

    Gorenstein, M. I.; Gazdzicki, M.

    2011-01-01

    Analysis of fluctuations of hadron production properties in collisions of relativistic particles profits from use of measurable intensive quantities which are independent of system size variations. The first family of such quantities was proposed in 1992; another is introduced in this paper. Furthermore we present a proof of independence of volume fluctuations for quantities from both families within the framework of the grand canonical ensemble. These quantities are referred to as strongly intensive ones. Influence of conservation laws and resonance decays is also discussed.

  4. Strong-coupling approximations

    International Nuclear Information System (INIS)

    Abbott, R.B.

    1984-03-01

    Standard path-integral techniques such as instanton calculations give good answers for weak-coupling problems, but become unreliable for strong-coupling. Here we consider a method of replacing the original potential by a suitably chosen harmonic oscillator potential. Physically this is motivated by the fact that potential barriers below the level of the ground-state energy of a quantum-mechanical system have little effect. Numerically, results are good, both for quantum-mechanical problems and for massive phi 4 field theory in 1 + 1 dimensions. 9 references, 6 figures

  5. Strongly disordered superconductors

    International Nuclear Information System (INIS)

    Muttalib, K.A.

    1982-01-01

    We examine some universal effects of strong non-magnetic disorder on the electron-phonon and electron-electron interactions in a superconductor. In particular we explicitly take into account the effect of slow diffusion of electrons in a disordered medium by working in an exact impurity eigenstate representation. We find that the normal diffusion of electrons characterized by a constant diffusion coefficient does not lead to any significant correction to the electron-phonon or the effective electron-electron interactions in a superconductor. We then consider sufficiently strong disorder where Anderson localization of electrons becomes important and determine the effect of localization on the electron-electron interactions. We find that due to localization, the diffusion of electrons becomes anomalous in the sense that the diffusion coefficient becomes scale dependent. This results in an increase in the effective electron-electron interaction with increasing disorder. We propose that this provides a natural explanation for the unusual sensitivity of the transition temperature T/sub c/ of the high T/sub c/ superconductors (T/sub c/ > 10 0 K) to damage effects

  6. Strong Coupling Holography

    CERN Document Server

    Dvali, Gia

    2009-01-01

    We show that whenever a 4-dimensional theory with N particle species emerges as a consistent low energy description of a 3-brane embedded in an asymptotically-flat (4+d)-dimensional space, the holographic scale of high-dimensional gravity sets the strong coupling scale of the 4D theory. This connection persists in the limit in which gravity can be consistently decoupled. We demonstrate this effect for orbifold planes, as well as for the solitonic branes and string theoretic D-branes. In all cases the emergence of a 4D strong coupling scale from bulk holography is a persistent phenomenon. The effect turns out to be insensitive even to such extreme deformations of the brane action that seemingly shield 4D theory from the bulk gravity effects. A well understood example of such deformation is given by large 4D Einstein term in the 3-brane action, which is known to suppress the strength of 5D gravity at short distances and change the 5D Newton's law into the four-dimensional one. Nevertheless, we observe that the ...

  7. The independent prokaryotic origins of eukaryotic fructose-1, 6-bisphosphatase and sedoheptulose-1, 7-bisphosphatase and the implications of their origins for the evolution of eukaryotic Calvin cycle

    Directory of Open Access Journals (Sweden)

    Jiang Yong-Hai

    2012-10-01

    Full Text Available Abstract Background In the Calvin cycle of eubacteria, the dephosphorylations of both fructose-1, 6-bisphosphate (FBP and sedoheptulose-1, 7-bisphosphate (SBP are catalyzed by the same bifunctional enzyme: fructose-1, 6-bisphosphatase/sedoheptulose-1, 7-bisphosphatase (F/SBPase, while in that of eukaryotic chloroplasts by two distinct enzymes: chloroplastic fructose-1, 6-bisphosphatase (FBPase and sedoheptulose-1, 7-bisphosphatase (SBPase, respectively. It was proposed that these two eukaryotic enzymes arose from the divergence of a common ancestral eubacterial bifunctional F/SBPase of mitochondrial origin. However, no specific affinity between SBPase and eubacterial FBPase or F/SBPase can be observed in the previous phylogenetic analyses, and it is hard to explain why SBPase and/or F/SBPase are/is absent from most extant nonphotosynthetic eukaryotes according to this scenario. Results Domain analysis indicated that eubacterial F/SBPase of two different resources contain distinct domains: proteobacterial F/SBPases contain typical FBPase domain, while cyanobacterial F/SBPases possess FBPase_glpX domain. Therefore, like prokaryotic FBPase, eubacterial F/SBPase can also be divided into two evolutionarily distant classes (Class I and II. Phylogenetic analysis based on a much larger taxonomic sampling than previous work revealed that all eukaryotic SBPase cluster together and form a close sister group to the clade of epsilon-proteobacterial Class I FBPase which are gluconeogenesis-specific enzymes, while all eukaryotic chloroplast FBPase group together with eukaryotic cytosolic FBPase and form another distinct clade which then groups with the Class I FBPase of diverse eubacteria. Motif analysis of these enzymes also supports these phylogenetic correlations. Conclusions There are two evolutionarily distant classes of eubacterial bifunctional F/SBPase. Eukaryotic FBPase and SBPase do not diverge from either of them but have two independent origins

  8. Beyond Agrobacterium-Mediated Transformation: Horizontal Gene Transfer from Bacteria to Eukaryotes.

    Science.gov (United States)

    Lacroix, Benoît; Citovsky, Vitaly

    2018-03-03

    Besides the massive gene transfer from organelles to the nuclear genomes, which occurred during the early evolution of eukaryote lineages, the importance of horizontal gene transfer (HGT) in eukaryotes remains controversial. Yet, increasing amounts of genomic data reveal many cases of bacterium-to-eukaryote HGT that likely represent a significant force in adaptive evolution of eukaryotic species. However, DNA transfer involved in genetic transformation of plants by Agrobacterium species has traditionally been considered as the unique example of natural DNA transfer and integration into eukaryotic genomes. Recent discoveries indicate that the repertoire of donor bacterial species and of recipient eukaryotic hosts potentially are much wider than previously thought, including donor bacterial species, such as plant symbiotic nitrogen-fixing bacteria (e.g., Rhizobium etli) and animal bacterial pathogens (e.g., Bartonella henselae, Helicobacter pylori), and recipient species from virtually all eukaryotic clades. Here, we review the molecular pathways and potential mechanisms of these trans-kingdom HGT events and discuss their utilization in biotechnology and research.

  9. Quantitative prediction of shrimp disease incidence via the profiles of gut eukaryotic microbiota.

    Science.gov (United States)

    Xiong, Jinbo; Yu, Weina; Dai, Wenfang; Zhang, Jinjie; Qiu, Qiongfen; Ou, Changrong

    2018-04-01

    One common notion is emerging that gut eukaryotes are commensal or beneficial, rather than detrimental. To date, however, surprisingly few studies have been taken to discern the factors that govern the assembly of gut eukaryotes, despite growing interest in the dysbiosis of gut microbiota-disease relationship. Herein, we firstly explored how the gut eukaryotic microbiotas were assembled over shrimp postlarval to adult stages and a disease progression. The gut eukaryotic communities changed markedly as healthy shrimp aged, and converged toward an adult-microbiota configuration. However, the adult-like stability was distorted by disease exacerbation. A null model untangled that the deterministic processes that governed the gut eukaryotic assembly tended to be more important over healthy shrimp development, whereas this trend was inverted as the disease progressed. After ruling out the baseline of gut eukaryotes over shrimp ages, we identified disease-discriminatory taxa (species level afforded the highest accuracy of prediction) that characteristic of shrimp health status. The profiles of these taxa contributed an overall 92.4% accuracy in predicting shrimp health status. Notably, this model can accurately diagnose the onset of shrimp disease. Interspecies interaction analysis depicted how the disease-discriminatory taxa interacted with one another in sustaining shrimp health. Taken together, our findings offer novel insights into the underlying ecological processes that govern the assembly of gut eukaryotes over shrimp postlarval to adult stages and a disease progression. Intriguingly, the established model can quantitatively and accurately predict the incidences of shrimp disease.

  10. Current Perspectives of Telomerase Structure and Function in Eukaryotes with Emerging Views on Telomerase in Human Parasites.

    Science.gov (United States)

    Dey, Abhishek; Chakrabarti, Kausik

    2018-01-24

    Replicative capacity of a cell is strongly correlated with telomere length regulation. Aberrant lengthening or reduction in the length of telomeres can lead to health anomalies, such as cancer or premature aging. Telomerase is a master regulator for maintaining replicative potential in most eukaryotic cells. It does so by controlling telomere length at chromosome ends. Akin to cancer cells, most single-cell eukaryotic pathogens are highly proliferative and require persistent telomerase activity to maintain constant length of telomere and propagation within their host. Although telomerase is key to unlimited cellular proliferation in both cases, not much was known about the role of telomerase in human parasites (malaria, Trypanosoma , etc.) until recently. Since telomerase regulation is mediated via its own structural components, interactions with catalytic reverse transcriptase and several factors that can recruit and assemble telomerase to telomeres in a cell cycle-dependent manner, we compare and discuss here recent findings in telomerase biology in cancer, aging and parasitic diseases to give a broader perspective of telomerase function in human diseases.

  11. LIGO: The strong belief

    CERN Multimedia

    Antonella Del Rosso

    2016-01-01

    Twenty years of designing, building and testing a number of innovative technologies, with the strong belief that the endeavour would lead to a historic breakthrough. The Bulletin publishes an abstract of the Courier’s interview with Barry Barish, one of the founding fathers of LIGO.   The plots show the signals of gravitational waves detected by the twin LIGO observatories at Livingston, Louisiana, and Hanford, Washington. (Image: Caltech/MIT/LIGO Lab) On 11 February, the Laser Interferometer Gravitational-Wave Observatory (LIGO) and Virgo collaborations published a historic paper in which they showed a gravitational signal emitted by the merger of two black holes. These results come after 20 years of hard work by a large collaboration of scientists operating the two LIGO observatories in the US. Barry Barish, Linde Professor of Physics, Emeritus at the California Institute of Technology and former Director of the Global Design Effort for the Internat...

  12. Strongly interacting Higgs bosons

    International Nuclear Information System (INIS)

    Appelquist, T.; Bernard, C.

    1980-01-01

    The sensitivity of present-energy weak interactions to a strongly interacting heavy-Higgs-boson sector is discussed. The gauged nonlinear sigma model, which is the limit of the linear model as the Higgs-boson mass goes to infinity, is used to organize and catalogue all possible heavy-Higgs-boson effects. As long as the SU(2)/sub L/ x SU(2)/sub R/ symmetry of the Higgs sector is preserved, these effects are found to be small, of the order of the square of the gauge coupling times logarithms (but not powers) of the Higgs-boson mass divided by the W mass. We work in the context of a simplified model with gauge group SU(2)/sub L/; the extension to SU(2)/sub L/ x U(1) is briefly discussed

  13. Structural and biomechanical basis of mitochondrial movement in eukaryotic cells

    Directory of Open Access Journals (Sweden)

    Wu M

    2013-10-01

    Full Text Available Min Wu,1 Aruna Kalyanasundaram,2 Jie Zhu1 1Laboratory of Biomechanics and Engineering, Institute of Biophysics, College of Science, Northwest A&F University, Yangling, Shaanxi, People's Republic of China; 2College of Pharmacology, University of Illinois at Chicago, Chicago, IL, USA Abstract: Mitochondria serve as energy-producing organelles in eukaryotic cells. In addition to providing the energy supply for cells, the mitochondria are also involved in other processes, such as proliferation, differentiation, information transfer, and apoptosis, and play an important role in regulation of cell growth and the cell cycle. In order to achieve these functions, the mitochondria need to move to the corresponding location. Therefore, mitochondrial movement has a crucial role in normal physiologic activity, and any mitochondrial movement disorder will cause irreparable damage to the organism. For example, recent studies have shown that abnormal movement of the mitochondria is likely to be the reason for Charcot–Marie–Tooth disease, amyotrophic lateral sclerosis, Alzheimer's disease, Huntington's disease, Parkinson's disease, and schizophrenia. So, in the cell, especially in the particular polarized cell, the appropriate distribution of mitochondria is crucial to the function and survival of the cell. Mitochondrial movement is mainly associated with the cytoskeleton and related proteins. However, those components play different roles according to cell type. In this paper, we summarize the structural basis of mitochondrial movement, including microtubules, actin filaments, motor proteins, and adaptin, and review studies of the biomechanical mechanisms of mitochondrial movement in different types of cells. Keywords: mitochondrial movement, microtubules, actin filaments, motor proteins, adaptin

  14. Positive selection for unpreferred codon usage in eukaryotic genomes

    Directory of Open Access Journals (Sweden)

    Galagan James E

    2007-07-01

    Full Text Available Abstract Background Natural selection has traditionally been understood as a force responsible for pushing genes to states of higher translational efficiency, whereas lower translational efficiency has been explained by neutral mutation and genetic drift. We looked for evidence of directional selection resulting in increased unpreferred codon usage (and presumably reduced translational efficiency in three divergent clusters of eukaryotic genomes using a simple optimal-codon-based metric (Kp/Ku. Results Here we show that for some genes natural selection is indeed responsible for causing accelerated unpreferred codon substitution, and document the scope of this selection. In Cryptococcus and to a lesser extent Drosophila, we find many genes showing a statistically significant signal of selection for unpreferred codon usage in one or more lineages. We did not find evidence for this type of selection in Saccharomyces. The signal of positive selection observed from unpreferred synonymous codon substitutions is coincident in Cryptococcus and Drosophila with the distribution of upstream open reading frames (uORFs, another genic feature known to reduce translational efficiency. Functional enrichment analysis of genes exhibiting low Kp/Ku ratios reveals that genes in regulatory roles are particularly subject to this type of selection. Conclusion Through genome-wide scans, we find recent selection for unpreferred codon usage at approximately 1% of genetic loci in a Cryptococcus and several genes in Drosophila. Unpreferred codons can impede translation efficiency, and we find that genes with translation-impeding uORFs are enriched for this selection signal. We find that regulatory genes are particularly likely to be subject to selection for unpreferred codon usage. Given that expression noise can propagate through regulatory cascades, and that low translational efficiency can reduce expression noise, this finding supports the hypothesis that translational

  15. MetWAMer: eukaryotic translation initiation site prediction

    Directory of Open Access Journals (Sweden)

    Brendel Volker

    2008-09-01

    Full Text Available Abstract Background Translation initiation site (TIS identification is an important aspect of the gene annotation process, requisite for the accurate delineation of protein sequences from transcript data. We have developed the MetWAMer package for TIS prediction in eukaryotic open reading frames of non-viral origin. MetWAMer can be used as a stand-alone, third-party tool for post-processing gene structure annotations generated by external computational programs and/or pipelines, or directly integrated into gene structure prediction software implementations. Results MetWAMer currently implements five distinct methods for TIS prediction, the most accurate of which is a routine that combines weighted, signal-based translation initiation site scores and the contrast in coding potential of sequences flanking TISs using a perceptron. Also, our program implements clustering capabilities through use of the k-medoids algorithm, thereby enabling cluster-specific TIS parameter utilization. In practice, our static weight array matrix-based indexing method for parameter set lookup can be used with good results in data sets exhibiting moderate levels of 5'-complete coverage. Conclusion We demonstrate that improvements in statistically-based models for TIS prediction can be achieved by taking the class of each potential start-methionine into account pending certain testing conditions, and that our perceptron-based model is suitable for the TIS identification task. MetWAMer represents a well-documented, extensible, and freely available software system that can be readily re-trained for differing target applications and/or extended with existing and novel TIS prediction methods, to support further research efforts in this area.

  16. Methyl labeling and TROSY NMR spectroscopy of proteins expressed in the eukaryote Pichia pastoris

    International Nuclear Information System (INIS)

    Clark, Lindsay; Zahm, Jacob A.; Ali, Rustam; Kukula, Maciej; Bian, Liangqiao; Patrie, Steven M.; Gardner, Kevin H.; Rosen, Michael K.; Rosenbaum, Daniel M.

    2015-01-01

    13 C Methyl TROSY NMR spectroscopy has emerged as a powerful method for studying the dynamics of large systems such as macromolecular assemblies and membrane proteins. Specific 13 C labeling of aliphatic methyl groups and perdeuteration has been limited primarily to proteins expressed in E. coli, preventing studies of many eukaryotic proteins of physiological and biomedical significance. We demonstrate the feasibility of efficient 13 C isoleucine δ1-methyl labeling in a deuterated background in an established eukaryotic expression host, Pichia pastoris, and show that this method can be used to label the eukaryotic protein actin, which cannot be expressed in bacteria. This approach will enable NMR studies of previously intractable targets

  17. Distinct Trajectories of Massive Recent Gene Gains and Losses in Populations of a Microbial Eukaryotic Pathogen.

    Science.gov (United States)

    Hartmann, Fanny E; Croll, Daniel

    2017-11-01

    Differences in gene content are a significant source of variability within species and have an impact on phenotypic traits. However, little is known about the mechanisms responsible for the most recent gene gains and losses. We screened the genomes of 123 worldwide isolates of the major pathogen of wheat Zymoseptoria tritici for robust evidence of gene copy number variation. Based on orthology relationships in three closely related fungi, we identified 599 gene gains and 1,024 gene losses that have not yet reached fixation within the focal species. Our analyses of gene gains and losses segregating in populations showed that gene copy number variation arose preferentially in subtelomeres and in proximity to transposable elements. Recently lost genes were enriched in virulence factors and secondary metabolite gene clusters. In contrast, recently gained genes encoded mostly secreted protein lacking a conserved domain. We analyzed the frequency spectrum at loci segregating a gene presence-absence polymorphism in four worldwide populations. Recent gene losses showed a significant excess in low-frequency variants compared with genome-wide single nucleotide polymorphism, which is indicative of strong negative selection against gene losses. Recent gene gains were either under weak negative selection or neutral. We found evidence for strong divergent selection among populations at individual loci segregating a gene presence-absence polymorphism. Hence, gene gains and losses likely contributed to local adaptation. Our study shows that microbial eukaryotes harbor extensive copy number variation within populations and that functional differences among recently gained and lost genes led to distinct evolutionary trajectories. © The Author 2017. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

  18. Strong-interaction nonuniversality

    International Nuclear Information System (INIS)

    Volkas, R.R.; Foot, R.; He, X.; Joshi, G.C.

    1989-01-01

    The universal QCD color theory is extended to an SU(3) 1 direct product SU(3) 2 direct product SU(3) 3 gauge theory, where quarks of the ith generation transform as triplets under SU(3)/sub i/ and singlets under the other two factors. The usual color group is then identified with the diagonal subgroup, which remains exact after symmetry breaking. The gauge bosons associated with the 16 broken generators then form two massive octets under ordinary color. The interactions between quarks and these heavy gluonlike particles are explicitly nonuniversal and thus an exploration of their physical implications allows us to shed light on the fundamental issue of strong-interaction universality. Nonuniversality and weak flavor mixing are shown to generate heavy-gluon-induced flavor-changing neutral currents. The phenomenology of these processes is studied, as they provide the major experimental constraint on the extended theory. Three symmetry-breaking scenarios are presented. The first has color breaking occurring at the weak scale, while the second and third divorce the two scales. The third model has the interesting feature of radiatively induced off-diagonal Kobayashi-Maskawa matrix elements

  19. John Strong (1941 - 2006)

    CERN Multimedia

    Wickens, F

    Our friend and colleague John Strong was cruelly taken from us by a brain tumour on Monday 31st July, a few days before his 65th birthday John started his career working with a group from Westfield College, under the leadership of Ted Bellamy. He obtained his PhD and spent the early part of his career on experiments at Rutherford Appleton Laboratory (RAL), but after the early 1970s his research was focussed on experiments in CERN. Over the years he made a number of notable contributions to experiments in CERN: The Omega spectrometer adopted a system John had originally developed for experiments at RAL using vidicon cameras to record the sparks in the spark chambers; He contributed to the success of NA1 and NA7, where he became heavily involved in the electronic trigger systems; He was responsible for the second level trigger system for the ALEPH detector and spent five years leading a team that designed and built the system, which ran for twelve years with only minor interventions. Following ALEPH he tur...

  20. Stirring Strongly Coupled Plasma

    CERN Document Server

    Fadafan, Kazem Bitaghsir; Rajagopal, Krishna; Wiedemann, Urs Achim

    2009-01-01

    We determine the energy it takes to move a test quark along a circle of radius L with angular frequency w through the strongly coupled plasma of N=4 supersymmetric Yang-Mills (SYM) theory. We find that for most values of L and w the energy deposited by stirring the plasma in this way is governed either by the drag force acting on a test quark moving through the plasma in a straight line with speed v=Lw or by the energy radiated by a quark in circular motion in the absence of any plasma, whichever is larger. There is a continuous crossover from the drag-dominated regime to the radiation-dominated regime. In the crossover regime we find evidence for significant destructive interference between energy loss due to drag and that due to radiation as if in vacuum. The rotating quark thus serves as a model system in which the relative strength of, and interplay between, two different mechanisms of parton energy loss is accessible via a controlled classical gravity calculation. We close by speculating on the implicati...

  1. Non-coding RNA regulation in pathogenic bacteria located inside eukaryotic cells

    NARCIS (Netherlands)

    Ortega, Alvaro D.; Quereda, Juan J; Pucciarelli, M Graciela; García-del Portillo, Francisco

    2014-01-01

    Intracellular bacterial pathogens have evolved distinct lifestyles inside eukaryotic cells. Some pathogens coexist with the infected cell in an obligate intracellular state, whereas others transit between the extracellular and intracellular environment. Adaptation to these intracellular lifestyles

  2. Extracellular DNA amplicon sequencing reveals high levels of benthic eukaryotic diversity in the central Red Sea

    KAUST Repository

    Pearman, John K.; Irigoien, Xabier; Carvalho, Susana

    2015-01-01

    The present study aims to characterize the benthic eukaryotic biodiversity patterns at a coarse taxonomic level in three areas of the central Red Sea (a lagoon, an offshore area in Thuwal and a shallow coastal area near Jeddah) based

  3. Meeting Report: Minutes from EMBO: Ten Years of Comparative Genomics of Eukaryotic Microorganisms

    Czech Academy of Sciences Publication Activity Database

    Lukeš, Julius; López-García, P.; Louis, E.; Boekhout, T.

    2016-01-01

    Roč. 167, č. 3 (2016), s. 217-221 ISSN 1434-4610 Institutional support: RVO:60077344 Keywords : protist * eukaryotic microorganisms * genomics Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.794, year: 2016

  4. Phylogenetic analysis of P5 P-type ATPases, a eukaryotic lineage of secretory pathway pumps

    DEFF Research Database (Denmark)

    Møller, Annette; Asp, Torben; Holm, Preben Bach

    2008-01-01

    prokaryotic genome. Based on a protein alignment we could group the P5 ATPases into two subfamilies, P5A and P5B that, based on the number of negative charges in conserved trans-membrane segment 4, are likely to have different ion specificities. P5A ATPases are present in all eukaryotic genomes sequenced so......Eukaryotes encompass a remarkable variety of organisms and unresolved lineages. Different phylogenetic analyses have lead to conflicting conclusions as to the origin and associations between lineages and species. In this work, we investigated evolutionary relationship of a family of cation pumps...... exclusive for the secretory pathway of eukaryotes by combining the identification of lineage-specific genes with phylogenetic evolution of common genes. Sequences of P5 ATPases, which are regarded to be cation pumps in the endoplasmic reticulum (ER), were identified in all eukaryotic lineages but not in any...

  5. Changes in bacterial and eukaryotic communities during sewage decomposition in Mississippi River water

    Science.gov (United States)

    Microbial decay processes are one of the mechanisms whereby sewage contamination is reduced in the environment. This decomposition process involves a highly complex array of bacterial and eukaryotic communities from both sewage and ambient waters. However, relatively little is kn...

  6. Automated Eukaryotic Gene Structure Annotation Using EVidenceModeler and the Program to Assemble Spliced Alignments

    Energy Technology Data Exchange (ETDEWEB)

    Haas, B J; Salzberg, S L; Zhu, W; Pertea, M; Allen, J E; Orvis, J; White, O; Buell, C R; Wortman, J R

    2007-12-10

    EVidenceModeler (EVM) is presented as an automated eukaryotic gene structure annotation tool that reports eukaryotic gene structures as a weighted consensus of all available evidence. EVM, when combined with the Program to Assemble Spliced Alignments (PASA), yields a comprehensive, configurable annotation system that predicts protein-coding genes and alternatively spliced isoforms. Our experiments on both rice and human genome sequences demonstrate that EVM produces automated gene structure annotation approaching the quality of manual curation.

  7. Chloride channels in the plasma membrane of a foetal Drosophila cell line, S2

    DEFF Research Database (Denmark)

    Asmild, Margit; Willumsen, Niels J.

    2000-01-01

    S2 cells, Cl- Channels, Expression system, Drosophila, Inward rectifier, Outward rectifier, Patch clamp......S2 cells, Cl- Channels, Expression system, Drosophila, Inward rectifier, Outward rectifier, Patch clamp...

  8. Mapping and characterizing N6-methyladenine in eukaryotic genomes using single molecule real-time sequencing.

    Science.gov (United States)

    Zhu, Shijia; Beaulaurier, John; Deikus, Gintaras; Wu, Tao; Strahl, Maya; Hao, Ziyang; Luo, Guanzheng; Gregory, James A; Chess, Andrew; He, Chuan; Xiao, Andrew; Sebra, Robert; Schadt, Eric E; Fang, Gang

    2018-05-15

    N6-methyladenine (m6dA) has been discovered as a novel form of DNA methylation prevalent in eukaryotes, however, methods for high resolution mapping of m6dA events are still lacking. Single-molecule real-time (SMRT) sequencing has enabled the detection of m6dA events at single-nucleotide resolution in prokaryotic genomes, but its application to detecting m6dA in eukaryotic genomes has not been rigorously examined. Herein, we identified unique characteristics of eukaryotic m6dA methylomes that fundamentally differ from those of prokaryotes. Based on these differences, we describe the first approach for mapping m6dA events using SMRT sequencing specifically designed for the study of eukaryotic genomes, and provide appropriate strategies for designing experiments and carrying out sequencing in future studies. We apply the novel approach to study two eukaryotic genomes. For green algae, we construct the first complete genome-wide map of m6dA at single nucleotide and single molecule resolution. For human lymphoblastoid cells (hLCLs), joint analyses of SMRT sequencing and independent sequencing data suggest that putative m6dA events are enriched in the promoters of young, full length LINE-1 elements (L1s). These analyses demonstrate a general method for rigorous mapping and characterization of m6dA events in eukaryotic genomes. Published by Cold Spring Harbor Laboratory Press.

  9. Diversity and distribution of eukaryotic microbes in and around a brine pool adjacent to the Thuwal cold seeps in the Red Sea

    KAUST Repository

    Wang, Yong

    2014-02-04

    A hypoxic/suboxic brine pool at a depth of about 850 m was discovered near the Thuwal cold seeps in the Red Sea. Filled with high concentrations of hydrogen sulfide and ammonia, such a brine pool might limit the spread of eukaryotic organisms. Here, we compared the communities of the eukaryotic microbes in a microbial mat, sediments and water samples distributed in 7 sites within and adjacent to the brine pool. Taxonomic classification of the pyrosequenced 18S rRNA amplicon reads showed that fungi highly similar to the species identified along the Arabic coast were almost ubiquitous in the water and sediment samples, supporting their wide distribution in various environments. The microbial mat displayed the highest species diversity and contained grazers and a considerable percentage of unclassified species. Phylogeny-based methods revealed novel lineages representing a majority of the reads from the interface between the sea water and brine pool. Phylogenetic relationships with more reference sequences suggest that the lineages were affiliated with novel Alveolata and Euglenozoa inhabiting the interface where chemosynthetic prokaryotes are highly proliferative due to the strong chemocline and halocline. The brine sediments harbored abundant species highly similar to invertebrate gregarine parasites identified in different oxygen-depleted sediments. Therefore, the present findings support the uniqueness of some microbial eukaryotic groups in this cold seep brine system. 2014 Wang, Zhang, Cao, Shek, Tian, Wong, Batang, Al-suwailem and Qian.

  10. The interplay between genome and network evolution in eukaryotes

    NARCIS (Netherlands)

    Fokkens, L.

    2013-01-01

    We assume that the functional relations that a protein engages in, influences it’s evolutionary dynamics. An extreme case is a functional module in which all components are strongly dependent on each other, for example a protein complex or metabolic pathway. If a protein’s function is only relevant

  11. Compound-specific amino acid δ15N patterns in marine algae: Tracer potential for cyanobacterial vs. eukaryotic organic nitrogen sources in the ocean

    Science.gov (United States)

    McCarthy, Matthew D.; Lehman, Jennifer; Kudela, Raphael

    2013-02-01

    Stable nitrogen isotopic analysis of individual amino acids (δ15N-AA) has unique potential to elucidate the complexities of food webs, track heterotrophic transformations, and understand diagenesis of organic nitrogen (ON). While δ15N-AA patterns of autotrophs have been shown to be generally similar, prior work has also suggested that differences may exist between cyanobacteria and eukaryotic algae. However, δ15N-AA patterns in differing oceanic algal groups have never been closely examined. The overarching goals of this study were first to establish a more quantitative understanding of algal δ15N-AA patterns, and second to examine whether δ15N-AA patterns have potential as a new tracer for distinguishing prokaryotic vs. eukaryotic N sources. We measured δ15N-AA from prokaryotic and eukaryotic phytoplankton cultures and used a complementary set of statistical approaches (simple normalization, regression-derived fractionation factors, and multivariate analyses) to test for variations. A generally similar δ15N-AA pattern was confirmed for all algae, however significant AA-specific variation was also consistently identified between the two groups. The relative δ15N fractionation of Glx (glutamine + glutamic acid combined) vs. total proteinaceous N appeared substantially different, which we hypothesize could be related to differing enzymatic forms. In addition, the several other AA (most notably glycine and leucine) appeared to have strong biomarker potential. Finally, we observed that overall patterns of δ15N values in algae correspond well with the Trophic vs. Source-AA division now commonly used to describe variable AA δ15N changes with trophic transfer, suggesting a common mechanistic basis. Overall, these results show that autotrophic δ15N-AA patterns can differ between major algal evolutionary groupings for many AA. The statistically significant multivariate results represent a first approach for testing ideas about relative eukaryotic vs. prokaryotic

  12. C-terminal motif prediction in eukaryotic proteomes using comparative genomics and statistical over-representation across protein families

    Directory of Open Access Journals (Sweden)

    Cutler Sean R

    2007-06-01

    between species, among kingdoms and across eukaryotes. Motifs of note include a serine-acidic peptide (DSD* as well as several lysine enriched motifs found in nearly all eukaryotic genomes examined. Conclusion We have successfully generated a high confidence representation of eukaryotic motifs anchored at the C-terminus. A high incidence of true-positives in our results suggests that several previously unidentified tripeptide patterns are strong candidates for representing novel peptide motifs of a widely employed nature in the C-terminal biology of eukaryotes. Our application of comparative genomics, statistical over-representation and the adjustment for protein family homology has generated several hypotheses concerning the C-terminal topology as it pertains to sorting and potential protein interaction signals. This approach to background reduction could be expanded for application to protein motif prediction in the protein interior. A parallel N-terminal analysis is presented as supplementary data.

  13. Eukaryotic evolutionary transitions are associated with extreme codon bias in functionally-related proteins.

    Directory of Open Access Journals (Sweden)

    Nicholas J Hudson

    Full Text Available Codon bias in the genome of an organism influences its phenome by changing the speed and efficiency of mRNA translation and hence protein abundance. We hypothesized that differences in codon bias, either between-species differences in orthologous genes, or within-species differences between genes, may play an evolutionary role. To explore this hypothesis, we compared the genome-wide codon bias in six species that occupy vital positions in the Eukaryotic Tree of Life. We acquired the entire protein coding sequences for these organisms, computed the codon bias for all genes in each organism and explored the output for relationships between codon bias and protein function, both within- and between-lineages. We discovered five notable coordinated patterns, with extreme codon bias most pronounced in traits considered highly characteristic of a given lineage. Firstly, the Homo sapiens genome had stronger codon bias for DNA-binding transcription factors than the Saccharomyces cerevisiae genome, whereas the opposite was true for ribosomal proteins--perhaps underscoring transcriptional regulation in the origin of complexity. Secondly, both mammalian species examined possessed extreme codon bias in genes relating to hair--a tissue unique to mammals. Thirdly, Arabidopsis thaliana showed extreme codon bias in genes implicated in cell wall formation and chloroplast function--which are unique to plants. Fourthly, Gallus gallus possessed strong codon bias in a subset of genes encoding mitochondrial proteins--perhaps reflecting the enhanced bioenergetic efficiency in birds that co-evolved with flight. And lastly, the G. gallus genome had extreme codon bias for the Ciliary Neurotrophic Factor--which may help to explain their spontaneous recovery from deafness. We propose that extreme codon bias in groups of genes that encode functionally related proteins has a pathway-level energetic explanation.

  14. Dissecting the expression relationships between RNA-binding proteins and their cognate targets in eukaryotic post-transcriptional regulatory networks

    Science.gov (United States)

    Nishtala, Sneha; Neelamraju, Yaseswini; Janga, Sarath Chandra

    2016-05-01

    RNA-binding proteins (RBPs) are pivotal in orchestrating several steps in the metabolism of RNA in eukaryotes thereby controlling an extensive network of RBP-RNA interactions. Here, we employed CLIP (cross-linking immunoprecipitation)-seq datasets for 60 human RBPs and RIP-ChIP (RNP immunoprecipitation-microarray) data for 69 yeast RBPs to construct a network of genome-wide RBP- target RNA interactions for each RBP. We show in humans that majority (~78%) of the RBPs are strongly associated with their target transcripts at transcript level while ~95% of the studied RBPs were also found to be strongly associated with expression levels of target transcripts when protein expression levels of RBPs were employed. At transcript level, RBP - RNA interaction data for the yeast genome, exhibited a strong association for 63% of the RBPs, confirming the association to be conserved across large phylogenetic distances. Analysis to uncover the features contributing to these associations revealed the number of target transcripts and length of the selected protein-coding transcript of an RBP at the transcript level while intensity of the CLIP signal, number of RNA-Binding domains, location of the binding site on the transcript, to be significant at the protein level. Our analysis will contribute to improved modelling and prediction of post-transcriptional networks.

  15. Metabarcoding analysis of eukaryotic microbiota in the gut of HIV-infected patients.

    Directory of Open Access Journals (Sweden)

    Ibrahim Hamad

    Full Text Available Research on the relationship between changes in the gut microbiota and human disease, including AIDS, is a growing field. However, studies on the eukaryotic component of the intestinal microbiota have just begun and have not yet been conducted in HIV-infected patients. Moreover, eukaryotic community profiling is influenced by the use of different methodologies at each step of culture-independent techniques. Herein, initially, four DNA extraction protocols were compared to test the efficiency of each method in recovering eukaryotic DNA from fecal samples. Our results revealed that recovering eukaryotic components from fecal samples differs significantly among DNA extraction methods. Subsequently, the composition of the intestinal eukaryotic microbiota was evaluated in HIV-infected patients and healthy volunteers through clone sequencing, high-throughput sequencing of nuclear ribosomal internal transcribed spacers 1 (ITS1 and 2 (ITS2 amplicons and real-time PCRs. Our results revealed that not only richness (Chao-1 index and alpha diversity (Shannon diversity differ between HIV-infected patients and healthy volunteers, depending on the molecular strategy used, but also the global eukaryotic community composition, with little overlapping taxa found between techniques. Moreover, our results based on cloning libraries and ITS1/ITS2 metabarcoding sequencing showed significant differences in fungal composition between HIV-infected patients and healthy volunteers, but without distinct clusters separating the two groups. Malassezia restricta was significantly more prevalent in fecal samples of HIV-infected patients, according to cloning libraries, whereas operational taxonomic units (OTUs belonging to Candida albicans and Candida tropicalis were significantly more abundant in fecal samples of HIV-infected patients compared to healthy subjects in both ITS subregions. Finally, real-time PCR showed the presence of Microsporidia, Giardia lamblia, Blastocystis

  16. Proton-pumping rhodopsins are abundantly expressed by microbial eukaryotes in a high-Arctic fjord.

    Science.gov (United States)

    Vader, Anna; Laughinghouse, Haywood D; Griffiths, Colin; Jakobsen, Kjetill S; Gabrielsen, Tove M

    2018-02-01

    Proton-pumping rhodopsins provide an alternative pathway to photosynthesis by which solar energy can enter the marine food web. Rhodopsin genes are widely found in marine bacteria, also in the Arctic, and were recently reported from several eukaryotic lineages. So far, little is known about rhodopsin expression in Arctic eukaryotes. In this study, we used metatranscriptomics and 18S rDNA tag sequencing to examine the mid-summer function and composition of marine protists (size 0.45-10 µm) in the high-Arctic Billefjorden (Spitsbergen), especially focussing on the expression of microbial proton-pumping rhodopsins. Rhodopsin transcripts were highly abundant, at a level similar to that of genes involved in photosynthesis. Phylogenetic analyses placed the environmental rhodopsins within disparate eukaryotic lineages, including dinoflagellates, stramenopiles, haptophytes and cryptophytes. Sequence comparison indicated the presence of several functional types, including xanthorhodopsins and a eukaryotic clade of proteorhodopsin. Transcripts belonging to the proteorhodopsin clade were also abundant in published metatranscriptomes from other oceanic regions, suggesting a global distribution. The diversity and abundance of rhodopsins show that these light-driven proton pumps play an important role in Arctic microbial eukaryotes. Understanding this role is imperative to predicting the future of the Arctic marine ecosystem faced by a changing light climate due to diminishing sea-ice. © 2017 Society for Applied Microbiology and John Wiley & Sons Ltd.

  17. Arginine deiminase pathway enzymes: evolutionary history in metamonads and other eukaryotes.

    Science.gov (United States)

    Novák, Lukáš; Zubáčová, Zuzana; Karnkowska, Anna; Kolisko, Martin; Hroudová, Miluše; Stairs, Courtney W; Simpson, Alastair G B; Keeling, Patrick J; Roger, Andrew J; Čepička, Ivan; Hampl, Vladimír

    2016-10-06

    Multiple prokaryotic lineages use the arginine deiminase (ADI) pathway for anaerobic energy production by arginine degradation. The distribution of this pathway among eukaryotes has been thought to be very limited, with only two specialized groups living in low oxygen environments (Parabasalia and Diplomonadida) known to possess the complete set of all three enzymes. We have performed an extensive survey of available sequence data in order to map the distribution of these enzymes among eukaryotes and to reconstruct their phylogenies. We have found genes for the complete pathway in almost all examined representatives of Metamonada, the anaerobic protist group that includes parabasalids and diplomonads. Phylogenetic analyses indicate the presence of the complete pathway in the last common ancestor of metamonads and heterologous transformation experiments suggest its cytosolic localization in the metamonad ancestor. Outside Metamonada, the complete pathway occurs rarely, nevertheless, it was found in representatives of most major eukaryotic clades. Phylogenetic relationships of complete pathways are consistent with the presence of the Archaea-derived ADI pathway in the last common ancestor of all eukaryotes, although other evolutionary scenarios remain possible. The presence of the incomplete set of enzymes is relatively common among eukaryotes and it may be related to the fact that these enzymes are involved in other cellular processes, such as the ornithine-urea cycle. Single protein phylogenies suggest that the evolutionary history of all three enzymes has been shaped by frequent gene losses and horizontal transfers, which may sometimes be connected with their diverse roles in cellular metabolism.

  18. Evolution of glutamate dehydrogenase genes: evidence for lateral gene transfer within and between prokaryotes and eukaryotes

    Directory of Open Access Journals (Sweden)

    Roger Andrew J

    2003-06-01

    Full Text Available Abstract Background Lateral gene transfer can introduce genes with novel functions into genomes or replace genes with functionally similar orthologs or paralogs. Here we present a study of the occurrence of the latter gene replacement phenomenon in the four gene families encoding different classes of glutamate dehydrogenase (GDH, to evaluate and compare the patterns and rates of lateral gene transfer (LGT in prokaryotes and eukaryotes. Results We extend the taxon sampling of gdh genes with nine new eukaryotic sequences and examine the phylogenetic distribution pattern of the various GDH classes in combination with maximum likelihood phylogenetic analyses. The distribution pattern analyses indicate that LGT has played a significant role in the evolution of the four gdh gene families. Indeed, a number of gene transfer events are identified by phylogenetic analyses, including numerous prokaryotic intra-domain transfers, some prokaryotic inter-domain transfers and several inter-domain transfers between prokaryotes and microbial eukaryotes (protists. Conclusion LGT has apparently affected eukaryotes and prokaryotes to a similar extent within the gdh gene families. In the absence of indications that the evolution of the gdh gene families is radically different from other families, these results suggest that gene transfer might be an important evolutionary mechanism in microbial eukaryote genome evolution.

  19. Diversity of Eukaryotic Translational Initiation Factor eIF4E in Protists.

    Science.gov (United States)

    Jagus, Rosemary; Bachvaroff, Tsvetan R; Joshi, Bhavesh; Place, Allen R

    2012-01-01

    The greatest diversity of eukaryotic species is within the microbial eukaryotes, the protists, with plants and fungi/metazoa representing just two of the estimated seventy five lineages of eukaryotes. Protists are a diverse group characterized by unusual genome features and a wide range of genome sizes from 8.2 Mb in the apicomplexan parasite Babesia bovis to 112,000-220,050 Mb in the dinoflagellate Prorocentrum micans. Protists possess numerous cellular, molecular and biochemical traits not observed in "text-book" model organisms. These features challenge some of the concepts and assumptions about the regulation of gene expression in eukaryotes. Like multicellular eukaryotes, many protists encode multiple eIF4Es, but few functional studies have been undertaken except in parasitic species. An earlier phylogenetic analysis of protist eIF4Es indicated that they cannot be grouped within the three classes that describe eIF4E family members from multicellular organisms. Many more protist sequences are now available from which three clades can be recognized that are distinct from the plant/fungi/metazoan classes. Understanding of the protist eIF4Es will be facilitated as more sequences become available particularly for the under-represented opisthokonts and amoebozoa. Similarly, a better understanding of eIF4Es within each clade will develop as more functional studies of protist eIF4Es are completed.

  20. Insights into the diversity of eukaryotes in acid mine drainage biofilm communities.

    Science.gov (United States)

    Baker, Brett J; Tyson, Gene W; Goosherst, Lindsey; Banfield, Jillian F

    2009-04-01

    Microscopic eukaryotes are known to have important ecosystem functions, but their diversity in most environments remains vastly unexplored. Here we analyzed an 18S rRNA gene library from a subsurface iron- and sulfur-oxidizing microbial community growing in highly acidic (pH morphological characterization. Results revealed that the populations vary significantly with the habitat and no group is ubiquitous. Surprisingly, many of the eukaryotic lineages (with the exception of the APC) are closely related to neutrophiles, suggesting that they recently adapted to this extreme environment. Molecular analyses presented here confirm that the number of eukaryotic species associated with the acid mine drainage (AMD) communities is low. This finding is consistent with previous results showing a limited diversity of archaea, bacteria, and viruses in AMD environments and suggests that the environmental pressures and interplay between the members of these communities limit species diversity at all trophic levels.

  1. A second pathway to degrade pyrimidine nucleic acid precursors in eukaryotes

    DEFF Research Database (Denmark)

    Andersen, Gorm; Bjornberg, Olof; Polakova, Silvia

    2008-01-01

    Pyrimidine bases are the central precursors for RNA and DNA, and their intracellular pools are determined by de novo, salvage and catabolic pathways. In eukaryotes, degradation of uracil has been believed to proceed only via the reduction to dihydrouracil. Using a yeast model, Saccharomyces kluyv...... of the eukaryotic or prokaryotic genes involved in pyrimidine degradation described to date.......Pyrimidine bases are the central precursors for RNA and DNA, and their intracellular pools are determined by de novo, salvage and catabolic pathways. In eukaryotes, degradation of uracil has been believed to proceed only via the reduction to dihydrouracil. Using a yeast model, Saccharomyces......, respectively. The gene products of URC1 and URC4 are highly conserved proteins with so far unknown functions and they are present in a variety of prokaryotes and fungi. In bacteria and in some fungi, URC1 and URC4 are linked on the genome together with the gene for uracil phosphoribosyltransferase (URC6). Urc1...

  2. Metabolism in anoxic permeable sediments is dominated by eukaryotic dark fermentation

    DEFF Research Database (Denmark)

    Bourke, Michael F.; Marriott, Philip J.; Glud, Ronnie N.

    2017-01-01

    Permeable sediments are common across continental shelves and are critical contributors to marine biogeochemical cycling. Organic matter in permeable sediments is dominated by microalgae, which as eukaryotes have different anaerobic metabolic pathways to prokaryotes such as bacteria and archaea....... Here we present analyses of flow-through reactor experiments showing that dissolved inorganic carbon is produced predominantly as a result of anaerobic eukaryotic metabolic activity. In our experiments, anaerobic production of dissolved inorganic carbon was consistently accompanied by large dissolved H....../hydrogenase pathway of fermentative eukaryotic H2 production, suggesting that pathway as the source of H2 and dissolved inorganic carbon production. Metabolomic analysis showed large increases in lipid production at the onset of anoxia, consistent with documented pathways of anoxic dark fermentation in microalgae...

  3. Large-scale analysis of phosphorylation site occupancy in eukaryotic proteins

    DEFF Research Database (Denmark)

    Rao, R Shyama Prasad; Møller, Ian Max

    2012-01-01

    in proteins is currently lacking. We have therefore analyzed the occurrence and occupancy of phosphorylated sites (~ 100,281) in a large set of eukaryotic proteins (~ 22,995). Phosphorylation probability was found to be much higher in both the  termini of protein sequences and this is much pronounced...... maximum randomness. An analysis of phosphorylation motifs indicated that just 40 motifs and a much lower number of associated kinases might account for nearly 50% of the known phosphorylations in eukaryotic proteins. Our results provide a broad picture of the phosphorylation sites in eukaryotic proteins.......Many recent high throughput technologies have enabled large-scale discoveries of new phosphorylation sites and phosphoproteins. Although they have provided a number of insights into protein phosphorylation and the related processes, an inclusive analysis on the nature of phosphorylated sites...

  4. Initiation of translation in bacteria by a structured eukaryotic IRES RNA.

    Science.gov (United States)

    Colussi, Timothy M; Costantino, David A; Zhu, Jianyu; Donohue, John Paul; Korostelev, Andrei A; Jaafar, Zane A; Plank, Terra-Dawn M; Noller, Harry F; Kieft, Jeffrey S

    2015-03-05

    The central dogma of gene expression (DNA to RNA to protein) is universal, but in different domains of life there are fundamental mechanistic differences within this pathway. For example, the canonical molecular signals used to initiate protein synthesis in bacteria and eukaryotes are mutually exclusive. However, the core structures and conformational dynamics of ribosomes that are responsible for the translation steps that take place after initiation are ancient and conserved across the domains of life. We wanted to explore whether an undiscovered RNA-based signal might be able to use these conserved features, bypassing mechanisms specific to each domain of life, and initiate protein synthesis in both bacteria and eukaryotes. Although structured internal ribosome entry site (IRES) RNAs can manipulate ribosomes to initiate translation in eukaryotic cells, an analogous RNA structure-based mechanism has not been observed in bacteria. Here we report our discovery that a eukaryotic viral IRES can initiate translation in live bacteria. We solved the crystal structure of this IRES bound to a bacterial ribosome to 3.8 Å resolution, revealing that despite differences between bacterial and eukaryotic ribosomes this IRES binds directly to both and occupies the space normally used by transfer RNAs. Initiation in both bacteria and eukaryotes depends on the structure of the IRES RNA, but in bacteria this RNA uses a different mechanism that includes a form of ribosome repositioning after initial recruitment. This IRES RNA bridges billions of years of evolutionary divergence and provides an example of an RNA structure-based translation initiation signal capable of operating in two domains of life.

  5. The reduced kinome of Ostreococcus tauri: core eukaryotic signalling components in a tractable model species.

    Science.gov (United States)

    Hindle, Matthew M; Martin, Sarah F; Noordally, Zeenat B; van Ooijen, Gerben; Barrios-Llerena, Martin E; Simpson, T Ian; Le Bihan, Thierry; Millar, Andrew J

    2014-08-02

    The current knowledge of eukaryote signalling originates from phenotypically diverse organisms. There is a pressing need to identify conserved signalling components among eukaryotes, which will lead to the transfer of knowledge across kingdoms. Two useful properties of a eukaryote model for signalling are (1) reduced signalling complexity, and (2) conservation of signalling components. The alga Ostreococcus tauri is described as the smallest free-living eukaryote. With less than 8,000 genes, it represents a highly constrained genomic palette. Our survey revealed 133 protein kinases and 34 protein phosphatases (1.7% and 0.4% of the proteome). We conducted phosphoproteomic experiments and constructed domain structures and phylogenies for the catalytic protein-kinases. For each of the major kinases families we review the completeness and divergence of O. tauri representatives in comparison to the well-studied kinomes of the laboratory models Arabidopsis thaliana and Saccharomyces cerevisiae, and of Homo sapiens. Many kinase clades in O. tauri were reduced to a single member, in preference to the loss of family diversity, whereas TKL and ABC1 clades were expanded. We also identified kinases that have been lost in A. thaliana but retained in O. tauri. For three, contrasting eukaryotic pathways - TOR, MAPK, and the circadian clock - we established the subset of conserved components and demonstrate conserved sites of substrate phosphorylation and kinase motifs. We conclude that O. tauri satisfies our two central requirements. Several of its kinases are more closely related to H. sapiens orthologs than S. cerevisiae is to H. sapiens. The greatly reduced kinome of O. tauri is therefore a suitable model for signalling in free-living eukaryotes.

  6. Convergent use of RhoGAP toxins by eukaryotic parasites and bacterial pathogens.

    Directory of Open Access Journals (Sweden)

    Dominique Colinet

    2007-12-01

    Full Text Available Inactivation of host Rho GTPases is a widespread strategy employed by bacterial pathogens to manipulate mammalian cellular functions and avoid immune defenses. Some bacterial toxins mimic eukaryotic Rho GTPase-activating proteins (GAPs to inactivate mammalian GTPases, probably as a result of evolutionary convergence. An intriguing question remains whether eukaryotic pathogens or parasites may use endogenous GAPs as immune-suppressive toxins to target the same key genes as bacterial pathogens. Interestingly, a RhoGAP domain-containing protein, LbGAP, was recently characterized from the parasitoid wasp Leptopilina boulardi, and shown to protect parasitoid eggs from the immune response of Drosophila host larvae. We demonstrate here that LbGAP has structural characteristics of eukaryotic RhoGAPs but that it acts similarly to bacterial RhoGAP toxins in mammals. First, we show by immunocytochemistry that LbGAP enters Drosophila immune cells, plasmatocytes and lamellocytes, and that morphological changes in lamellocytes are correlated with the quantity of LbGAP they contain. Demonstration that LbGAP displays a GAP activity and specifically interacts with the active, GTP-bound form of the two Drosophila Rho GTPases Rac1 and Rac2, both required for successful encapsulation of Leptopilina eggs, was then achieved using biochemical tests, yeast two-hybrid analysis, and GST pull-down assays. In addition, we show that the overall structure of LbGAP is similar to that of eukaryotic RhoGAP domains, and we identify distinct residues involved in its interaction with Rac GTPases. Altogether, these results show that eukaryotic parasites can use endogenous RhoGAPs as virulence factors and that despite their differences in sequence and structure, eukaryotic and bacterial RhoGAP toxins are similarly used to target the same immune pathways in insects and mammals.

  7. Revisiting the Relationship between Transposable Elements and the Eukaryotic Stress Response.

    Science.gov (United States)

    Horváth, Vivien; Merenciano, Miriam; González, Josefa

    2017-11-01

    A relationship between transposable elements (TEs) and the eukaryotic stress response was suggested in the first publications describing TEs. Since then, it has often been assumed that TEs are activated by stress, and that this activation is often beneficial for the organism. In recent years, the availability of new high-throughput experimental techniques has allowed further interrogation of the relationship between TEs and stress. By reviewing the recent literature, we conclude that although there is evidence for a beneficial effect of TE activation under stress conditions, the relationship between TEs and the eukaryotic stress response is quite complex. Copyright © 2017 Elsevier Ltd. All rights reserved.

  8. Comparative radiobiology of genetic loci of eukaryots as the basis of the general theory of mutations

    International Nuclear Information System (INIS)

    Aleksandrov, I.D.

    1983-01-01

    One of the fundamental problems of modern molecular cellular radiobiology is to reveal general and peculiar processes of the formation of gene mutations and chromosome aberrations in each stage of their formation in the irradiated genome of the higher eukaryots. The solution of the problems depends on the development of research within the framework of comparative radiobiology of genetic loci of the higher eukaryots that makes it possible to study quantitative regularities in the formation of gene (point) mutations and chromosome aberrations in one object and in the same experiment

  9. Once in a lifetime: strategies for preventing re-replication in prokaryotic and eukaryotic cells

    DEFF Research Database (Denmark)

    Nielsen, Olaf; Løbner-Olesen, Anders

    2008-01-01

    DNA replication is an extremely accurate process and cells have evolved intricate control mechanisms to ensure that each region of their genome is replicated only once during S phase. Here, we compare what is known about the processes that prevent re-replication in prokaryotic and eukaryotic cells...... prokaryotes and eukaryotes are inactivated until the next cell cycle. Furthermore, in both systems the beta-clamp of the replicative polymerase associates with enzymatic activities that contribute to the inactivation of the helicase loaders. Finally, recent studies suggest that the control mechanism...

  10. Signaling mechanisms of apoptosis-like programmed cell death in unicellular eukaryotes.

    Science.gov (United States)

    Shemarova, Irina V

    2010-04-01

    In unicellular eukaryotes, apoptosis-like cell death occurs during development, aging and reproduction, and can be induced by environmental stresses and exposure to toxic agents. The essence of the apoptotic machinery in unicellular organisms is similar to that in mammals, but the apoptotic signal network is less complex and of more ancient origin. The review summarizes current data about key apoptotic proteins and mechanisms of the transduction of apoptotic signals by caspase-like proteases and mitochondrial apoptogenic proteins in unicellular eukaryotes. The roles of receptor-dependent and receptor-independent caspase cascades are reviewed. 2010 Elsevier Inc. All rights reserved.

  11. Eukaryotic RNA polymerase subunit RPB8 is a new relative of the OB family.

    Science.gov (United States)

    Krapp, S; Kelly, G; Reischl, J; Weinzierl, R O; Matthews, S

    1998-02-01

    RNA polymerase II subunit RPB8 is an essential subunit that is highly conserved throughout eukaryotic evolution and is present in all three types of nuclear RNA polymerases. We report the first high resolution structural insight into eukaryotic RNA polymerase architecture with the solution structure of RPB8 from Saccharomyces cerevisiae. It consists of an eight stranded, antiparallel beta-barrel, four short helical regions and a large, unstructured omega-loop. The strands are connected in classic Greek-key fashion. The overall topology is unusual and contains a striking C2 rotational symmetry. Furthermore, it is most likely a novel associate of the oligonucleotide/oligosaccharide (OB) binding protein class.

  12. Quantum electrodynamics of strong fields

    International Nuclear Information System (INIS)

    Greiner, W.

    1983-01-01

    Quantum Electrodynamics of Strong Fields provides a broad survey of the theoretical and experimental work accomplished, presenting papers by a group of international researchers who have made significant contributions to this developing area. Exploring the quantum theory of strong fields, the volume focuses on the phase transition to a charged vacuum in strong electric fields. The contributors also discuss such related topics as QED at short distances, precision tests of QED, nonperturbative QCD and confinement, pion condensation, and strong gravitational fields In addition, the volume features a historical paper on the roots of quantum field theory in the history of quantum physics by noted researcher Friedrich Hund

  13. Instabilities in strongly coupled plasmas

    CERN Document Server

    Kalman, G J

    2003-01-01

    The conventional Vlasov treatment of beam-plasma instabilities is inappropriate when the plasma is strongly coupled. In the strongly coupled liquid state, the strong correlations between the dust grains fundamentally affect the conditions for instability. In the crystalline state, the inherent anisotropy couples the longitudinal and transverse polarizations, and results in unstable excitations in both polarizations. We summarize analyses of resonant and non-resonant, as well as resistive instabilities. We consider both ion-dust streaming and dust beam-plasma instabilities. Strong coupling, in general, leads to an enhancement of the growth rates. In the crystalline phase, a resonant transverse instability can be excited.

  14. The scale-free dynamics of eukaryotic cells.

    Directory of Open Access Journals (Sweden)

    Miguel A Aon

    Full Text Available Temporal organization of biological processes requires massively parallel processing on a synchronized time-base. We analyzed time-series data obtained from the bioenergetic oscillatory outputs of Saccharomyces cerevisiae and isolated cardiomyocytes utilizing Relative Dispersional (RDA and Power Spectral (PSA analyses. These analyses revealed broad frequency distributions and evidence for long-term memory in the observed dynamics. Moreover RDA and PSA showed that the bioenergetic dynamics in both systems show fractal scaling over at least 3 orders of magnitude, and that this scaling obeys an inverse power law. Therefore we conclude that in S. cerevisiae and cardiomyocytes the dynamics are scale-free in vivo. Applying RDA and PSA to data generated from an in silico model of mitochondrial function indicated that in yeast and cardiomyocytes the underlying mechanisms regulating the scale-free behavior are similar. We validated this finding in vivo using single cells, and attenuating the activity of the mitochondrial inner membrane anion channel with 4-chlorodiazepam to show that the oscillation of NAD(PH and reactive oxygen species (ROS can be abated in these two evolutionarily distant species. Taken together these data strongly support our hypothesis that the generation of ROS, coupled to redox cycling, driven by cytoplasmic and mitochondrial processes, are at the core of the observed rhythmicity and scale-free dynamics. We argue that the operation of scale-free bioenergetic dynamics plays a fundamental role to integrate cellular function, while providing a framework for robust, yet flexible, responses to the environment.

  15. Eukaryotic ribonucleases P/MRP: the crystal structure of the P3 domain.

    Science.gov (United States)

    Perederina, Anna; Esakova, Olga; Quan, Chao; Khanova, Elena; Krasilnikov, Andrey S

    2010-02-17

    Ribonuclease (RNase) P is a site-specific endoribonuclease found in all kingdoms of life. Typical RNase P consists of a catalytic RNA component and a protein moiety. In the eukaryotes, the RNase P lineage has split into two, giving rise to a closely related enzyme, RNase MRP, which has similar components but has evolved to have different specificities. The eukaryotic RNases P/MRP have acquired an essential helix-loop-helix protein-binding RNA domain P3 that has an important function in eukaryotic enzymes and distinguishes them from bacterial and archaeal RNases P. Here, we present a crystal structure of the P3 RNA domain from Saccharomyces cerevisiae RNase MRP in a complex with RNase P/MRP proteins Pop6 and Pop7 solved to 2.7 A. The structure suggests similar structural organization of the P3 RNA domains in RNases P/MRP and possible functions of the P3 domains and proteins bound to them in the stabilization of the holoenzymes' structures as well as in interactions with substrates. It provides the first insight into the structural organization of the eukaryotic enzymes of the RNase P/MRP family.

  16. Mathematical model of reproductive death of irradiated eukaryotic cells, which considers saturation of DNA reparation system

    International Nuclear Information System (INIS)

    Knyigavko, V.G.; Ponomarenko, N.S.; Meshcheryakova, O.P.; Protasenya, S.Yu.

    2009-01-01

    A mathematical model of the processes determining reproductive death of the exposed cells was built. The model takes into account the phenomenon of saturation of the system of DNA radiation lesion reparation and structural functional peculiarities of chromatin structure in eukaryotes. The problem of assessment of the model parameters using experimental data was discussed.

  17. Avian leukosis virus is a versatile eukaryotic platform for polypeptide display

    International Nuclear Information System (INIS)

    Khare, Pranay D.; Russell, Stephen J.; Federspiel, Mark J.

    2003-01-01

    Display technology refers to methods of generating libraries of modularly coded biomolecules and screening them for particular properties. Retroviruses are good candidates to be a eukaryotic viral platform for the display of polypeptides synthesized in eukaryotic cells. Here we demonstrate that avian leukosis virus (ALV) provides an ideal platform for display of nonviral polyaeptides expressed in a eukaryotic cell substrate. Different sizes of polypeptides were genetically fused to the extreme N-terminus of the ALV envelope glycoprotein in an ALV infectious clone containing an alkaline phosphatase reporter gene. The chimeric envelope glycoproteins were efficiently incorporated into virions and were stably displayed on the surface of the virions through multiple virus replication cycles. The foreign polypeptides did not interfere with the attachment and entry functions of the underlying ALV envelope glycoproteins. The displayed polypeptides were fully functional and could efficiently mediate attachment of the recombinant viruses to their respective cognate receptors. This study demonstrates that ALV is an ideal display platform for the generation and selection of libraries of polypeptides where there is a need for expression, folding, and posttranslational modification in the endoplasmic reticulum of eukaryotic cells

  18. Distribution and Diversity of Microbial Eukaryotes in Bathypelagic Waters of the South China Sea.

    Science.gov (United States)

    Xu, Dapeng; Jiao, Nianzhi; Ren, Rui; Warren, Alan

    2017-05-01

    Little is known about the biodiversity of microbial eukaryotes in the South China Sea, especially in waters at bathyal depths. Here, we employed SSU rDNA gene sequencing to reveal the diversity and community structure across depth and distance gradients in the South China Sea. Vertically, the highest alpha diversity was found at 75-m depth. The communities of microbial eukaryotes were clustered into shallow-, middle-, and deep-water groups according to the depth from which they were collected, indicating a depth-related diversity and distribution pattern. Rhizaria sequences dominated the microeukaryote community and occurred in all samples except those from less than 50-m deep, being most abundant near the sea floor where they contributed ca. 64-97% and 40-74% of the total sequences and OTUs recovered, respectively. A large portion of rhizarian OTUs has neither a nearest named neighbor nor a nearest neighbor in the GenBank database which indicated the presence of new phylotypes in the South China Sea. Given their overwhelming abundance and richness, further phylogenetic analysis of rhizarians were performed and three new genetic clusters were revealed containing sequences retrieved from the deep waters of the South China Sea. Our results shed light on the diversity and community structure of microbial eukaryotes in this not yet fully explored area. © 2016 The Author(s) Journal of Eukaryotic Microbiology © 2016 International Society of Protistologists.

  19. EuMicroSatdb: A database for microsatellites in the sequenced genomes of eukaryotes

    Directory of Open Access Journals (Sweden)

    Grover Atul

    2007-07-01

    Full Text Available Abstract Background Microsatellites have immense utility as molecular markers in different fields like genome characterization and mapping, phylogeny and evolutionary biology. Existing microsatellite databases are of limited utility for experimental and computational biologists with regard to their content and information output. EuMicroSatdb (Eukaryotic MicroSatellite database http://ipu.ac.in/usbt/EuMicroSatdb.htm is a web based relational database for easy and efficient positional mining of microsatellites from sequenced eukaryotic genomes. Description A user friendly web interface has been developed for microsatellite data retrieval using Active Server Pages (ASP. The backend database codes for data extraction and assembly have been written using Perl based scripts and C++. Precise need based microsatellites data retrieval is possible using different input parameters like microsatellite type (simple perfect or compound perfect, repeat unit length (mono- to hexa-nucleotide, repeat number, microsatellite length and chromosomal location in the genome. Furthermore, information about clustering of different microsatellites in the genome can also be retrieved. Finally, to facilitate primer designing for PCR amplification of any desired microsatellite locus, 200 bp upstream and downstream sequences are provided. Conclusion The database allows easy systematic retrieval of comprehensive information about simple and compound microsatellites, microsatellite clusters and their locus coordinates in 31 sequenced eukaryotic genomes. The information content of the database is useful in different areas of research like gene tagging, genome mapping, population genetics, germplasm characterization and in understanding microsatellite dynamics in eukaryotic genomes.

  20. Horizontal gene transfer of an entire metabolic pathway between a eukaryotic alga and its DNA virus

    Science.gov (United States)

    Monier, Adam; Pagarete, António; de Vargas, Colomban; Allen, Michael J.; Read, Betsy; Claverie, Jean-Michel; Ogata, Hiroyuki

    2009-01-01

    Interactions between viruses and phytoplankton, the main primary producers in the oceans, affect global biogeochemical cycles and climate. Recent studies are increasingly revealing possible cases of gene transfers between cyanobacteria and phages, which might have played significant roles in the evolution of cyanobacteria/phage systems. However, little has been documented about the occurrence of horizontal gene transfer in eukaryotic phytoplankton/virus systems. Here we report phylogenetic evidence for the transfer of seven genes involved in the sphingolipid biosynthesis pathway between the cosmopolitan eukaryotic microalga Emiliania huxleyi and its large DNA virus EhV. PCR assays indicate that these genes are prevalent in E. huxleyi and EhV strains isolated from different geographic locations. Patterns of protein and gene sequence conservation support that these genes are functional in both E. huxleyi and EhV. This is the first clear case of horizontal gene transfer of multiple functionally linked enzymes in a eukaryotic phytoplankton–virus system. We examine arguments for the possible direction of the gene transfer. The virus-to-host direction suggests the existence of ancient viruses that controlled the complex metabolic pathway in order to infect primitive eukaryotic cells. In contrast, the host-to-virus direction suggests that the serial acquisition of genes involved in the same metabolic pathway might have been a strategy for the ancestor of EhVs to stay ahead of their closest relatives in the great evolutionary race for survival. PMID:19451591

  1. Eukaryotic resistance to fluoride toxicity mediated by a widespread family of fluoride export proteins.

    Science.gov (United States)

    Li, Sanshu; Smith, Kathryn D; Davis, Jared H; Gordon, Patricia B; Breaker, Ronald R; Strobel, Scott A

    2013-11-19

    Fluorine is an abundant element and is toxic to organisms from bacteria to humans, but the mechanisms by which eukaryotes resist fluoride toxicity are unknown. The Escherichia coli gene crcB was recently shown to be regulated by a fluoride-responsive riboswitch, implicating it in fluoride response. There are >8,000 crcB homologs across all domains of life, indicating that it has an important role in biology. Here we demonstrate that eukaryotic homologs [renamed FEX (fluoride exporter)] function in fluoride export. FEX KOs in three eukaryotic model organisms, Neurospora crassa, Saccharomyces cerevisiae, and Candida albicans, are highly sensitized to fluoride (>200-fold) but not to other halides. Some of these KO strains are unable to grow in fluoride concentrations found in tap water. Using the radioactive isotope of fluoride, (18)F, we developed an assay to measure the intracellular fluoride concentration and show that the FEX deletion strains accumulate fluoride in excess of the external concentration, providing direct evidence of FEX function in fluoride efflux. In addition, they are more sensitive to lower pH in the presence of fluoride. These results demonstrate that eukaryotic FEX genes encode a previously unrecognized class of fluoride exporter necessary for survival in standard environmental conditions.

  2. Genome-wide mapping reveals single-origin chromosome replication in Leishmania, a eukaryotic microbe.

    Science.gov (United States)

    Marques, Catarina A; Dickens, Nicholas J; Paape, Daniel; Campbell, Samantha J; McCulloch, Richard

    2015-10-19

    DNA replication initiates on defined genome sites, termed origins. Origin usage appears to follow common rules in the eukaryotic organisms examined to date: all chromosomes are replicated from multiple origins, which display variations in firing efficiency and are selected from a larger pool of potential origins. To ask if these features of DNA replication are true of all eukaryotes, we describe genome-wide origin mapping in the parasite Leishmania. Origin mapping in Leishmania suggests a striking divergence in origin usage relative to characterized eukaryotes, since each chromosome appears to be replicated from a single origin. By comparing two species of Leishmania, we find evidence that such origin singularity is maintained in the face of chromosome fusion or fission events during evolution. Mapping Leishmania origins suggests that all origins fire with equal efficiency, and that the genomic sites occupied by origins differ from related non-origins sites. Finally, we provide evidence that origin location in Leishmania displays striking conservation with Trypanosoma brucei, despite the latter parasite replicating its chromosomes from multiple, variable strength origins. The demonstration of chromosome replication for a single origin in Leishmania, a microbial eukaryote, has implications for the evolution of origin multiplicity and associated controls, and may explain the pervasive aneuploidy that characterizes Leishmania chromosome architecture.

  3. Selfish operons: the evolutionary impact of gene clustering in prokaryotes and eukaryotes.

    Science.gov (United States)

    Lawrence, J

    1999-12-01

    The Selfish Operon Model postulates that the organization of bacterial genes into operons is beneficial to the constituent genes in that proximity allows horizontal cotransfer of all genes required for a selectable phenotype; eukaryotic operons formed for very different reasons. Horizontal transfer of selfish operons most probably promotes bacterial diversification.

  4. Are maternal mitochondria the selfish entities that are masters of the cells of eukaryotic multicellular organisms?

    Science.gov (United States)

    Barlow, Peter W; Baldelli, E; Baluška, Frantisek

    2009-01-01

    The Energide concept, as well as the endosymbiotic theory of eukaryotic cell organization and evolution, proposes that present-day cells of eukaryotic organisms are mosaics of specialized and cooperating units, or organelles. Some of these units were originally free-living prokaryotes, which were engulfed during evolutionary time. Mitochondria represent one of these types of previously independent organisms, the Energide, is another type. This new perspective on the organization of the cell has been further expanded to reveal the concept of a public milieu, the cytosol, in which Energides and mitochondria live, each with their own private internal milieu. The present paper discusses how the endosymbiotic theory implicates a new hypothesis about the hierarchical and communicational organization of the integrated prokaryotic components of the eukaryotic cell and provides a new angle from which to consider the theory of evolution and its bearing upon cellular complexity. Thus, it is proposed that the “selfish gene” hypothesis of Dawkins1 is not the only possible perspective for comprehending genomic and cellular evolution. Our proposal is that maternal mitochondria are the selfish “master” entities of the eukaryotic cell with respect not only to their propagation from cell-to-cell and from generation-to-generation but also to their regulation of all other cellular functions. However, it should be recognized that the concept of “master” and “servant” cell components is a metaphor; in present-day living organisms their organellar components are considered to be interdependent and inseparable. PMID:19513277

  5. Neural Network Prediction of Translation Initiation Sites in Eukaryotes: Perspectives for EST and Genome analysis

    DEFF Research Database (Denmark)

    Pedersen, Anders Gorm; Nielsen, Henrik

    1997-01-01

    Translation in eukaryotes does not always start at the first AUG in an mRNA, implying that context information also plays a role.This makes prediction of translation initiation sites a non-trivial task, especially when analysing EST and genome data where the entire mature mRNA sequence is not known...

  6. Lateral gene transfer between prokaryotes and multicellular eukaryotes: ongoing and significant?

    NARCIS (Netherlands)

    Ros, V.I.D.; Hurst, G.D.D.

    2009-01-01

    The expansion of genome sequencing projects has produced accumulating evidence for lateral transfer of genes between prokaryotic and eukaryotic genomes. However, it remains controversial whether these genes are of functional importance in their recipient host. Nikoh and Nakabachi, in a recent paper

  7. Identification of prokaryotic and eukaryotic signal peptides and prediction of their cleavage sites

    DEFF Research Database (Denmark)

    Nielsen, Henrik; Engelbrecht, Jacob; Brunak, Søren

    1997-01-01

    We have developed a new method for the identification of signal peptides and their cleavage based on neural networks trained on separate sets of prokaryotic and eukaryotic sequence. The method performs significantly better than previous prediction schemes and can easily be applied on genome...

  8. Sex is a ubiquitous, ancient, and inherent attribute of eukaryotic life

    Czech Academy of Sciences Publication Activity Database

    Speijer, D.; Lukeš, Julius; Eliáš, M.

    2015-01-01

    Roč. 112, č. 29 (2015), s. 8827-8834 ISSN 0027-8424 R&D Projects: GA MŠk LH12104; GA ČR GA15-21974S Institutional support: RVO:60077344 Keywords : reactive oxygen species * evolution * protists * eukaryotes * sex Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 9.423, year: 2015

  9. A versatile selection system for folding competent proteins using genetic complementation in a eukaryotic host

    DEFF Research Database (Denmark)

    Lyngsø, C.; Kjaerulff, S.; Muller, S.

    2010-01-01

    in vivo selection system for folded proteins. It is based on genetic complementation of the Schizosaccharomyces pombe growth marker gene invertase fused C-terminally to a protein library. The fusion proteins are directed to the secretion system, utilizing the ability of the eukaryotic protein quality...

  10. The New Higher Level Classification of Eukaryotes with Emphasis on the Taxonomy of Protists

    Science.gov (United States)

    SINA M. ADL; ALASTAIR G. B. SIMPSON; MARK A. FARMER; ROBERT A. ANDERSEN; O. ROGER ANDERSON; JOHN R. BARTA; SAMUEL S. BOWSER; GUY BRUGEROLLE; ROBERT A. FENSOME; SUZANNE FREDERICQ; TIMOTHY Y. JAMES; SERGEI KARPOV; PAUL KUGRENS; JOHN KRUG; CHRISTOPHER E. LANE; LOUISE A. LEWIS; JEAN LODGE; DENIS H. LYNN; DAVID G. MANN; RICHARD M. MCCOURT; LEONEL MENDOZA; ØJVIND MOESTRUP; SHARON E. MOZLEY-STANDRIDGE; THOMAS A. NERAD; CAROL A. SHEARER; ALEXEY V. SMIRNOV; FREDERICK W. SPIEGEL; MAX F.J.R. TAYLOR

    2005-01-01

    This revision of the classification of unicellular eukaryotes updates that of Levine et al. (1980) for the protozoa and expands it to include other protists. Whereas the previous revision was primarily to incorporate the results of ultrastructural studies, this revision incorporates results from both ultrastructural research since 1980 and molecular phylogenetic...

  11. The new higher level classification of eukaryotes with emphasis on the taxonomy of protists

    Science.gov (United States)

    Sina M. Adl; Alastair G.B. Simpson; Mark A. Farmer; Robert A. Andersen; O. Roger Anderson; John R. Barta; Samuel S. Bowser; Guy Brugerolle; Robert A. Fensome; Suzanne Fredericq; Timothy Y. James; Sergei Karpov; Paul Kugrens; John Krug; Christopher E. Lane; Louise A. Lewis; Jean Lodge; Denis H. Lynn; David G. Mann; Richard M. McCourt; Leonel Mendoza; Ojvind Moestrup; Sharon E. Mozley-Standridge; Thomas A. Nerad; Carol A. Shearer; Alexey V. Smirnov; Frederick W. Speigel; Max F.J.R. Taylor

    2005-01-01

    This revision of the classification of unicellular eukaryotes updates that of Levine et al. (1980) for the protozoa and expands it to include other protists. Whereas the previous revision was primarily to incorporate the results of ultrastructural studies, this revision incorporates results from both ultrastructural research since 1980 and molecular phylogenetic...

  12. Short proofs of strong normalization

    OpenAIRE

    Wojdyga, Aleksander

    2008-01-01

    This paper presents simple, syntactic strong normalization proofs for the simply-typed lambda-calculus and the polymorphic lambda-calculus (system F) with the full set of logical connectives, and all the permutative reductions. The normalization proofs use translations of terms and types to systems, for which strong normalization property is known.

  13. Strong-back safety latch

    International Nuclear Information System (INIS)

    DeSantis, G.N.

    1995-01-01

    The calculation decides the integrity of the safety latch that will hold the strong-back to the pump during lifting. The safety latch will be welded to the strong-back and will latch to a 1.5-in. dia cantilever rod welded to the pump baseplate. The static and dynamic analysis shows that the safety latch will hold the strong-back to the pump if the friction clamps fail and the pump become free from the strong-back. Thus, the safety latch will meet the requirements of the Lifting and Rigging Manual for under the hook lifting for static loading; it can withstand shock loads from the strong-back falling 0.25 inch

  14. Bacterial Signaling Nucleotides Inhibit Yeast Cell Growth by Impacting Mitochondrial and Other Specifically Eukaryotic Functions.

    Science.gov (United States)

    Hesketh, Andy; Vergnano, Marta; Wan, Chris; Oliver, Stephen G

    2017-07-25

    We have engineered Saccharomyces cerevisiae to inducibly synthesize the prokaryotic signaling nucleotides cyclic di-GMP (cdiGMP), cdiAMP, and ppGpp in order to characterize the range of effects these nucleotides exert on eukaryotic cell function during bacterial pathogenesis. Synthetic genetic array (SGA) and transcriptome analyses indicated that, while these compounds elicit some common reactions in yeast, there are also complex and distinctive responses to each of the three nucleotides. All three are capable of inhibiting eukaryotic cell growth, with the guanine nucleotides exhibiting stronger effects than cdiAMP. Mutations compromising mitochondrial function and chromatin remodeling show negative epistatic interactions with all three nucleotides. In contrast, certain mutations that cause defects in chromatin modification and ribosomal protein function show positive epistasis, alleviating growth inhibition by at least two of the three nucleotides. Uniquely, cdiGMP is lethal both to cells growing by respiration on acetate and to obligately fermentative petite mutants. cdiGMP is also synthetically lethal with the ribonucleotide reductase (RNR) inhibitor hydroxyurea. Heterologous expression of the human ppGpp hydrolase Mesh1p prevented the accumulation of ppGpp in the engineered yeast and restored cell growth. Extensive in vivo interactions between bacterial signaling molecules and eukaryotic gene function occur, resulting in outcomes ranging from growth inhibition to death. cdiGMP functions through a mechanism that must be compensated by unhindered RNR activity or by functionally competent mitochondria. Mesh1p may be required for abrogating the damaging effects of ppGpp in human cells subjected to bacterial infection. IMPORTANCE During infections, pathogenic bacteria can release nucleotides into the cells of their eukaryotic hosts. These nucleotides are recognized as signals that contribute to the initiation of defensive immune responses that help the infected

  15. Molecular detection of eukaryotes in a single human stool sample from Senegal.

    Directory of Open Access Journals (Sweden)

    Ibrahim Hamad

    Full Text Available BACKGROUND: Microbial eukaryotes represent an important component of the human gut microbiome, with different beneficial or harmful roles; some species are commensal or mutualistic, whereas others are opportunistic or parasitic. The diversity of eukaryotes inhabiting humans remains relatively unexplored because of either the low abundance of these organisms in human gut or because they have received limited attention from a whole-community perspective. METHODOLOGY/PRINCIPAL FINDING: In this study, a single fecal sample from a healthy African male was studied using both culture-dependent methods and extended molecular methods targeting the 18S rRNA and ITS sequences. Our results revealed that very few fungi, including Candida spp., Galactomyces spp., and Trichosporon asahii, could be isolated using culture-based methods. In contrast, a relatively a high number of eukaryotic species could be identified in this fecal sample when culture-independent methods based on various primer sets were used. A total of 27 species from one sample were found among the 977 analyzed clones. The clone libraries were dominated by fungi (716 clones/977, 73.3%, corresponding to 16 different species. In addition, 187 sequences out of 977 (19.2% corresponded to 9 different species of plants; 59 sequences (6% belonged to other micro-eukaryotes in the gut, including Entamoeba hartmanni and Blastocystis sp; and only 15 clones/977 (1.5% were related to human 18S rRNA sequences. CONCLUSION: Our results revealed a complex eukaryotic community in the volunteer's gut, with fungi being the most abundant species in the stool sample. Larger investigations are needed to assess the generality of these results and to understand their roles in human health and disease.

  16. Critical analysis of eukaryotic phylogeny: a case study based on the HSP70 family.

    Science.gov (United States)

    Germot, A; Philippe, H

    1999-01-01

    Trichomonads, together with diplomonads and microsporidia, emerge at the base of the eukaryotic tree, on the basis of the small subunit rRNA phylogeny. However, phylogenies based on protein sequences such as tubulin are markedly different with these protists emerging much later. We have investigated 70 kDa heat-shock protein (HSP70), which could be a reliable phylogenetic marker. In eukaryotes, HSP70s are found in cytosol, endoplasmic reticulum, and organelles (mitochondria and chloroplasts). In Trichomonas vaginalis we identified nine different HSP70-encoding genes and sequenced three nearly complete cDNAs corresponding to cytosolic, endoplasmic reticulum, and mitochondrial-type HSP70. Phylogenies of eukaryotes were reconstructed using the classical methods while varying the number of species and characters considered. Almost all the undoubtedly monophyletic groups, defined by ultrastructural characters, were recovered. However, due to the long branch attraction phenomenon, the evolutionary rates were the main factor determining the position of species, even with the use of a close outgroup, which is an important advantage of HSP70 with respect to many other markers. Numerous variable sites are peculiar to Trichomonas and probably generated the artefactual placement of this species at the base of the eukaryotes or as the sister group of fast-evolving species. The inter-phyla relationships were not well supported and were sensitive to the reconstruction method, the number of species; and the quantity of information used. This lack of resolution could be explained by the very rapid diversification of eukaryotes, likely after the mitochondrial endosymbiosis.

  17. Metabolic profiles of prokaryotic and eukaryotic communities in deep-sea sponge Neamphius huxleyi indicated by metagenomics

    Science.gov (United States)

    Li, Zhi-Yong; Wang, Yue-Zhu; He, Li-Ming; Zheng, Hua-Jun

    2014-01-01

    The whole metabolism of a sponge holobiont and the respective contributions of prokaryotic and eukaryotic symbionts and their associations with the sponge host remain largely unclear. Meanwhile, compared with shallow water sponges, deep-sea sponges are rarely understood. Here we report the metagenomic exploration of deep-sea sponge Neamphius huxleyi at the whole community level. Metagenomic data showed phylogenetically diverse prokaryotes and eukaryotes in Neamphius huxleyi. MEGAN and gene enrichment analyses indicated different metabolic potentials of prokaryotic symbionts from eukaryotic symbionts, especially in nitrogen and carbon metabolisms, and their molecular interactions with the sponge host. These results supported the hypothesis that prokaryotic and eukaryotic symbionts have different ecological roles and relationships with sponge host. Moreover, vigorous denitrification, and CO2 fixation by chemoautotrophic prokaryotes were suggested for this deep-sea sponge. The study provided novel insights into the respective potentials of prokaryotic and eukaryotic symbionts and their associations with deep-sea sponge Neamphius huxleyi. PMID:24463735

  18. Metabolic profiles of prokaryotic and eukaryotic communities in deep-sea sponge Neamphius huxleyi [corrected]. indicated by metagenomics.

    Science.gov (United States)

    Li, Zhi-Yong; Wang, Yue-Zhu; He, Li-Ming; Zheng, Hua-Jun

    2014-01-27

    The whole metabolism of a sponge holobiont and the respective contributions of prokaryotic and eukaryotic symbionts and their associations with the sponge host remain largely unclear. Meanwhile, compared with shallow water sponges, deep-sea sponges are rarely understood. Here we report the metagenomic exploration of deep-sea sponge Neamphius huxleyi [corrected] . at the whole community level. Metagenomic data showed phylogenetically diverse prokaryotes and eukaryotes in Neamphius huxleyi [corrected]. MEGAN and gene enrichment analyses indicated different metabolic potentials of prokaryotic symbionts from eukaryotic symbionts, especially in nitrogen and carbon metabolisms, and their molecular interactions with the sponge host. These results supported the hypothesis that prokaryotic and eukaryotic symbionts have different ecological roles and relationships with sponge host. Moreover, vigorous denitrification, and CO2 fixation by chemoautotrophic prokaryotes were suggested for this deep-sea sponge. The study provided novel insights into the respective potentials of prokaryotic and eukaryotic symbionts and their associations with deep-sea sponge Neamphius huxleyi [corrected].

  19. Metabolic profiles of prokaryotic and eukaryotic communities in deep-sea sponge Lamellomorpha sp. indicated by metagenomics

    Science.gov (United States)

    Li, Zhi-Yong; Wang, Yue-Zhu; He, Li-Ming; Zheng, Hua-Jun

    2014-01-01

    The whole metabolism of a sponge holobiont and the respective contributions of prokaryotic and eukaryotic symbionts and their associations with the sponge host remain largely unclear. Meanwhile, compared with shallow water sponges, deep-sea sponges are rarely understood. Here we report the metagenomic exploration of deep-sea sponge Lamellomorpha sp. at the whole community level. Metagenomic data showed phylogenetically diverse prokaryotes and eukaryotes in Lamellomorpha sp.. MEGAN and gene enrichment analyses indicated different metabolic potentials of prokaryotic symbionts from eukaryotic symbionts, especially in nitrogen and carbon metabolisms, and their molecular interactions with the sponge host. These results supported the hypothesis that prokaryotic and eukaryotic symbionts have different ecological roles and relationships with sponge host. Moreover, vigorous denitrification, and CO2 fixation by chemoautotrophic prokaryotes were suggested for this deep-sea sponge. The study provided novel insights into the respective potentials of prokaryotic and eukaryotic symbionts and their associations with deep-sea sponge Lamellomorpha sp..

  20. Lateral transfer of tetrahymanol-synthesizing genes has allowed multiple diverse eukaryote lineages to independently adapt to environments without oxygen

    Directory of Open Access Journals (Sweden)

    Takishita Kiyotaka

    2012-02-01

    Full Text Available Abstract Sterols are key components of eukaryotic cellular membranes that are synthesized by multi-enzyme pathways that require molecular oxygen. Because prokaryotes fundamentally lack sterols, it is unclear how the vast diversity of bacterivorous eukaryotes that inhabit hypoxic environments obtain, or synthesize, sterols. Here we show that tetrahymanol, a triterpenoid that does not require molecular oxygen for its biosynthesis, likely functions as a surrogate of sterol in eukaryotes inhabiting oxygen-poor environments. Genes encoding the tetrahymanol synthesizing enzyme squalene-tetrahymanol cyclase were found from several phylogenetically diverged eukaryotes that live in oxygen-poor environments and appear to have been laterally transferred among such eukaryotes. Reviewers This article was reviewed by Eric Bapteste and Eugene Koonin.

  1. Different roles of eukaryotic MutS and MutL complexes in repair of small insertion and deletion loops in yeast.

    Directory of Open Access Journals (Sweden)

    Nina V Romanova

    2013-10-01

    Full Text Available DNA mismatch repair greatly increases genome fidelity by recognizing and removing replication errors. In order to understand how this fidelity is maintained, it is important to uncover the relative specificities of the different components of mismatch repair. There are two major mispair recognition complexes in eukaryotes that are homologues of bacterial MutS proteins, MutSα and MutSβ, with MutSα recognizing base-base mismatches and small loop mispairs and MutSβ recognizing larger loop mispairs. Upon recognition of a mispair, the MutS complexes then interact with homologues of the bacterial MutL protein. Loops formed on the primer strand during replication lead to insertion mutations, whereas loops on the template strand lead to deletions. We show here in yeast, using oligonucleotide transformation, that MutSα has a strong bias toward repair of insertion loops, while MutSβ has an even stronger bias toward repair of deletion loops. Our results suggest that this bias in repair is due to the different interactions of the MutS complexes with the MutL complexes. Two mutants of MutLα, pms1-G882E and pms1-H888R, repair deletion mispairs but not insertion mispairs. Moreover, we find that a different MutL complex, MutLγ, is extremely important, but not sufficient, for deletion repair in the presence of either MutLα mutation. MutSβ is present in many eukaryotic organisms, but not in prokaryotes. We suggest that the biased repair of deletion mispairs may reflect a critical eukaryotic function of MutSβ in mismatch repair.

  2. Genome-wide analyses and functional classification of proline repeat-rich proteins: potential role of eIF5A in eukaryotic evolution.

    Directory of Open Access Journals (Sweden)

    Ajeet Mandal

    Full Text Available The eukaryotic translation factor, eIF5A has been recently reported as a sequence-specific elongation factor that facilitates peptide bond formation at consecutive prolines in Saccharomyces cerevisiae, as its ortholog elongation factor P (EF-P does in bacteria. We have searched the genome databases of 35 representative organisms from six kingdoms of life for PPP (Pro-Pro-Pro and/or PPG (Pro-Pro-Gly-encoding genes whose expression is expected to depend on eIF5A. We have made detailed analyses of proteome data of 5 selected species, Escherichia coli, Saccharomyces cerevisiae, Drosophila melanogaster, Mus musculus and Homo sapiens. The PPP and PPG motifs are low in the prokaryotic proteomes. However, their frequencies markedly increase with the biological complexity of eukaryotic organisms, and are higher in newly derived proteins than in those orthologous proteins commonly shared in all species. Ontology classifications of S. cerevisiae and human genes encoding the highest level of polyprolines reveal their strong association with several specific biological processes, including actin/cytoskeletal associated functions, RNA splicing/turnover, DNA binding/transcription and cell signaling. Previously reported phenotypic defects in actin polarity and mRNA decay of eIF5A mutant strains are consistent with the proposed role for eIF5A in the translation of the polyproline-containing proteins. Of all the amino acid tandem repeats (≥3 amino acids, only the proline repeat frequency correlates with functional complexity of the five organisms examined. Taken together, these findings suggest the importance of proline repeat-rich proteins and a potential role for eIF5A and its hypusine modification pathway in the course of eukaryotic evolution.

  3. Structural analyses of Legionella LepB reveal a new GAP fold that catalytically mimics eukaryotic RasGAP.

    Science.gov (United States)

    Yu, Qin; Hu, Liyan; Yao, Qing; Zhu, Yongqun; Dong, Na; Wang, Da-Cheng; Shao, Feng

    2013-06-01

    Rab GTPases are emerging targets of diverse bacterial pathogens. Here, we perform biochemical and structural analyses of LepB, a Rab GTPase-activating protein (GAP) effector from Legionella pneumophila. We map LepB GAP domain to residues 313-618 and show that the GAP domain is Rab1 specific with a catalytic activity higher than the canonical eukaryotic TBC GAP and the newly identified VirA/EspG family of bacterial RabGAP effectors. Exhaustive mutation analyses identify Arg444 as the arginine finger, but no catalytically essential glutamine residues. Crystal structures of LepB313-618 alone and the GAP domain of Legionella drancourtii LepB in complex with Rab1-GDP-AlF3 support the catalytic role of Arg444, and also further reveal a 3D architecture and a GTPase-binding mode distinct from all known GAPs. Glu449, structurally equivalent to TBC RabGAP glutamine finger in apo-LepB, undergoes a drastic movement upon Rab1 binding, which induces Rab1 Gln70 side-chain flipping towards GDP-AlF3 through a strong ionic interaction. This conformationally rearranged Gln70 acts as the catalytic cis-glutamine, therefore uncovering an unexpected RasGAP-like catalytic mechanism for LepB. Our studies highlight an extraordinary structural and catalytic diversity of RabGAPs, particularly those from bacterial pathogens.

  4. Bacteriophage T5 encodes a homolog of the eukaryotic transcription coactivator PC4 implicated in recombination-dependent DNA replication.

    Science.gov (United States)

    Steigemann, Birthe; Schulz, Annina; Werten, Sebastiaan

    2013-11-15

    The RNA polymerase II cofactor PC4 globally regulates transcription of protein-encoding genes through interactions with unwinding DNA, the basal transcription machinery and transcription activators. Here, we report the surprising identification of PC4 homologs in all sequenced representatives of the T5 family of bacteriophages, as well as in an archaeon and seven phyla of eubacteria. We have solved the crystal structure of the full-length T5 protein at 1.9Å, revealing a striking resemblance to the characteristic single-stranded DNA (ssDNA)-binding core domain of PC4. Intriguing novel structural features include a potential regulatory region at the N-terminus and a C-terminal extension of the homodimerisation interface. The genome organisation of T5-related bacteriophages points at involvement of the PC4 homolog in recombination-dependent DNA replication, strongly suggesting that the protein corresponds to the hitherto elusive replicative ssDNA-binding protein of the T5 family. Our findings imply that PC4-like factors intervene in multiple unwinding-related processes by acting as versatile modifiers of nucleic acid conformation and raise the possibility that the eukaryotic transcription coactivator derives from ancestral DNA replication, recombination and repair factors. © 2013.

  5. Strong coupling phase in QED

    International Nuclear Information System (INIS)

    Aoki, Ken-ichi

    1988-01-01

    Existence of a strong coupling phase in QED has been suggested in solutions of the Schwinger-Dyson equation and in Monte Carlo simulation of lattice QED. In this article we recapitulate the previous arguments, and formulate the problem in the modern framework of the renormalization theory, Wilsonian renormalization. This scheme of renormalization gives the best understanding of the basic structure of a field theory especially when it has a multi-phase structure. We resolve some misleading arguments in the previous literature. Then we set up a strategy to attack the strong phase, if any. We describe a trial; a coupled Schwinger-Dyson equation. Possible picture of the strong coupling phase QED is presented. (author)

  6. Guanine nucleotide binding to the Bateman domain mediates the allosteric inhibition of eukaryotic IMP dehydrogenases

    Science.gov (United States)

    Buey, Rubén M.; Ledesma-Amaro, Rodrigo; Velázquez-Campoy, Adrián; Balsera, Mónica; Chagoyen, Mónica; de Pereda, José M.; Revuelta, José L.

    2015-11-01

    Inosine-5'-monophosphate dehydrogenase (IMPDH) plays key roles in purine nucleotide metabolism and cell proliferation. Although IMPDH is a widely studied therapeutic target, there is limited information about its physiological regulation. Using Ashbya gossypii as a model, we describe the molecular mechanism and the structural basis for the allosteric regulation of IMPDH by guanine nucleotides. We report that GTP and GDP bind to the regulatory Bateman domain, inducing octamers with compromised catalytic activity. Our data suggest that eukaryotic and prokaryotic IMPDHs might have developed different regulatory mechanisms, with GTP/GDP inhibiting only eukaryotic IMPDHs. Interestingly, mutations associated with human retinopathies map into the guanine nucleotide-binding sites including a previously undescribed non-canonical site and disrupt allosteric inhibition. Together, our results shed light on the mechanisms of the allosteric regulation of enzymes mediated by Bateman domains and provide a molecular basis for certain retinopathies, opening the door to new therapeutic approaches.

  7. Reassigning stop codons via translation termination: How a few eukaryotes broke the dogma.

    Science.gov (United States)

    Alkalaeva, Elena; Mikhailova, Tatiana

    2017-03-01

    The genetic code determines how amino acids are encoded within mRNA. It is universal among the vast majority of organisms, although several exceptions are known. Variant genetic codes are found in ciliates, mitochondria, and numerous other organisms. All revealed genetic codes (standard and variant) have at least one codon encoding a translation stop signal. However, recently two new genetic codes with a reassignment of all three stop codons were revealed in studies examining the protozoa transcriptomes. Here, we discuss this finding and the recent studies of variant genetic codes in eukaryotes. We consider the possible molecular mechanisms allowing the use of certain codons as sense and stop signals simultaneously. The results obtained by studying these amazing organisms represent a new and exciting insight into the mechanism of stop codon decoding in eukaryotes. Also see the video abstract here. © 2017 WILEY Periodicals, Inc.

  8. Three-dimensional structural analysis of eukaryotic flagella/cilia by electron cryo-tomography

    International Nuclear Information System (INIS)

    Bui, Khanh Huy; Pigino, Gaia; Ishikawa, Takashi

    2011-01-01

    Based on the molecular architecture revealed by electron cryo-tomography, the mechanism of the bending motion of eukaryotic flagella/cilia is discussed. Electron cryo-tomography is a potential approach to analyzing the three-dimensional conformation of frozen hydrated biological macromolecules using electron microscopy. Since projections of each individual object illuminated from different orientations are merged, electron tomography is capable of structural analysis of such heterogeneous environments as in vivo or with polymorphism, although radiation damage and the missing wedge are severe problems. Here, recent results on the structure of eukaryotic flagella, which is an ATP-driven bending organelle, from green algae Chlamydomonas are presented. Tomographic analysis reveals asymmetric molecular arrangements, especially that of the dynein motor proteins, in flagella, giving insight into the mechanism of planar asymmetric bending motion. Methodological challenges to obtaining higher-resolution structures from this technique are also discussed

  9. Production of isotopically labeled heterologous proteins in non-E. coli prokaryotic and eukaryotic cells

    International Nuclear Information System (INIS)

    Takahashi, Hideo; Shimada, Ichio

    2010-01-01

    The preparation of stable isotope-labeled proteins is necessary for the application of a wide variety of NMR methods, to study the structures and dynamics of proteins and protein complexes. The E. coli expression system is generally used for the production of isotope-labeled proteins, because of the advantages of ease of handling, rapid growth, high-level protein production, and low cost for isotope-labeling. However, many eukaryotic proteins are not functionally expressed in E. coli, due to problems related to disulfide bond formation, post-translational modifications, and folding. In such cases, other expression systems are required for producing proteins for biomolecular NMR analyses. In this paper, we review the recent advances in expression systems for isotopically labeled heterologous proteins, utilizing non-E. coli prokaryotic and eukaryotic cells.

  10. Incoherent feedforward control governs adaptation of activated ras in a eukaryotic chemotaxis pathway.

    Science.gov (United States)

    Takeda, Kosuke; Shao, Danying; Adler, Micha; Charest, Pascale G; Loomis, William F; Levine, Herbert; Groisman, Alex; Rappel, Wouter-Jan; Firtel, Richard A

    2012-01-03

    Adaptation in signaling systems, during which the output returns to a fixed baseline after a change in the input, often involves negative feedback loops and plays a crucial role in eukaryotic chemotaxis. We determined the dynamical response to a uniform change in chemoattractant concentration of a eukaryotic chemotaxis pathway immediately downstream from G protein-coupled receptors. The response of an activated Ras showed near-perfect adaptation, leading us to attempt to fit the results using mathematical models for the two possible simple network topologies that can provide perfect adaptation. Only the incoherent feedforward network accurately described the experimental results. This analysis revealed that adaptation in this Ras pathway is achieved through the proportional activation of upstream components and not through negative feedback loops. Furthermore, these results are consistent with a local excitation, global inhibition mechanism for gradient sensing, possibly with a Ras guanosine triphosphatase-activating protein acting as a global inhibitor.

  11. GFFview: A Web Server for Parsing and Visualizing Annotation Information of Eukaryotic Genome.

    Science.gov (United States)

    Deng, Feilong; Chen, Shi-Yi; Wu, Zhou-Lin; Hu, Yongsong; Jia, Xianbo; Lai, Song-Jia

    2017-10-01

    Owing to wide application of RNA sequencing (RNA-seq) technology, more and more eukaryotic genomes have been extensively annotated, such as the gene structure, alternative splicing, and noncoding loci. Annotation information of genome is prevalently stored as plain text in General Feature Format (GFF), which could be hundreds or thousands Mb in size. Therefore, it is a challenge for manipulating GFF file for biologists who have no bioinformatic skill. In this study, we provide a web server (GFFview) for parsing the annotation information of eukaryotic genome and then generating statistical description of six indices for visualization. GFFview is very useful for investigating quality and difference of the de novo assembled transcriptome in RNA-seq studies.

  12. The Big Bang of picorna-like virus evolution antedates the radiation of eukaryotic supergroups.

    Science.gov (United States)

    Koonin, Eugene V; Wolf, Yuri I; Nagasaki, Keizo; Dolja, Valerian V

    2008-12-01

    The recent discovery of RNA viruses in diverse unicellular eukaryotes and developments in evolutionary genomics have provided the means for addressing the origin of eukaryotic RNA viruses. The phylogenetic analyses of RNA polymerases and helicases presented in this Analysis article reveal close evolutionary relationships between RNA viruses infecting hosts from the Chromalveolate and Excavate supergroups and distinct families of picorna-like viruses of plants and animals. Thus, diversification of picorna-like viruses probably occurred in a 'Big Bang' concomitant with key events of eukaryogenesis. The origins of the conserved genes of picorna-like viruses are traced to likely ancestors including bacterial group II retroelements, the family of HtrA proteases and DNA bacteriophages.

  13. Deciphering DNA replication dynamics in eukaryotic cell populations in relation with their averaged chromatin conformations

    Science.gov (United States)

    Goldar, A.; Arneodo, A.; Audit, B.; Argoul, F.; Rappailles, A.; Guilbaud, G.; Petryk, N.; Kahli, M.; Hyrien, O.

    2016-03-01

    We propose a non-local model of DNA replication that takes into account the observed uncertainty on the position and time of replication initiation in eukaryote cell populations. By picturing replication initiation as a two-state system and considering all possible transition configurations, and by taking into account the chromatin’s fractal dimension, we derive an analytical expression for the rate of replication initiation. This model predicts with no free parameter the temporal profiles of initiation rate, replication fork density and fraction of replicated DNA, in quantitative agreement with corresponding experimental data from both S. cerevisiae and human cells and provides a quantitative estimate of initiation site redundancy. This study shows that, to a large extent, the program that regulates the dynamics of eukaryotic DNA replication is a collective phenomenon that emerges from the stochastic nature of replication origins initiation.

  14. The Persistent Contributions of RNA to Eukaryotic Gen(om)e Architecture and Cellular Function

    Science.gov (United States)

    Brosius, Jürgen

    2014-01-01

    Currently, the best scenario for earliest forms of life is based on RNA molecules as they have the proven ability to catalyze enzymatic reactions and harbor genetic information. Evolutionary principles valid today become apparent in such models already. Furthermore, many features of eukaryotic genome architecture might have their origins in an RNA or RNA/protein (RNP) world, including the onset of a further transition, when DNA replaced RNA as the genetic bookkeeper of the cell. Chromosome maintenance, splicing, and regulatory function via RNA may be deeply rooted in the RNA/RNP worlds. Mostly in eukaryotes, conversion from RNA to DNA is still ongoing, which greatly impacts the plasticity of extant genomes. Raw material for novel genes encoding protein or RNA, or parts of genes including regulatory elements that selection can act on, continues to enter the evolutionary lottery. PMID:25081515

  15. The eukaryotic genome is structurally and functionally more like a social insect colony than a book.

    Science.gov (United States)

    Qiu, Guo-Hua; Yang, Xiaoyan; Zheng, Xintian; Huang, Cuiqin

    2017-11-01

    Traditionally, the genome has been described as the 'book of life'. However, the metaphor of a book may not reflect the dynamic nature of the structure and function of the genome. In the eukaryotic genome, the number of centrally located protein-coding sequences is relatively constant across species, but the amount of noncoding DNA increases considerably with the increase of organismal evolutional complexity. Therefore, it has been hypothesized that the abundant peripheral noncoding DNA protects the genome and the central protein-coding sequences in the eukaryotic genome. Upon comparison with the habitation, sociality and defense mechanisms of a social insect colony, it is found that the genome is similar to a social insect colony in various aspects. A social insect colony may thus be a better metaphor than a book to describe the spatial organization and physical functions of the genome. The potential implications of the metaphor are also discussed.

  16. Strong interactions at high energy

    International Nuclear Information System (INIS)

    Anselmino, M.

    1995-01-01

    Spin effects in strong interaction high energy processes are subtle phenomena which involve both short and long distance physics and test perturbative and non perturbative aspects of QCD. Moreover, depending on quantities like interferences between different amplitudes and relative phases, spin observables always test a theory at a fundamental quantum mechanical level; it is then no surprise that spin data are often difficult to accommodate within the existing models. A report is made on the main issues and contributions discussed in the parallel Session on the open-quote open-quote Strong interactions at high energy close-quote close-quote in this Conference. copyright 1995 American Institute of Physics

  17. Strong-field dissociation dynamics

    International Nuclear Information System (INIS)

    DiMauro, L.F.; Yang, Baorui.

    1993-01-01

    The strong-field dissociation behavior of diatomic molecules is examined under two distinctive physical scenarios. In the first scenario, the dissociation of the isolated hydrogen and deuterium molecular ions is discussed. The dynamics of above-threshold dissociation (ATD) are investigated over a wide range of green and infrared intensities and compared to a dressed-state model. The second situation arises when strong-field neutral dissociation is followed by ionization of the atomic fragments. The study results in a direct measure of the atomic fragment's ac-Stark shift by observing the intensity-dependent shifts in the electron or nuclear fragment kinetic energy. 8 figs., 14 refs

  18. A Polycomb complex remains bound through DNA replication in the absence of other eukaryotic proteins

    KAUST Repository

    Lengsfeld, Bettina M.; Berry, Kayla N.; Ghosh, Sharmistha; Takahashi, Masateru; Francis, Nicole J.

    2012-01-01

    Propagation of chromatin states through DNA replication is central to epigenetic regulation and can involve recruitment of chromatin proteins to replicating chromatin through interactions with replication fork components. Here we show using a fully reconstituted T7 bacteriophage system that eukaryotic proteins are not required to tether the Polycomb complex PRC1 to templates during DNA replication. Instead, DNA binding by PRC1 can withstand passage of a simple replication fork.

  19. Inhibition of Ribosome Recruitment Induces Stress Granule Formation Independently of Eukaryotic Initiation Factor 2α Phosphorylation

    OpenAIRE

    Mazroui, Rachid; Sukarieh, Rami; Bordeleau, Marie-Eve; Kaufman, Randal J.; Northcote, Peter; Tanaka, Junichi; Gallouzi, Imed; Pelletier, Jerry

    2006-01-01

    Cytoplasmic aggregates known as stress granules (SGs) arise as a consequence of cellular stress and contain stalled translation preinitiation complexes. These foci are thought to serve as sites of mRNA storage or triage during the cell stress response. SG formation has been shown to require induction of eukaryotic initiation factor (eIF)2α phosphorylation. Herein, we investigate the potential role of other initiation factors in this process and demonstrate that interfering with eIF4A activity...

  20. Functional and evolutionary characterization of Ohr proteins in eukaryotes reveals many active homologs among pathogenic fungi

    Directory of Open Access Journals (Sweden)

    D.A. Meireles

    2017-08-01

    Full Text Available Ohr and OsmC proteins comprise two subfamilies within a large group of proteins that display Cys-based, thiol dependent peroxidase activity. These proteins were previously thought to be restricted to prokaryotes, but we show here, using iterated sequence searches, that Ohr/OsmC homologs are also present in 217 species of eukaryotes with a massive presence in Fungi (186 species. Many of these eukaryotic Ohr proteins possess an N-terminal extension that is predicted to target them to mitochondria. We obtained recombinant proteins for four eukaryotic members of the Ohr/OsmC family and three of them displayed lipoyl peroxidase activity. Further functional and biochemical characterization of the Ohr homologs from the ascomycete fungus Mycosphaerella fijiensis Mf_1 (MfOhr, the causative agent of Black Sigatoka disease in banana plants, was pursued. Similarly to what has been observed for the bacterial proteins, we found that: (i the peroxidase activity of MfOhr was supported by DTT or dihydrolipoamide (dithiols, but not by β-mercaptoethanol or GSH (monothiols, even in large excess; (ii MfOhr displayed preference for organic hydroperoxides (CuOOH and tBOOH over hydrogen peroxide; (iii MfOhr presented extraordinary reactivity towards linoleic acid hydroperoxides (k=3.18 (±2.13×108 M−1 s−1. Both Cys87 and Cys154 were essential to the peroxidase activity, since single mutants for each Cys residue presented no activity and no formation of intramolecular disulfide bond upon treatment with hydroperoxides. The pKa value of the Cysp residue was determined as 5.7±0.1 by a monobromobimane alkylation method. Therefore, eukaryotic Ohr peroxidases share several biochemical features with prokaryotic orthologues and are preferentially located in mitochondria. Keywords: Ohr/OsmC, Thiol-dependent peroxidases, Phylogeny

  1. Eukaryotic resistance to fluoride toxicity mediated by a widespread family of fluoride export proteins

    OpenAIRE

    Li, Sanshu; Smith, Kathryn D.; Davis, Jared H.; Gordon, Patricia B.; Breaker, Ronald R.; Strobel, Scott A.

    2013-01-01

    Although fluoride is plentiful in the environment and is commonly used at high concentrations in oral hygiene products, little has been known about how biological systems overcome the toxic effects of this anion. We demonstrate that a protein called FEX in many fungi is essential for cell survival in the presence of high fluoride concentrations. The protein is required for the rapid expulsion of cytoplasmic fluoride, indicating that many eukaryotic species that carry FEX genes likely avoid fl...

  2. Functional and evolutionary characterization of Ohr proteins in eukaryotes reveals many active homologs among pathogenic fungi.

    Science.gov (United States)

    Meireles, D A; Domingos, R M; Gaiarsa, J W; Ragnoni, E G; Bannitz-Fernandes, R; da Silva Neto, J F; de Souza, R F; Netto, L E S

    2017-08-01

    Ohr and OsmC proteins comprise two subfamilies within a large group of proteins that display Cys-based, thiol dependent peroxidase activity. These proteins were previously thought to be restricted to prokaryotes, but we show here, using iterated sequence searches, that Ohr/OsmC homologs are also present in 217 species of eukaryotes with a massive presence in Fungi (186 species). Many of these eukaryotic Ohr proteins possess an N-terminal extension that is predicted to target them to mitochondria. We obtained recombinant proteins for four eukaryotic members of the Ohr/OsmC family and three of them displayed lipoyl peroxidase activity. Further functional and biochemical characterization of the Ohr homologs from the ascomycete fungus Mycosphaerella fijiensis Mf_1 (MfOhr), the causative agent of Black Sigatoka disease in banana plants, was pursued. Similarly to what has been observed for the bacterial proteins, we found that: (i) the peroxidase activity of MfOhr was supported by DTT or dihydrolipoamide (dithiols), but not by β-mercaptoethanol or GSH (monothiols), even in large excess; (ii) MfOhr displayed preference for organic hydroperoxides (CuOOH and tBOOH) over hydrogen peroxide; (iii) MfOhr presented extraordinary reactivity towards linoleic acid hydroperoxides (k=3.18 (±2.13)×10 8 M -1 s -1 ). Both Cys 87 and Cys 154 were essential to the peroxidase activity, since single mutants for each Cys residue presented no activity and no formation of intramolecular disulfide bond upon treatment with hydroperoxides. The pK a value of the Cys p residue was determined as 5.7±0.1 by a monobromobimane alkylation method. Therefore, eukaryotic Ohr peroxidases share several biochemical features with prokaryotic orthologues and are preferentially located in mitochondria. Copyright © 2017. Published by Elsevier B.V.

  3. Expression of the lysostaphin gene of Staphylococcus simulans in a eukaryotic system.

    OpenAIRE

    Williamson, C M; Bramley, A J; Lax, A J

    1994-01-01

    The lysostaphin gene of Staphylococcus simulans was cloned into Escherichia coli. The 5' end of the gene was modified to include a eukaryotic start codon, the Kozak expression start site consensus sequence, and an enzyme site to facilitate manipulation of the gene. Transcription of the modified gene in vitro yielded an RNA transcript which, when added to a rabbit reticulocyte cell-free translation system, directed the synthesis of several products. The largest product, migrating at approximat...

  4. Molecular Dynamics Investigation of Cl− and Water Transport through a Eukaryotic CLC Transporter

    OpenAIRE

    Cheng, Mary Hongying; Coalson, Rob D.

    2012-01-01

    Early crystal structures of prokaryotic CLC proteins identified three Cl– binding sites: internal (Sint), central (Scen), and external (Sext). A conserved external GLU (GLUex) residue acts as a gate competing for Sext. Recently, the first crystal structure of a eukaryotic transporter, CmCLC, revealed that in this transporter GLUex competes instead for Scen. Here, we use molecular dynamics simulations to investigate Cl– transport through CmCLC. The gating and Cl–/H+ transport cycle are inferre...

  5. A Polycomb complex remains bound through DNA replication in the absence of other eukaryotic proteins

    KAUST Repository

    Lengsfeld, Bettina M.

    2012-09-17

    Propagation of chromatin states through DNA replication is central to epigenetic regulation and can involve recruitment of chromatin proteins to replicating chromatin through interactions with replication fork components. Here we show using a fully reconstituted T7 bacteriophage system that eukaryotic proteins are not required to tether the Polycomb complex PRC1 to templates during DNA replication. Instead, DNA binding by PRC1 can withstand passage of a simple replication fork.

  6. New insights into the structural organization of eukaryotic and prokaryotic cytoskeletons using cryo-electron tomography

    International Nuclear Information System (INIS)

    Kuerner, Julia; Medalia, Ohad; Linaroudis, Alexandros A.; Baumeister, Wolfgang

    2004-01-01

    Cryo-electron tomography (cryo-ET) is an emerging imaging technology that combines the potential of three-dimensional (3-D) imaging at molecular resolution (<5 nm) with a close-to-life preservation of the specimen. In conjunction with pattern recognition techniques, it enables us to map the molecular landscape inside cells. The application of cryo-ET to intact cells provides novel insights into the structure and the spatial organization of the cytoskeleton in prokaryotic and eukaryotic cells

  7. Microbial eukaryote diversity in the marine oxygen minimum zone off northern Chile

    OpenAIRE

    Parris, Darren J.; Ganesh, Sangita; Edgcomb, Virginia P.; Stewart, Frank J.; DeLong, Edward

    2014-01-01

    Molecular surveys are revealing diverse eukaryotic assemblages in oxygen-limited ocean waters. These communities may play pivotal ecological roles through autotrophy, feeding, and a wide range of symbiotic associations with prokaryotes. We used 18S rRNA gene sequencing to provide the first snapshot of pelagic microeukaryotic community structure in two cellular size fractions (0.2-1.6 µm, >1.6 µm) from seven depths through the anoxic oxygen minimum zone (OMZ) off northern Chile. Sequencing ...

  8. Six subgroups and extensive recent duplications characterize the evolution of the eukaryotic tubulin protein family.

    Science.gov (United States)

    Findeisen, Peggy; Mühlhausen, Stefanie; Dempewolf, Silke; Hertzog, Jonny; Zietlow, Alexander; Carlomagno, Teresa; Kollmar, Martin

    2014-08-27

    Tubulins belong to the most abundant proteins in eukaryotes providing the backbone for many cellular substructures like the mitotic and meiotic spindles, the intracellular cytoskeletal network, and the axonemes of cilia and flagella. Homologs have even been reported for archaea and bacteria. However, a taxonomically broad and whole-genome-based analysis of the tubulin protein family has never been performed, and thus, the number of subfamilies, their taxonomic distribution, and the exact grouping of the supposed archaeal and bacterial homologs are unknown. Here, we present the analysis of 3,524 tubulins from 504 species. The tubulins formed six major subfamilies, α to ζ. Species of all major kingdoms of the eukaryotes encode members of these subfamilies implying that they must have already been present in the last common eukaryotic ancestor. The proposed archaeal homologs grouped together with the bacterial TubZ proteins as sister clade to the FtsZ proteins indicating that tubulins are unique to eukaryotes. Most species contained α- and/or β-tubulin gene duplicates resulting from recent branch- and species-specific duplication events. This shows that tubulins cannot be used for constructing species phylogenies without resolving their ortholog-paralog relationships. The many gene duplicates and also the independent loss of the δ-, ε-, or ζ-tubulins, which have been shown to be part of the triplet microtubules in basal bodies, suggest that tubulins can functionally substitute each other. © The Author(s) 2014. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

  9. A study of eukaryotic response mechanisms to atmospheric pressure cold plasma by using Saccharomyces cerevisiae single gene mutants

    International Nuclear Information System (INIS)

    Feng Hongqing; Wang Ruixue; Sun Peng; Wu Haiyan; Liu Qi; Li Fangting; Fang Jing; Zhang Jue; Zhu Weidong

    2010-01-01

    The mechanisms of eukaryotic cell response to cold plasma are studied. A series of single gene mutants of eukaryotic model organism Saccharomyces cerevisiae are used to compare their sensitivity to plasma treatment with the wild type. We examined 12 mutants in the oxidative stress pathway and the cell cycle pathway, in which 8 are found to be hypersensitive to plasma processing. The mutated genes' roles in the two pathways are analyzed to understand the biological response mechanisms of plasma treatment. The results demonstrate that genes from both pathways are needed for the eukaryotic cells to survive the complex plasma treatment.

  10. Phylogenetic analysis of the core histone doublet and DNA topo II genes of Marseilleviridae: evidence of proto-eukaryotic provenance.

    Science.gov (United States)

    Erives, Albert J

    2017-11-28

    While the genomes of eukaryotes and Archaea both encode the histone-fold domain, only eukaryotes encode the core histone paralogs H2A, H2B, H3, and H4. With DNA, these core histones assemble into the nucleosomal octamer underlying eukaryotic chromatin. Importantly, core histones for H2A and H3 are maintained as neofunctionalized paralogs adapted for general bulk chromatin (canonical H2 and H3) or specialized chromatin (H2A.Z enriched at gene promoters and cenH3s enriched at centromeres). In this context, the identification of core histone-like "doublets" in the cytoplasmic replication factories of the Marseilleviridae (MV) is a novel finding with possible relevance to understanding the origin of eukaryotic chromatin. Here, we analyze and compare the core histone doublet genes from all known MV genomes as well as other MV genes relevant to the origin of the eukaryotic replisome. Using different phylogenetic approaches, we show that MV histone domains encode obligate H2B-H2A and H4-H3 dimers of possible proto-eukaryotic origin. MV core histone moieties form sister clades to each of the four eukaryotic clades of canonical and variant core histones. This suggests that MV core histone moieties diverged prior to eukaryotic neofunctionalizations associated with paired linear chromosomes and variant histone octamer assembly. We also show that MV genomes encode a proto-eukaryotic DNA topoisomerase II enzyme that forms a sister clade to eukaryotes. This is a relevant finding given that DNA topo II influences histone deposition and chromatin compaction and is the second most abundant nuclear protein after histones. The combined domain architecture and phylogenomic analyses presented here suggest that a primitive origin for MV histone genes is a more parsimonious explanation than horizontal gene transfers + gene fusions + sufficient divergence to eliminate relatedness to eukaryotic neofunctionalizations within the H2A and H3 clades without loss of relatedness to each of

  11. Generation of an induced pluripotent stem cell (iPSC line from a patient with maturity-onset diabetes of the young type 13 (MODY13 with a the potassium inwardly-rectifying channel, subfamily J, member 11 (KCNJ11 mutation

    Directory of Open Access Journals (Sweden)

    Frank Griscelli

    2017-08-01

    Full Text Available Heterozygous activating mutation (p.Glu227Lys in KCNJ11 leads to maturity-onset diabetes of the young (MODY type 13, that is a subtype of dominant inherited young-onset non-autoimmune diabetes due to a primary defect in pancreatic beta cells. We generated induced pluripotent stem cells (iPSCs from a patient with KCNJ11p.Glu227Lys mutation who developed MODY at 13 years old. KCNJ11p.Glu227Lys-mutated cells that were reprogrammed by non-integrative viral transduction had normal karyotype, harboured the KCNJ11p.Glu227Lys mutation, expressed pluripotency hallmarks and had the differentiation capacity into the three germ layers.

  12. Glucose- and interleukin-1beta-induced beta-cell apoptosis requires Ca2+ influx and extracellular signal-regulated kinase (ERK) 1/2 activation and is prevented by a sulfonylurea receptor 1/inwardly rectifying K+ channel 6.2 (SUR/Kir6.2) selective potassium channel opener in human islets

    DEFF Research Database (Denmark)

    Maedler, Kathrin; Størling, Joachim; Sturis, Jeppe

    2004-01-01

    Increasing evidence indicates that a progressive decrease in the functional beta-cell mass is the hallmark of both type 1 and type 2 diabetes. The underlying causes, beta-cell apoptosis and impaired secretory function, seem to be partly mediated by macrophage production of interleukin (IL)-1beta ...

  13. Snapshot of the Eukaryotic Gene Expression in Muskoxen Rumen—A Metatranscriptomic Approach

    Science.gov (United States)

    O'Toole, Nicholas; Barboza, Perry S.; Ungerfeld, Emilio; Leigh, Mary Beth; Selinger, L. Brent; Butler, Greg; Tsang, Adrian; McAllister, Tim A.; Forster, Robert J.

    2011-01-01

    Background Herbivores rely on digestive tract lignocellulolytic microorganisms, including bacteria, fungi and protozoa, to derive energy and carbon from plant cell wall polysaccharides. Culture independent metagenomic studies have been used to reveal the genetic content of the bacterial species within gut microbiomes. However, the nature of the genes encoded by eukaryotic protozoa and fungi within these environments has not been explored using metagenomic or metatranscriptomic approaches. Methodology/Principal Findings In this study, a metatranscriptomic approach was used to investigate the functional diversity of the eukaryotic microorganisms within the rumen of muskoxen (Ovibos moschatus), with a focus on plant cell wall degrading enzymes. Polyadenylated RNA (mRNA) was sequenced on the Illumina Genome Analyzer II system and 2.8 gigabases of sequences were obtained and 59129 contigs assembled. Plant cell wall degrading enzyme modules including glycoside hydrolases, carbohydrate esterases and polysaccharide lyases were identified from over 2500 contigs. These included a number of glycoside hydrolase family 6 (GH6), GH48 and swollenin modules, which have rarely been described in previous gut metagenomic studies. Conclusions/Significance The muskoxen rumen metatranscriptome demonstrates a much higher percentage of cellulase enzyme discovery and an 8.7x higher rate of total carbohydrate active enzyme discovery per gigabase of sequence than previous rumen metagenomes. This study provides a snapshot of eukaryotic gene expression in the muskoxen rumen, and identifies a number of candidate genes coding for potentially valuable lignocellulolytic enzymes. PMID:21655220

  14. Type VI secretion system MIX-effectors carry both antibacterial and anti-eukaryotic activities.

    Science.gov (United States)

    Ray, Ann; Schwartz, Nika; de Souza Santos, Marcela; Zhang, Junmei; Orth, Kim; Salomon, Dor

    2017-11-01

    Most type VI secretion systems (T6SSs) described to date are protein delivery apparatuses that mediate bactericidal activities. Several T6SSs were also reported to mediate virulence activities, although only few anti-eukaryotic effectors have been described. Here, we identify three T6SSs in the marine bacterium Vibrio proteolyticus and show that T6SS1 mediates bactericidal activities under warm marine-like conditions. Using comparative proteomics, we find nine potential T6SS1 effectors, five of which belong to the polymorphic MIX-effector class. Remarkably, in addition to six predicted bactericidal effectors, the T6SS1 secretome includes three putative anti-eukaryotic effectors. One of these is a MIX-effector containing a cytotoxic necrotizing factor 1 domain. We demonstrate that T6SS1 can use this MIX-effector to target phagocytic cells, resulting in morphological changes and actin cytoskeleton rearrangements. In conclusion, the V. proteolyticus T6SS1, a system homologous to one found in pathogenic vibrios, uses a suite of polymorphic effectors that target both bacteria and eukaryotic neighbors. © 2017 The Authors. Published under the terms of the CC BY 4.0 license.

  15. Death of a dogma: eukaryotic mRNAs can code for more than one protein.

    Science.gov (United States)

    Mouilleron, Hélène; Delcourt, Vivian; Roucou, Xavier

    2016-01-08

    mRNAs carry the genetic information that is translated by ribosomes. The traditional view of a mature eukaryotic mRNA is a molecule with three main regions, the 5' UTR, the protein coding open reading frame (ORF) or coding sequence (CDS), and the 3' UTR. This concept assumes that ribosomes translate one ORF only, generally the longest one, and produce one protein. As a result, in the early days of genomics and bioinformatics, one CDS was associated with each protein-coding gene. This fundamental concept of a single CDS is being challenged by increasing experimental evidence indicating that annotated proteins are not the only proteins translated from mRNAs. In particular, mass spectrometry (MS)-based proteomics and ribosome profiling have detected productive translation of alternative open reading frames. In several cases, the alternative and annotated proteins interact. Thus, the expression of two or more proteins translated from the same mRNA may offer a mechanism to ensure the co-expression of proteins which have functional interactions. Translational mechanisms already described in eukaryotic cells indicate that the cellular machinery is able to translate different CDSs from a single viral or cellular mRNA. In addition to summarizing data showing that the protein coding potential of eukaryotic mRNAs has been underestimated, this review aims to challenge the single translated CDS dogma. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

  16. Large variability of bathypelagic microbial eukaryotic communities across the world's oceans.

    Science.gov (United States)

    Pernice, Massimo C; Giner, Caterina R; Logares, Ramiro; Perera-Bel, Júlia; Acinas, Silvia G; Duarte, Carlos M; Gasol, Josep M; Massana, Ramon

    2016-04-01

    In this work, we study the diversity of bathypelagic microbial eukaryotes (0.8-20 μm) in the global ocean. Seawater samples from 3000 to 4000 m depth from 27 stations in the Atlantic, Pacific and Indian Oceans were analyzed by pyrosequencing the V4 region of the 18S ribosomal DNA. The relative abundance of the most abundant operational taxonomic units agreed with the results of a parallel metagenomic analysis, suggesting limited PCR biases in the tag approach. Although rarefaction curves for single stations were seldom saturated, the global analysis of all sequences together suggested an adequate recovery of bathypelagic diversity. Community composition presented a large variability among samples, which was poorly explained by linear geographic distance. In fact, the similarity between communities was better explained by water mass composition (26% of the variability) and the ratio in cell abundance between prokaryotes and microbial eukaryotes (21%). Deep diversity appeared dominated by four taxonomic groups (Collodaria, Chrysophytes, Basidiomycota and MALV-II) appearing in different proportions in each sample. Novel diversity amounted to 1% of the pyrotags and was lower than expected. Our study represents an essential step in the investigation of bathypelagic microbial eukaryotes, indicating dominating taxonomic groups and suggesting idiosyncratic assemblages in distinct oceanic regions.

  17. Solution structure of an archaeal DNA binding protein with an eukaryotic zinc finger fold.

    Directory of Open Access Journals (Sweden)

    Florence Guillière

    Full Text Available While the basal transcription machinery in archaea is eukaryal-like, transcription factors in archaea and their viruses are usually related to bacterial transcription factors. Nevertheless, some of these organisms show predicted classical zinc fingers motifs of the C2H2 type, which are almost exclusively found in proteins of eukaryotes and most often associated with transcription regulators. In this work, we focused on the protein AFV1p06 from the hyperthermophilic archaeal virus AFV1. The sequence of the protein consists of the classical eukaryotic C2H2 motif with the fourth histidine coordinating zinc missing, as well as of N- and C-terminal extensions. We showed that the protein AFV1p06 binds zinc and solved its solution structure by NMR. AFV1p06 displays a zinc finger fold with a novel structure extension and disordered N- and C-termini. Structure calculations show that a glutamic acid residue that coordinates zinc replaces the fourth histidine of the C2H2 motif. Electromobility gel shift assays indicate that the protein binds to DNA with different affinities depending on the DNA sequence. AFV1p06 is the first experimentally characterised archaeal zinc finger protein with a DNA binding activity. The AFV1p06 protein family has homologues in diverse viruses of hyperthermophilic archaea. A phylogenetic analysis points out a common origin of archaeal and eukaryotic C2H2 zinc fingers.

  18. Colorimetric sensor for triphosphates and their application as a viable staining agent for prokaryotes and eukaryotes.

    Science.gov (United States)

    Ghosh, Amrita; Shrivastav, Anupama; Jose, D Amilan; Mishra, Sanjiv K; Chandrakanth, C K; Mishra, Sandhya; Das, Amitava

    2008-07-15

    The chromogenic complex 1 x Zn (where 1 is (E)-4-(4-dimethylamino-phenylazo)-N,N-bispyridin-2-ylmethyl-benzenesulfonamide) showed high affinity toward the phosphate ion in tetrabutylammonium phosphate in acetonitrile solution and could preferentially bind to adenosine triphosphate (ATP) in aqueous solution at physiological pH. This binding caused a visual change in color, whereas no such change was noticed with other related anions (adenosine monophosphate, adenosine diphosphate, pyrophosphate, and phosphate) of biological significance. Thus, 1 x Zn could be used as a staining agent for different biological cells through binding to the ATP, generated in situ by the mitochondria (in eukaryotes). For prokaryotes (bacteria) the cell membrane takes care of the cells' energy conversion, since they lack mitochondria. ATP is produced in their unique cell structure on the cell membrane, which is not found in any eukaryotes. These stained cells could be viewed with normal light microscopy. This reagent could even be used for distinguishing the gram-positive and the gram-negative bacteria (prokaryotes). This dye was found to be nonlipophilic in nature and nontoxic to living microbes (eukaryotes and prokaryotes). Further, stained cells were found to grow in their respective media, and this confirmed the maintenance of viability of the microbes even after staining, unlike with many other dyes available commercially.

  19. Large variability of bathypelagic microbial eukaryotic communities across the world’s oceans

    KAUST Repository

    Pernice, Massimo C.

    2015-10-09

    In this work, we study the diversity of bathypelagic microbial eukaryotes (0.8–20 μm) in the global ocean. Seawater samples from 3000 to 4000 m depth from 27 stations in the Atlantic, Pacific and Indian Oceans were analyzed by pyrosequencing the V4 region of the 18S ribosomal DNA. The relative abundance of the most abundant operational taxonomic units agreed with the results of a parallel metagenomic analysis, suggesting limited PCR biases in the tag approach. Although rarefaction curves for single stations were seldom saturated, the global analysis of all sequences together suggested an adequate recovery of bathypelagic diversity. Community composition presented a large variability among samples, which was poorly explained by linear geographic distance. In fact, the similarity between communities was better explained by water mass composition (26% of the variability) and the ratio in cell abundance between prokaryotes and microbial eukaryotes (21%). Deep diversity appeared dominated by four taxonomic groups (Collodaria, Chrysophytes, Basidiomycota and MALV-II) appearing in different proportions in each sample. Novel diversity amounted to 1% of the pyrotags and was lower than expected. Our study represents an essential step in the investigation of bathypelagic microbial eukaryotes, indicating dominating taxonomic groups and suggesting idiosyncratic assemblages in distinct oceanic regions.

    The ISME Journal advance online publication, 9 October 2015; doi:10.1038/ismej.2015.170

  20. Phylogenomic analyses support the monophyly of Excavata and resolve relationships among eukaryotic "supergroups".

    Science.gov (United States)

    Hampl, Vladimir; Hug, Laura; Leigh, Jessica W; Dacks, Joel B; Lang, B Franz; Simpson, Alastair G B; Roger, Andrew J

    2009-03-10

    Nearly all of eukaryotic diversity has been classified into 6 suprakingdom-level groups (supergroups) based on molecular and morphological/cell-biological evidence; these are Opisthokonta, Amoebozoa, Archaeplastida, Rhizaria, Chromalveolata, and Excavata. However, molecular phylogeny has not provided clear evidence that either Chromalveolata or Excavata is monophyletic, nor has it resolved the relationships among the supergroups. To establish the affinities of Excavata, which contains parasites of global importance and organisms regarded previously as primitive eukaryotes, we conducted a phylogenomic analysis of a dataset of 143 proteins and 48 taxa, including 19 excavates. Previous phylogenomic studies have not included all major subgroups of Excavata, and thus have not definitively addressed their interrelationships. The enigmatic flagellate Andalucia is sister to typical jakobids. Jakobids (including Andalucia), Euglenozoa and Heterolobosea form a major clade that we name Discoba. Analyses of the complete dataset group Discoba with the mitochondrion-lacking excavates or "metamonads" (diplomonads, parabasalids, and Preaxostyla), but not with the final excavate group, Malawimonas. This separation likely results from a long-branch attraction artifact. Gradual removal of rapidly-evolving taxa from the dataset leads to moderate bootstrap support (69%) for the monophyly of all Excavata, and 90% support once all metamonads are removed. Most importantly, Excavata robustly emerges between unikonts (Amoebozoa + Opisthokonta) and "megagrouping" of Archaeplastida, Rhizaria, and chromalveolates. Our analyses indicate that Excavata forms a monophyletic suprakingdom-level group that is one of the 3 primary divisions within eukaryotes, along with unikonts and a megagroup of Archaeplastida, Rhizaria, and the chromalveolate lineages.

  1. Visualizing Patterns of Marine Eukaryotic Diversity from Metabarcoding Data Using QIIME.

    Science.gov (United States)

    Leray, Matthieu; Knowlton, Nancy

    2016-01-01

    PCR amplification followed by deep sequencing of homologous gene regions is increasingly used to characterize the diversity and taxonomic composition of marine eukaryotic communities. This approach may generate millions of sequences for hundreds of samples simultaneously. Therefore, tools that researchers can use to visualize complex patterns of diversity for these massive datasets are essential. Efforts by microbiologists to understand the Earth and human microbiomes using high-throughput sequencing of the 16S rRNA gene has led to the development of several user-friendly, open-source software packages that can be similarly used to analyze eukaryotic datasets. Quantitative Insights Into Microbial Ecology (QIIME) offers some of the most helpful data visualization tools. Here, we describe functionalities to import OTU tables generated with any molecular marker (e.g., 18S, COI, ITS) and associated metadata into QIIME. We then present a range of analytical tools implemented within QIIME that can be used to obtain insights about patterns of alpha and beta diversity for marine eukaryotes.

  2. EUPAN enables pan-genome studies of a large number of eukaryotic genomes.

    Science.gov (United States)

    Hu, Zhiqiang; Sun, Chen; Lu, Kuang-Chen; Chu, Xixia; Zhao, Yue; Lu, Jinyuan; Shi, Jianxin; Wei, Chaochun

    2017-08-01

    Pan-genome analyses are routinely carried out for bacteria to interpret the within-species gene presence/absence variations (PAVs). However, pan-genome analyses are rare for eukaryotes due to the large sizes and higher complexities of their genomes. Here we proposed EUPAN, a eukaryotic pan-genome analysis toolkit, enabling automatic large-scale eukaryotic pan-genome analyses and detection of gene PAVs at a relatively low sequencing depth. In the previous studies, we demonstrated the effectiveness and high accuracy of EUPAN in the pan-genome analysis of 453 rice genomes, in which we also revealed widespread gene PAVs among individual rice genomes. Moreover, EUPAN can be directly applied to the current re-sequencing projects primarily focusing on single nucleotide polymorphisms. EUPAN is implemented in Perl, R and C ++. It is supported under Linux and preferred for a computer cluster with LSF and SLURM job scheduling system. EUPAN together with its standard operating procedure (SOP) is freely available for non-commercial use (CC BY-NC 4.0) at http://cgm.sjtu.edu.cn/eupan/index.html . ccwei@sjtu.edu.cn or jianxin.shi@sjtu.edu.cn. Supplementary data are available at Bioinformatics online. © The Author (2017). Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com

  3. GenColors-based comparative genome databases for small eukaryotic genomes.

    Science.gov (United States)

    Felder, Marius; Romualdi, Alessandro; Petzold, Andreas; Platzer, Matthias; Sühnel, Jürgen; Glöckner, Gernot

    2013-01-01

    Many sequence data repositories can give a quick and easily accessible overview on genomes and their annotations. Less widespread is the possibility to compare related genomes with each other in a common database environment. We have previously described the GenColors database system (http://gencolors.fli-leibniz.de) and its applications to a number of bacterial genomes such as Borrelia, Legionella, Leptospira and Treponema. This system has an emphasis on genome comparison. It combines data from related genomes and provides the user with an extensive set of visualization and analysis tools. Eukaryote genomes are normally larger than prokaryote genomes and thus pose additional challenges for such a system. We have, therefore, adapted GenColors to also handle larger datasets of small eukaryotic genomes and to display eukaryotic gene structures. Further recent developments include whole genome views, genome list options and, for bacterial genome browsers, the display of horizontal gene transfer predictions. Two new GenColors-based databases for two fungal species (http://fgb.fli-leibniz.de) and for four social amoebas (http://sacgb.fli-leibniz.de) were set up. Both new resources open up a single entry point for related genomes for the amoebozoa and fungal research communities and other interested users. Comparative genomics approaches are greatly facilitated by these resources.

  4. Snapshot of the eukaryotic gene expression in muskoxen rumen--a metatranscriptomic approach.

    Directory of Open Access Journals (Sweden)

    Meng Qi

    Full Text Available BACKGROUND: Herbivores rely on digestive tract lignocellulolytic microorganisms, including bacteria, fungi and protozoa, to derive energy and carbon from plant cell wall polysaccharides. Culture independent metagenomic studies have been used to reveal the genetic content of the bacterial species within gut microbiomes. However, the nature of the genes encoded by eukaryotic protozoa and fungi within these environments has not been explored using metagenomic or metatranscriptomic approaches. METHODOLOGY/PRINCIPAL FINDINGS: In this study, a metatranscriptomic approach was used to investigate the functional diversity of the eukaryotic microorganisms within the rumen of muskoxen (Ovibos moschatus, with a focus on plant cell wall degrading enzymes. Polyadenylated RNA (mRNA was sequenced on the Illumina Genome Analyzer II system and 2.8 gigabases of sequences were obtained and 59129 contigs assembled. Plant cell wall degrading enzyme modules including glycoside hydrolases, carbohydrate esterases and polysaccharide lyases were identified from over 2500 contigs. These included a number of glycoside hydrolase family 6 (GH6, GH48 and swollenin modules, which have rarely been described in previous gut metagenomic studies. CONCLUSIONS/SIGNIFICANCE: The muskoxen rumen metatranscriptome demonstrates a much higher percentage of cellulase enzyme discovery and an 8.7x higher rate of total carbohydrate active enzyme discovery per gigabase of sequence than previous rumen metagenomes. This study provides a snapshot of eukaryotic gene expression in the muskoxen rumen, and identifies a number of candidate genes coding for potentially valuable lignocellulolytic enzymes.

  5. Evolution of an intricate J-protein network driving protein disaggregation in eukaryotes.

    Science.gov (United States)

    Nillegoda, Nadinath B; Stank, Antonia; Malinverni, Duccio; Alberts, Niels; Szlachcic, Anna; Barducci, Alessandro; De Los Rios, Paolo; Wade, Rebecca C; Bukau, Bernd

    2017-05-15

    Hsp70 participates in a broad spectrum of protein folding processes extending from nascent chain folding to protein disaggregation. This versatility in function is achieved through a diverse family of J-protein cochaperones that select substrates for Hsp70. Substrate selection is further tuned by transient complexation between different classes of J-proteins, which expands the range of protein aggregates targeted by metazoan Hsp70 for disaggregation. We assessed the prevalence and evolutionary conservation of J-protein complexation and cooperation in disaggregation. We find the emergence of a eukaryote-specific signature for interclass complexation of canonical J-proteins. Consistently, complexes exist in yeast and human cells, but not in bacteria, and correlate with cooperative action in disaggregation in vitro. Signature alterations exclude some J-proteins from networking, which ensures correct J-protein pairing, functional network integrity and J-protein specialization. This fundamental change in J-protein biology during the prokaryote-to-eukaryote transition allows for increased fine-tuning and broadening of Hsp70 function in eukaryotes.

  6. Diatoms dominate the eukaryotic metatranscriptome during spring in coastal 'dead zone' sediments.

    Science.gov (United States)

    Broman, Elias; Sachpazidou, Varvara; Dopson, Mark; Hylander, Samuel

    2017-10-11

    An important characteristic of marine sediments is the oxygen concentration that affects many central metabolic processes. There has been a widespread increase in hypoxia in coastal systems (referred to as 'dead zones') mainly caused by eutrophication. Hence, it is central to understand the metabolism and ecology of eukaryotic life in sediments during changing oxygen conditions. Therefore, we sampled coastal 'dead zone' Baltic Sea sediment during autumn and spring, and analysed the eukaryotic metatranscriptome from field samples and after incubation in the dark under oxic or anoxic conditions. Bacillariophyta (diatoms) dominated the eukaryotic metatranscriptome in spring and were also abundant during autumn. A large fraction of the diatom RNA reads was associated with the photosystems suggesting a constitutive expression in darkness. Microscope observation showed intact diatom cells and these would, if hatched, represent a significant part of the pelagic phytoplankton biomass. Oxygenation did not significantly change the relative proportion of diatoms nor resulted in any major shifts in metabolic 'signatures'. By contrast, diatoms rapidly responded when exposed to light suggesting that light is limiting diatom development in hypoxic sediments. Hence, it is suggested that diatoms in hypoxic sediments are on 'standby' to exploit the environment if they reach suitable habitats. © 2017 The Author(s).

  7. Metabolism in anoxic permeable sediments is dominated by eukaryotic dark fermentation

    Science.gov (United States)

    Bourke, Michael F.; Marriott, Philip J.; Glud, Ronnie N.; Hasler-Sheetal, Harald; Kamalanathan, Manoj; Beardall, John; Greening, Chris; Cook, Perran L. M.

    2017-01-01

    Permeable sediments are common across continental shelves and are critical contributors to marine biogeochemical cycling. Organic matter in permeable sediments is dominated by microalgae, which as eukaryotes have different anaerobic metabolic pathways to bacteria and archaea. Here we present analyses of flow-through reactor experiments showing that dissolved inorganic carbon is produced predominantly as a result of anaerobic eukaryotic metabolic activity. In our experiments, anaerobic production of dissolved inorganic carbon was consistently accompanied by large dissolved H2 production rates, suggesting the presence of fermentation. The production of both dissolved inorganic carbon and H2 persisted following administration of broad spectrum bactericidal antibiotics, but ceased following treatment with metronidazole. Metronidazole inhibits the ferredoxin/hydrogenase pathway of fermentative eukaryotic H2 production, suggesting that pathway as the source of H2 and dissolved inorganic carbon production. Metabolomic analysis showed large increases in lipid production at the onset of anoxia, consistent with documented pathways of anoxic dark fermentation in microalgae. Cell counts revealed a predominance of microalgae in the sediments. H2 production was observed in dark anoxic cultures of diatoms (Fragilariopsis sp.) and a chlorophyte (Pyramimonas) isolated from the study site, substantiating the hypothesis that microalgae undertake fermentation. We conclude that microalgal dark fermentation could be an important energy-conserving pathway in permeable sediments.

  8. Oxygenation of the Mesoproterozoic ocean and the evolution of complex eukaryotes

    Science.gov (United States)

    Zhang, Kan; Zhu, Xiangkun; Wood, Rachel A.; Shi, Yao; Gao, Zhaofu; Poulton, Simon W.

    2018-05-01

    The Mesoproterozoic era (1,600-1,000 million years ago (Ma)) has long been considered a period of relative environmental stasis, with persistently low levels of atmospheric oxygen. There remains much uncertainty, however, over the evolution of ocean chemistry during this period, which may have been of profound significance for the early evolution of eukaryotic life. Here we present rare earth element, iron-speciation and inorganic carbon isotope data to investigate the redox evolution of the 1,600-1,550 Ma Yanliao Basin, North China Craton. These data confirm that the ocean at the start of the Mesoproterozoic was dominantly anoxic and ferruginous. Significantly, however, we find evidence for a progressive oxygenation event starting at 1,570 Ma, immediately prior to the occurrence of complex multicellular eukaryotes in shelf areas of the Yanliao Basin. Our study thus demonstrates that oxygenation of the Mesoproterozoic environment was far more dynamic and intense than previously envisaged, and establishes an important link between rising oxygen and the emerging record of diverse, multicellular eukaryotic life in the early Mesoproterozoic.

  9. Tetrapyrroles as Endogenous TSPO Ligands in Eukaryotes and Prokaryotes: Comparisons with Synthetic Ligands

    Directory of Open Access Journals (Sweden)

    Leo Veenman

    2016-06-01

    Full Text Available The 18 kDa translocator protein (TSPO is highly 0conserved in eukaryotes and prokaryotes. Since its discovery in 1977, numerous studies established the TSPO’s importance for life essential functions. For these studies, synthetic TSPO ligands typically are applied. Tetrapyrroles present endogenous ligands for the TSPO. Tetrapyrroles are also evolutionarily conserved and regulate multiple functions. TSPO and tetrapyrroles regulate each other. In animals TSPO-tetrapyrrole interactions range from effects on embryonic development to metabolism, programmed cell death, response to stress, injury and disease, and even to life span extension. In animals TSPOs are primarily located in mitochondria. In plants TSPOs are also present in plastids, the nuclear fraction, the endoplasmic reticulum, and Golgi stacks. This may contribute to translocation of tetrapyrrole intermediates across organelles’ membranes. As in animals, plant TSPO binds heme and protoporphyrin IX. TSPO-tetrapyrrole interactions in plants appear to relate to development as well as stress conditions, including salt tolerance, abscisic acid-induced stress, reactive oxygen species homeostasis, and finally cell death regulation. In bacteria, TSPO is important for switching from aerobic to anaerobic metabolism, including the regulation of photosynthesis. As in mitochondria, in bacteria TSPO is located in the outer membrane. TSPO-tetrapyrrole interactions may be part of the establishment of the bacterial-eukaryote relationships, i.e., mitochondrial-eukaryote and plastid-plant endosymbiotic relationships.

  10. Heterologous Expression of Toxins from Bacterial Toxin-Antitoxin Systems in Eukaryotic Cells: Strategies and Applications

    Directory of Open Access Journals (Sweden)

    Chew Chieng Yeo

    2016-02-01

    Full Text Available Toxin-antitoxin (TA systems are found in nearly all prokaryotic genomes and usually consist of a pair of co-transcribed genes, one of which encodes a stable toxin and the other, its cognate labile antitoxin. Certain environmental and physiological cues trigger the degradation of the antitoxin, causing activation of the toxin, leading either to the death or stasis of the host cell. TA systems have a variety of functions in the bacterial cell, including acting as mediators of programmed cell death, the induction of a dormant state known as persistence and the stable maintenance of plasmids and other mobile genetic elements. Some bacterial TA systems are functional when expressed in eukaryotic cells and this has led to several innovative applications, which are the subject of this review. Here, we look at how bacterial TA systems have been utilized for the genetic manipulation of yeasts and other eukaryotes, for the containment of genetically modified organisms, and for the engineering of high expression eukaryotic cell lines. We also examine how TA systems have been adopted as an important tool in developmental biology research for the ablation of specific cells and the potential for utility of TA systems in antiviral and anticancer gene therapies.

  11. Heterologous Expression of Toxins from Bacterial Toxin-Antitoxin Systems in Eukaryotic Cells: Strategies and Applications

    Science.gov (United States)

    Yeo, Chew Chieng; Abu Bakar, Fauziah; Chan, Wai Ting; Espinosa, Manuel; Harikrishna, Jennifer Ann

    2016-01-01

    Toxin-antitoxin (TA) systems are found in nearly all prokaryotic genomes and usually consist of a pair of co-transcribed genes, one of which encodes a stable toxin and the other, its cognate labile antitoxin. Certain environmental and physiological cues trigger the degradation of the antitoxin, causing activation of the toxin, leading either to the death or stasis of the host cell. TA systems have a variety of functions in the bacterial cell, including acting as mediators of programmed cell death, the induction of a dormant state known as persistence and the stable maintenance of plasmids and other mobile genetic elements. Some bacterial TA systems are functional when expressed in eukaryotic cells and this has led to several innovative applications, which are the subject of this review. Here, we look at how bacterial TA systems have been utilized for the genetic manipulation of yeasts and other eukaryotes, for the containment of genetically modified organisms, and for the engineering of high expression eukaryotic cell lines. We also examine how TA systems have been adopted as an important tool in developmental biology research for the ablation of specific cells and the potential for utility of TA systems in antiviral and anticancer gene therapies. PMID:26907343

  12. Tracking the rise of eukaryotes to ecological dominance with zinc isotopes.

    Science.gov (United States)

    Isson, Terry T; Love, Gordon D; Dupont, Christopher L; Reinhard, Christopher T; Zumberge, Alex J; Asael, Dan; Gueguen, Bleuenn; McCrow, John; Gill, Ben C; Owens, Jeremy; Rainbird, Robert H; Rooney, Alan D; Zhao, Ming-Yu; Stueeken, Eva E; Konhauser, Kurt O; John, Seth G; Lyons, Timothy W; Planavsky, Noah J

    2018-06-05

    The biogeochemical cycling of zinc (Zn) is intimately coupled with organic carbon in the ocean. Based on an extensive new sedimentary Zn isotope record across Earth's history, we provide evidence for a fundamental shift in the marine Zn cycle ~800 million years ago. We discuss a wide range of potential drivers for this transition and propose that, within available constraints, a restructuring of marine ecosystems is the most parsimonious explanation for this shift. Using a global isotope mass balance approach, we show that a change in the organic Zn/C ratio is required to account for observed Zn isotope trends through time. Given the higher affinity of eukaryotes for Zn relative to prokaryotes, we suggest that a shift toward a more eukaryote-rich ecosystem could have provided a means of more efficiently sequestering organic-derived Zn. Despite the much earlier appearance of eukaryotes in the microfossil record (~1700 to 1600 million years ago), our data suggest a delayed rise to ecological prominence during the Neoproterozoic, consistent with the currently accepted organic biomarker records. © 2018 John Wiley & Sons Ltd.

  13. Unexpected Importance of Potential Parasites in the Composition of the Freshwater Small-Eukaryote Community▿

    Science.gov (United States)

    Lepère, Cécile; Domaizon, Isabelle; Debroas, Didier

    2008-01-01

    The diversity of small eukaryotes (0.2 to 5 μm) in a mesotrophic lake (Lake Bourget) was investigated using 18S rRNA gene library construction and fluorescent in situ hybridization coupled with tyramide signal amplification (TSA-FISH). Samples collected from the epilimnion on two dates were used to extend a data set previously obtained using similar approaches for lakes with a range of trophic types. A high level of diversity was recorded for this system with intermediate trophic status, and the main sequences from Lake Bourget were affiliated with ciliates (maximum, 19% of the operational taxonomic units [OTUs]), cryptophytes (33%), stramenopiles (13.2%), and cercozoa (9%). Although the comparison of TSA-FISH results and clone libraries suggested that the level of Chlorophyceae may have been underestimated using PCR with 18S rRNA primers, heterotrophic organisms dominated the small-eukaryote assemblage. We found that a large fraction of the sequences belonged to potential parasites of freshwater phytoplankton, including sequences affiliated with fungi and Perkinsozoa. On average, these sequences represented 30% of the OTUs (40% of the clones) obtained for each of two dates for Lake Bourget. Our results provide information on lacustrine small-eukaryote diversity and structure, adding to the phylogenetic data available for lakes with various trophic types. PMID:18359836

  14. Strong Decomposition of Random Variables

    DEFF Research Database (Denmark)

    Hoffmann-Jørgensen, Jørgen; Kagan, Abram M.; Pitt, Loren D.

    2007-01-01

    A random variable X is stongly decomposable if X=Y+Z where Y=Φ(X) and Z=X-Φ(X) are independent non-degenerated random variables (called the components). It is shown that at least one of the components is singular, and we derive a necessary and sufficient condition for strong decomposability...... of a discrete random variable....

  15. Strong coupling electroweak symmetry breaking

    International Nuclear Information System (INIS)

    Barklow, T.L.; Burdman, G.; Chivukula, R.S.

    1997-04-01

    The authors review models of electroweak symmetry breaking due to new strong interactions at the TeV energy scale and discuss the prospects for their experimental tests. They emphasize the direct observation of the new interactions through high-energy scattering of vector bosons. They also discuss indirect probes of the new interactions and exotic particles predicted by specific theoretical models

  16. Strong coupling electroweak symmetry breaking

    Energy Technology Data Exchange (ETDEWEB)

    Barklow, T.L. [Stanford Linear Accelerator Center, Menlo Park, CA (United States); Burdman, G. [Univ. of Wisconsin, Madison, WI (United States). Dept. of Physics; Chivukula, R.S. [Boston Univ., MA (United States). Dept. of Physics

    1997-04-01

    The authors review models of electroweak symmetry breaking due to new strong interactions at the TeV energy scale and discuss the prospects for their experimental tests. They emphasize the direct observation of the new interactions through high-energy scattering of vector bosons. They also discuss indirect probes of the new interactions and exotic particles predicted by specific theoretical models.

  17. The colours of strong interaction

    International Nuclear Information System (INIS)

    1995-01-01

    The aim of this session is to draw a consistent framework about the different ways to consider strong interaction. A large part is dedicated to theoretical work and the latest experimental results obtained at the first electron collider HERA are discussed. (A.C.)

  18. Strong cosmic censorship and the strong curvature singularities

    International Nuclear Information System (INIS)

    Krolak, A.

    1987-01-01

    Conditions are given under which any asymptotically simple and empty space-time that has a partial Cauchy surface with an asymptotically simple past is globally hyperbolic. It is shown that this result suggests that the Cauchy horizons of the type occurring in Reissner--Nordstroem and Kerr space-times are unstable. This in turn gives support for the validity of the strong cosmic censorship hypothesis

  19. Strongly Correlated Systems Theoretical Methods

    CERN Document Server

    Avella, Adolfo

    2012-01-01

    The volume presents, for the very first time, an exhaustive collection of those modern theoretical methods specifically tailored for the analysis of Strongly Correlated Systems. Many novel materials, with functional properties emerging from macroscopic quantum behaviors at the frontier of modern research in physics, chemistry and materials science, belong to this class of systems. Any technique is presented in great detail by its own inventor or by one of the world-wide recognized main contributors. The exposition has a clear pedagogical cut and fully reports on the most relevant case study where the specific technique showed to be very successful in describing and enlightening the puzzling physics of a particular strongly correlated system. The book is intended for advanced graduate students and post-docs in the field as textbook and/or main reference, but also for other researchers in the field who appreciates consulting a single, but comprehensive, source or wishes to get acquainted, in a as painless as po...

  20. Strongly correlated systems numerical methods

    CERN Document Server

    Mancini, Ferdinando

    2013-01-01

    This volume presents, for the very first time, an exhaustive collection of those modern numerical methods specifically tailored for the analysis of Strongly Correlated Systems. Many novel materials, with functional properties emerging from macroscopic quantum behaviors at the frontier of modern research in physics, chemistry and material science, belong to this class of systems. Any technique is presented in great detail by its own inventor or by one of the world-wide recognized main contributors. The exposition has a clear pedagogical cut and fully reports on the most relevant case study where the specific technique showed to be very successful in describing and enlightening the puzzling physics of a particular strongly correlated system. The book is intended for advanced graduate students and post-docs in the field as textbook and/or main reference, but also for other researchers in the field who appreciate consulting a single, but comprehensive, source or wishes to get acquainted, in a as painless as possi...

  1. Strongly correlated systems experimental techniques

    CERN Document Server

    Mancini, Ferdinando

    2015-01-01

    The continuous evolution and development of experimental techniques is at the basis of any fundamental achievement in modern physics. Strongly correlated systems (SCS), more than any other, need to be investigated through the greatest variety of experimental techniques in order to unveil and crosscheck the numerous and puzzling anomalous behaviors characterizing them. The study of SCS fostered the improvement of many old experimental techniques, but also the advent of many new ones just invented in order to analyze the complex behaviors of these systems. Many novel materials, with functional properties emerging from macroscopic quantum behaviors at the frontier of modern research in physics, chemistry and materials science, belong to this class of systems. The volume presents a representative collection of the modern experimental techniques specifically tailored for the analysis of strongly correlated systems. Any technique is presented in great detail by its own inventor or by one of the world-wide recognize...

  2. Flavour Democracy in Strong Unification

    CERN Document Server

    Abel, S A; Abel, Steven; King, Steven

    1998-01-01

    We show that the fermion mass spectrum may naturally be understood in terms of flavour democratic fixed points in supersymmetric theories which have a large domain of attraction in the presence of "strong unification". Our approach provides an alternative to the approximate Yukawa texture zeroes of the Froggatt-Nielsen mechanism. We discuss a particular model based on a broken gauged $SU(3)_L\\times SU(3)_R$ family symmetry which illustrates our approach.

  3. String dynamics at strong coupling

    International Nuclear Information System (INIS)

    Hull, C.M.

    1996-01-01

    The dynamics of superstring, supergravity and M-theories and their compactifications are probed by studying the various perturbation theories that emerge in the strong and weak-coupling limits for various directions in coupling constant space. The results support the picture of an underlying non-perturbative theory that, when expanded perturbatively in different coupling constants, gives different perturbation theories, which can be perturbative superstring theories or superparticle theories. The p-brane spectrum is considered in detail and a criterion found to establish which p-branes govern the strong-coupling dynamics. In many cases there are competing conjectures in the literature, and this analysis decides between them. In other cases, new results are found. The chiral 6-dimensional theory resulting from compactifying the type IIB string on K 3 is studied in detail and it is found that certain strong-coupling limits appear to give new theories, some of which hint at the possibility of a 12-dimensional origin. (orig.)

  4. Next-Generation Sequencing Assessment of Eukaryotic Diversity in Oil Sands Tailings Ponds Sediments and Surface Water.

    Science.gov (United States)

    Aguilar, Maria; Richardson, Elisabeth; Tan, BoonFei; Walker, Giselle; Dunfield, Peter F; Bass, David; Nesbø, Camilla; Foght, Julia; Dacks, Joel B

    2016-11-01

    Tailings ponds in the Athabasca oil sands (Canada) contain fluid wastes, generated by the extraction of bitumen from oil sands ores. Although the autochthonous prokaryotic communities have been relatively well characterized, almost nothing is known about microbial eukaryotes living in the anoxic soft sediments of tailings ponds or in the thin oxic layer of water that covers them. We carried out the first next-generation sequencing study of microbial eukaryotic diversity in oil sands tailings ponds. In metagenomes prepared from tailings sediment and surface water, we detected very low numbers of sequences encoding eukaryotic small subunit ribosomal RNA representing seven major taxonomic groups of protists. We also produced and analysed three amplicon-based 18S rRNA libraries prepared from sediment samples. These revealed a more diverse set of taxa, 169 different OTUs encompassing up to eleven higher order groups of eukaryotes, according to detailed classification using homology searching and phylogenetic methods. The 10 most abundant OTUs accounted for > 90% of the total of reads, vs. large numbers of rare OTUs (< 1% abundance). Despite the anoxic and hydrocarbon-enriched nature of the environment, the tailings ponds harbour complex communities of microbial eukaryotes indicating that these organisms should be taken into account when studying the microbiology of the oil sands. © 2016 The Author(s) Journal of Eukaryotic Microbiology © 2016 International Society of Protistologists.

  5. Intermediary metabolism in protists: a sequence-based view of facultative anaerobic metabolism in evolutionarily diverse eukaryotes.

    Science.gov (United States)

    Ginger, Michael L; Fritz-Laylin, Lillian K; Fulton, Chandler; Cande, W Zacheus; Dawson, Scott C

    2010-12-01

    Protists account for the bulk of eukaryotic diversity. Through studies of gene and especially genome sequences the molecular basis for this diversity can be determined. Evident from genome sequencing are examples of versatile metabolism that go far beyond the canonical pathways described for eukaryotes in textbooks. In the last 2-3 years, genome sequencing and transcript profiling has unveiled several examples of heterotrophic and phototrophic protists that are unexpectedly well-equipped for ATP production using a facultative anaerobic metabolism, including some protists that can (Chlamydomonas reinhardtii) or are predicted (Naegleria gruberi, Acanthamoeba castellanii, Amoebidium parasiticum) to produce H(2) in their metabolism. It is possible that some enzymes of anaerobic metabolism were acquired and distributed among eukaryotes by lateral transfer, but it is also likely that the common ancestor of eukaryotes already had far more metabolic versatility than was widely thought a few years ago. The discussion of core energy metabolism in unicellular eukaryotes is the subject of this review. Since genomic sequencing has so far only touched the surface of protist diversity, it is anticipated that sequences of additional protists may reveal an even wider range of metabolic capabilities, while simultaneously enriching our understanding of the early evolution of eukaryotes. Copyright © 2010 Elsevier GmbH. All rights reserved.

  6. PREFACE: Strongly correlated electron systems Strongly correlated electron systems

    Science.gov (United States)

    Saxena, Siddharth S.; Littlewood, P. B.

    2012-07-01

    This special section is dedicated to the Strongly Correlated Electron Systems Conference (SCES) 2011, which was held from 29 August-3 September 2011, in Cambridge, UK. SCES'2011 is dedicated to 100 years of superconductivity and covers a range of topics in the area of strongly correlated systems. The correlated electronic and magnetic materials featured include f-electron based heavy fermion intermetallics and d-electron based transition metal compounds. The selected papers derived from invited presentations seek to deepen our understanding of the rich physical phenomena that arise from correlation effects. The focus is on quantum phase transitions, non-Fermi liquid phenomena, quantum magnetism, unconventional superconductivity and metal-insulator transitions. Both experimental and theoretical work is presented. Based on fundamental advances in the understanding of electronic materials, much of 20th century materials physics was driven by miniaturisation and integration in the electronics industry to the current generation of nanometre scale devices. The achievements of this industry have brought unprecedented advances to society and well-being, and no doubt there is much further to go—note that this progress is founded on investments and studies in the fundamentals of condensed matter physics from more than 50 years ago. Nevertheless, the defining challenges for the 21st century will lie in the discovery in science, and deployment through engineering, of technologies that can deliver the scale needed to have an impact on the sustainability agenda. Thus the big developments in nanotechnology may lie not in the pursuit of yet smaller transistors, but in the design of new structures that can revolutionise the performance of solar cells, batteries, fuel cells, light-weight structural materials, refrigeration, water purification, etc. The science presented in the papers of this special section also highlights the underlying interest in energy-dense materials, which

  7. Atoms in strong laser fields

    International Nuclear Information System (INIS)

    L'Huillier, A.

    2002-01-01

    When a high-power laser focuses into a gas of atoms, the electromagnetic field becomes of the same magnitude as the Coulomb field which binds a 1s electron in a hydrogen atom. 3 highly non-linear phenomena can happen: 1) ATI (above threshold ionization): electrons initially in the ground state absorb a large number of photons, many more than the minimum number required for ionization; 2) multiple ionization: many electrons can be emitted one at a time, in a sequential process, or simultaneously in a mechanism called direct or non-sequential; and 3) high order harmonic generation (HHG): efficient photon emission in the extreme ultraviolet range, in the form of high-order harmonics of the fundamental laser field can occur. The theoretical problem consists in solving the time dependent Schroedinger equation (TDSE) that describes the interaction of a many-electron atom with a laser field. A number of methods have been proposed to solve this problem in the case of a hydrogen atom or a single-active electron atom in a strong laser field. A large effort is presently being devoted to go beyond the single-active approximation. The understanding of the physics of the interaction between atoms and strong laser fields has been provided by a very simple model called ''simple man's theory''. A unified view of HHG, ATI, and non-sequential ionization, originating from the simple man's model and the strong field approximation, expressed in terms of electrons trajectories or quantum paths is slowly emerging. (A.C.)

  8. Rydberg atoms in strong fields

    International Nuclear Information System (INIS)

    Kleppner, D.; Tsimmerman, M.

    1985-01-01

    Experimental and theoretical achievements in studying Rydberg atoms in external fields are considered. Only static (or quasistatic) fields and ''one-electron'' atoms, i.e. atoms that are well described by one-electron states, are discussed. Mainly behaviour of alkali metal atoms in electric field is considered. The state of theoretical investigations for hydrogen atom in magnetic field is described, but experimental data for atoms of alkali metals are presented as an illustration. Results of the latest experimental and theoretical investigations into the structure of Rydberg atoms in strong fields are presented

  9. Strong versions of Bell's theorem

    International Nuclear Information System (INIS)

    Stapp, H.P.

    1994-01-01

    Technical aspects of a recently constructed strong version of Bell's theorem are discussed. The theorem assumes neither hidden variables nor factorization, and neither determinism nor counterfactual definiteness. It deals directly with logical connections. Hence its relationship with modal logic needs to be described. It is shown that the proof can be embedded in an orthodox modal logic, and hence its compatibility with modal logic assured, but that this embedding weakens the theorem by introducing as added assumptions the conventionalities of the particular modal logic that is adopted. This weakening is avoided in the recent proof by using directly the set-theoretic conditions entailed by the locality assumption

  10. Strongly interacting light dark matter

    International Nuclear Information System (INIS)

    Bruggisser, Sebastian; Riva, Francesco; Urbano, Alfredo

    2016-07-01

    In the presence of approximate global symmetries that forbid relevant interactions, strongly coupled light Dark Matter (DM) can appear weakly coupled at small-energy and generate a sizable relic abundance. Fundamental principles like unitarity restrict these symmetries to a small class, where the leading interactions are captured by effective operators up to dimension-8. Chiral symmetry, spontaneously broken global symmetries and non-linearly realized supersymmetry are examples of this. Their DM candidates (composite fermions, pseudo-Nambu-Goldstone Bosons and Goldstini) are interesting targets for LHC missing-energy searches.

  11. Weak consistency and strong paraconsistency

    Directory of Open Access Journals (Sweden)

    Gemma Robles

    2009-11-01

    Full Text Available In a standard sense, consistency and paraconsistency are understood as, respectively, the absence of any contradiction and as the absence of the ECQ (“E contradictione quodlibet” rule that allows us to conclude any well formed formula from any contradiction. The aim of this paper is to explain the concepts of weak consistency alternative to the standard one, the concepts of paraconsistency related to them and the concept of strong paraconsistency, all of which have been defined by the author together with José M. Méndez.

  12. On the strong CP problem

    Energy Technology Data Exchange (ETDEWEB)

    Dowrick, N.J. (Dept. of Physics, Oxford (United Kingdom)); McDougall, N.A. (National Lab. for High Energy Physics, Tsukuba, Ibaraki (Japan))

    1992-07-09

    We show that two well-known solutions to the strong CP problem, the axion and a massless quark, may be understood in terms of the mechanism recently proposed by Samuel where long-range interactions between topological charges may be responsible for the removal of CP violation. We explain how the axion and a QCD meson (identified as the {eta}' if all quarks are massless) suppress fluctuations in global topological charge by almost identical dynamical although the masses, couplings and relevant length scales are very different. Furthermore, we elucidate the precise origin of the {eta}' mass. (orig.).

  13. Scalar strong interaction hadron theory

    CERN Document Server

    Hoh, Fang Chao

    2015-01-01

    The scalar strong interaction hadron theory, SSI, is a first principles' and nonlocal theory at quantum mechanical level that provides an alternative to low energy QCD and Higgs related part of the standard model. The quark-quark interaction is scalar rather than color-vectorial. A set of equations of motion for mesons and another set for baryons have been constructed. This book provides an account of the present state of a theory supposedly still at its early stage of development. This work will facilitate researchers interested in entering into this field and serve as a basis for possible future development of this theory.

  14. Estimation of strong ground motion

    International Nuclear Information System (INIS)

    Watabe, Makoto

    1993-01-01

    Fault model has been developed to estimate a strong ground motion in consideration of characteristics of seismic source and propagation path of seismic waves. There are two different approaches in the model. The first one is a theoretical approach, while the second approach is a semi-empirical approach. Though the latter is more practical than the former to be applied to the estimation of input motions, it needs at least the small-event records, the value of the seismic moment of the small event and the fault model of the large event

  15. Strong Mechanoluminescence from Oxynitridosilicate Phosphors

    Energy Technology Data Exchange (ETDEWEB)

    Zhang Lin; Xu Chaonan; Yamada, Hiroshi, E-mail: cn-xu@aist.go.jp [National Institute of Advanced Industrial Science and Technology (AIST), 807-1 Shuku, Tosu, Saga 841-0052 (Japan)

    2011-10-29

    We successfully developed a novel Mechanoluminescence (ML) material with water resistance, oxynitridosilicate; BaSi{sub 2}O{sub 2}N{sub 2}: Eu{sup 2+}. The crystal structure, photoluminescence (PL) and ML properties were characterized. The ML of BaSi{sub 2}O{sub 2}N{sub 2}: Eu{sup 2+} is so strong that the blue-green emission can be observed by the naked eyes clearly. In addition, it shows superior water resistance property. No changes were found in the ML intensities during the total water treatment test.

  16. Effective lagrangian for strong interactions

    International Nuclear Information System (INIS)

    Jain, P.

    1988-01-01

    We attempt to construct a realistic phenomenological Lagrangian in order to describe strong interactions. This is in general a very complicated problem and we shall explore its various aspects. We first include the vector mesons by writing down the most general chiral invariant terms proportional to the Levi-Civita symbol ε μναβ . These terms involve three unknown coefficients, which are calculated by using the experimental results of strong interaction processes. We then calculate the static nucleon properties by finding the solitonic excitations of this model. The results turn out to be, as is also the case for most other vector-pseudoscalar Lagrangians, better than the Skyrme model but are still somewhat different from the experiments. Another aspect that we shall study is the incorporation of scale anomaly of QCD into the Skyrme model. We thus introduce a scalar glueball in our Lagrangian. Here we find an interesting result that the effective glue field dynamically forms a bag for the soliton. Depending on the values of the parameters, we get either a deep bag or a shallow bag. However by including the scalar meson, we find that to get realistic scalar sector we must have the shallow bag. Finally we show some intriguing connections between the chiral quark model, in which the nucleon is described as a solitonic excitation, and the ordinary potential binding quark model

  17. EDITORIAL: Strongly correlated electron systems Strongly correlated electron systems

    Science.gov (United States)

    Ronning, Filip; Batista, Cristian

    2011-03-01

    Strongly correlated electrons is an exciting and diverse field in condensed matter physics. This special issue aims to capture some of that excitement and recent developments in the field. Given that this issue was inspired by the 2010 International Conference on Strongly Correlated Electron Systems (SCES 2010), we briefly give some history in order to place this issue in context. The 2010 International Conference on Strongly Correlated Electron Systems was held in Santa Fe, New Mexico, a reunion of sorts from the 1989 International Conference on the Physics of Highly Correlated Electron Systems that also convened in Santa Fe. SCES 2010—co-chaired by John Sarrao and Joe Thompson—followed the tradition of earlier conferences, in this century, hosted by Buzios (2008), Houston (2007), Vienna (2005), Karlsruhe (2004), Krakow (2002) and Ann Arbor (2001). Every three years since 1997, SCES has joined the International Conference on Magnetism (ICM), held in Recife (2000), Rome (2003), Kyoto (2006) and Karlsruhe (2009). Like its predecessors, SCES 2010 topics included strongly correlated f- and d-electron systems, heavy-fermion behaviors, quantum-phase transitions, non-Fermi liquid phenomena, unconventional superconductivity, and emergent states that arise from electronic correlations. Recent developments from studies of quantum magnetism and cold atoms complemented the traditional subjects and were included in SCES 2010. 2010 celebrated the 400th anniversary of Santa Fe as well as the birth of astronomy. So what's the connection to SCES? The Dutch invention of the first practical telescope and its use by Galileo in 1610 and subsequent years overturned dogma that the sun revolved about the earth. This revolutionary, and at the time heretical, conclusion required innovative combinations of new instrumentation, observation and mathematics. These same combinations are just as important 400 years later and are the foundation of scientific discoveries that were discussed

  18. Strong Selective Adsorption of Polymers.

    Science.gov (United States)

    Ge, Ting; Rubinstein, Michael

    2015-06-09

    A scaling theory is developed for selective adsorption of polymers induced by the strong binding between specific monomers and complementary surface adsorption sites. By "selective" we mean specific attraction between a subset of all monomers, called "sticky", and a subset of surface sites, called "adsorption sites". We demonstrate that, in addition to the expected dependence on the polymer volume fraction ϕ bulk in the bulk solution, selective adsorption strongly depends on the ratio between two characteristic length scales, the root-mean-square distance l between neighboring sticky monomers along the polymer, and the average distance d between neighboring surface adsorption sites. The role of the ratio l / d arises from the fact that a polymer needs to deform to enable the spatial commensurability between its sticky monomers and the surface adsorption sites for selective adsorption. We study strong selective adsorption of both telechelic polymers with two end monomers being sticky and multisticker polymers with many sticky monomers between sticky ends. For telechelic polymers, we identify four adsorption regimes at l / d 1, we expect that the adsorption layer at exponentially low ϕ bulk consists of separated unstretched loops, while as ϕ bulk increases the layer crosses over to a brush of extended loops with a second layer of weakly overlapping tails. For multisticker chains, in the limit of exponentially low ϕ bulk , adsorbed polymers are well separated from each other. As l / d increases, the conformation of an individual polymer changes from a single-end-adsorbed "mushroom" to a random walk of loops. For high ϕ bulk , adsorbed polymers at small l / d are mushrooms that cover all the adsorption sites. At sufficiently large l / d , adsorbed multisticker polymers strongly overlap. We anticipate the formation of a self-similar carpet and with increasing l / d a two-layer structure with a brush of loops covered by a self-similar carpet. As l / d exceeds the

  19. Identification of eukaryotic open reading frames in metagenomic cDNA libraries made from environmental samples.

    Science.gov (United States)

    Grant, Susan; Grant, William D; Cowan, Don A; Jones, Brian E; Ma, Yanhe; Ventosa, Antonio; Heaphy, Shaun

    2006-01-01

    Here we describe the application of metagenomic technologies to construct cDNA libraries from RNA isolated from environmental samples. RNAlater (Ambion) was shown to stabilize RNA in environmental samples for periods of at least 3 months at -20 degrees C. Protocols for library construction were established on total RNA extracted from Acanthamoeba polyphaga trophozoites. The methodology was then used on algal mats from geothermal hot springs in Tengchong county, Yunnan Province, People's Republic of China, and activated sludge from a sewage treatment plant in Leicestershire, United Kingdom. The Tenchong libraries were dominated by RNA from prokaryotes, reflecting the mainly prokaryote microbial composition. The majority of these clones resulted from rRNA; only a few appeared to be derived from mRNA. In contrast, many clones from the activated sludge library had significant similarity to eukaryote mRNA-encoded protein sequences. A library was also made using polyadenylated RNA isolated from total RNA from activated sludge; many more clones in this library were related to eukaryotic mRNA sequences and proteins. Open reading frames (ORFs) up to 378 amino acids in size could be identified. Some resembled known proteins over their full length, e.g., 36% match to cystatin, 49% match to ribosomal protein L32, 63% match to ribosomal protein S16, 70% to CPC2 protein. The methodology described here permits the polyadenylated transcriptome to be isolated from environmental samples with no knowledge of the identity of the microorganisms in the sample or the necessity to culture them. It has many uses, including the identification of novel eukaryotic ORFs encoding proteins and enzymes.

  20. A set of ligation-independent in vitro translation vectors for eukaryotic protein production

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    Endo Yaeta

    2008-03-01

    Full Text Available Abstract Background The last decade has brought the renaissance of protein studies and accelerated the development of high-throughput methods in all aspects of proteomics. Presently, most protein synthesis systems exploit the capacity of living cells to translate proteins, but their application is limited by several factors. A more flexible alternative protein production method is the cell-free in vitro protein translation. Currently available in vitro translation systems are suitable for high-throughput robotic protein production, fulfilling the requirements of proteomics studies. Wheat germ extract based in vitro translation system is likely the most promising method, since numerous eukaryotic proteins can be cost-efficiently synthesized in their native folded form. Although currently available vectors for wheat embryo in vitro translation systems ensure high productivity, they do not meet the requirements of state-of-the-art proteomics. Target genes have to be inserted using restriction endonucleases and the plasmids do not encode cleavable affinity purification tags. Results We designed four ligation independent cloning (LIC vectors for wheat germ extract based in vitro protein translation. In these constructs, the RNA transcription is driven by T7 or SP6 phage polymerase and two TEV protease cleavable affinity tags can be added to aid protein purification. To evaluate our improved vectors, a plant mitogen activated protein kinase was cloned in all four constructs. Purification of this eukaryotic protein kinase demonstrated that all constructs functioned as intended: insertion of PCR fragment by LIC worked efficiently, affinity purification of translated proteins by GST-Sepharose or MagneHis particles resulted in high purity kinase, and the affinity tags could efficiently be removed under different reaction conditions. Furthermore, high in vitro kinase activity testified of proper folding of the purified protein. Conclusion Four newly

  1. How MCM loading and spreading specify eukaryotic DNA replication initiation sites [version 1; referees: 4 approved

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    Olivier Hyrien

    2016-08-01

    Full Text Available DNA replication origins strikingly differ between eukaryotic species and cell types. Origins are localized and can be highly efficient in budding yeast, are randomly located in early fly and frog embryos, which do not transcribe their genomes, and are clustered in broad (10-100 kb non-transcribed zones, frequently abutting transcribed genes, in mammalian cells. Nonetheless, in all cases, origins are established during the G1-phase of the cell cycle by the loading of double hexamers of the Mcm 2-7 proteins (MCM DHs, the core of the replicative helicase. MCM DH activation in S-phase leads to origin unwinding, polymerase recruitment, and initiation of bidirectional DNA synthesis. Although MCM DHs are initially loaded at sites defined by the binding of the origin recognition complex (ORC, they ultimately bind chromatin in much greater numbers than ORC and only a fraction are activated in any one S-phase. Data suggest that the multiplicity and functional redundancy of MCM DHs provide robustness to the replication process and affect replication time and that MCM DHs can slide along the DNA and spread over large distances around the ORC. Recent studies further show that MCM DHs are displaced along the DNA by collision with transcription complexes but remain functional for initiation after displacement. Therefore, eukaryotic DNA replication relies on intrinsically mobile and flexible origins, a strategy fundamentally different from bacteria but conserved from yeast to human. These properties of MCM DHs likely contribute to the establishment of broad, intergenic replication initiation zones in higher eukaryotes.

  2. How MCM loading and spreading specify eukaryotic DNA replication initiation sites.

    Science.gov (United States)

    Hyrien, Olivier

    2016-01-01

    DNA replication origins strikingly differ between eukaryotic species and cell types. Origins are localized and can be highly efficient in budding yeast, are randomly located in early fly and frog embryos, which do not transcribe their genomes, and are clustered in broad (10-100 kb) non-transcribed zones, frequently abutting transcribed genes, in mammalian cells. Nonetheless, in all cases, origins are established during the G1-phase of the cell cycle by the loading of double hexamers of the Mcm 2-7 proteins (MCM DHs), the core of the replicative helicase. MCM DH activation in S-phase leads to origin unwinding, polymerase recruitment, and initiation of bidirectional DNA synthesis. Although MCM DHs are initially loaded at sites defined by the binding of the origin recognition complex (ORC), they ultimately bind chromatin in much greater numbers than ORC and only a fraction are activated in any one S-phase. Data suggest that the multiplicity and functional redundancy of MCM DHs provide robustness to the replication process and affect replication time and that MCM DHs can slide along the DNA and spread over large distances around the ORC. Recent studies further show that MCM DHs are displaced along the DNA by collision with transcription complexes but remain functional for initiation after displacement. Therefore, eukaryotic DNA replication relies on intrinsically mobile and flexible origins, a strategy fundamentally different from bacteria but conserved from yeast to human. These properties of MCM DHs likely contribute to the establishment of broad, intergenic replication initiation zones in higher eukaryotes.

  3. The nature and origin of nucleus-like intracellular inclusions in Paleoproterozoic eukaryote microfossils.

    Science.gov (United States)

    Pang, K; Tang, Q; Schiffbauer, J D; Yao, J; Yuan, X; Wan, B; Chen, L; Ou, Z; Xiao, S

    2013-11-01

    The well-known debate on the nature and origin of intracellular inclusions (ICIs) in silicified microfossils from the early Neoproterozoic Bitter Springs Formation has recently been revived by reports of possible fossilized nuclei in phosphatized animal embryo-like fossils from the Ediacaran Doushantuo Formation of South China. The revisitation of this discussion prompted a critical and comprehensive investigation of ICIs in some of the oldest indisputable eukaryote microfossils-the ornamented acritarchs Dictyosphaera delicata and Shuiyousphaeridium macroreticulatum from the Paleoproterozoic Ruyang Group of North China-using a suite of characterization approaches: scanning electron microscopy (SEM), transmission electron microscopy (TEM), and focused ion beam scanning electron microscopy (FIB-SEM). Although the Ruyang acritarchs must have had nuclei when alive, our data suggest that their ICIs represent neither fossilized nuclei nor taphonomically condensed cytoplasm. We instead propose that these ICIs likely represent biologically contracted and consolidated eukaryotic protoplasts (the combination of the nucleus, surrounding cytoplasm, and plasma membrane). As opposed to degradational contraction of prokaryotic cells within a mucoidal sheath-a model proposed to explain the Bitter Springs ICIs-our model implies that protoplast condensation in the Ruyang acritarchs was an in vivo biologically programmed response to adverse conditions in preparation for encystment. While the discovery of bona fide nuclei in Paleoproterozoic acritarchs would be a substantial landmark in our understanding of eukaryote evolution, the various processes (such as degradational and biological condensation of protoplasts) capable of producing nuclei-mimicking structures require that interpretation of ICIs as fossilized nuclei be based on comprehensive investigations. © 2013 John Wiley & Sons Ltd.

  4. Structural basis for the initiation of eukaryotic transcription-coupled DNA repair.

    Science.gov (United States)

    Xu, Jun; Lahiri, Indrajit; Wang, Wei; Wier, Adam; Cianfrocco, Michael A; Chong, Jenny; Hare, Alissa A; Dervan, Peter B; DiMaio, Frank; Leschziner, Andres E; Wang, Dong

    2017-11-30

    Eukaryotic transcription-coupled repair (TCR) is an important and well-conserved sub-pathway of nucleotide excision repair that preferentially removes DNA lesions from the template strand that block translocation of RNA polymerase II (Pol II). Cockayne syndrome group B (CSB, also known as ERCC6) protein in humans (or its yeast orthologues, Rad26 in Saccharomyces cerevisiae and Rhp26 in Schizosaccharomyces pombe) is among the first proteins to be recruited to the lesion-arrested Pol II during the initiation of eukaryotic TCR. Mutations in CSB are associated with the autosomal-recessive neurological disorder Cockayne syndrome, which is characterized by progeriod features, growth failure and photosensitivity. The molecular mechanism of eukaryotic TCR initiation remains unclear, with several long-standing unanswered questions. How cells distinguish DNA lesion-arrested Pol II from other forms of arrested Pol II, the role of CSB in TCR initiation, and how CSB interacts with the arrested Pol II complex are all unknown. The lack of structures of CSB or the Pol II-CSB complex has hindered our ability to address these questions. Here we report the structure of the S. cerevisiae Pol II-Rad26 complex solved by cryo-electron microscopy. The structure reveals that Rad26 binds to the DNA upstream of Pol II, where it markedly alters its path. Our structural and functional data suggest that the conserved Swi2/Snf2-family core ATPase domain promotes the forward movement of Pol II, and elucidate key roles for Rad26 in both TCR and transcription elongation.

  5. Bacterial Signaling Nucleotides Inhibit Yeast Cell Growth by Impacting Mitochondrial and Other Specifically Eukaryotic Functions

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    Andy Hesketh

    2017-07-01

    Full Text Available We have engineered Saccharomyces cerevisiae to inducibly synthesize the prokaryotic signaling nucleotides cyclic di-GMP (cdiGMP, cdiAMP, and ppGpp in order to characterize the range of effects these nucleotides exert on eukaryotic cell function during bacterial pathogenesis. Synthetic genetic array (SGA and transcriptome analyses indicated that, while these compounds elicit some common reactions in yeast, there are also complex and distinctive responses to each of the three nucleotides. All three are capable of inhibiting eukaryotic cell growth, with the guanine nucleotides exhibiting stronger effects than cdiAMP. Mutations compromising mitochondrial function and chromatin remodeling show negative epistatic interactions with all three nucleotides. In contrast, certain mutations that cause defects in chromatin modification and ribosomal protein function show positive epistasis, alleviating growth inhibition by at least two of the three nucleotides. Uniquely, cdiGMP is lethal both to cells growing by respiration on acetate and to obligately fermentative petite mutants. cdiGMP is also synthetically lethal with the ribonucleotide reductase (RNR inhibitor hydroxyurea. Heterologous expression of the human ppGpp hydrolase Mesh1p prevented the accumulation of ppGpp in the engineered yeast and restored cell growth. Extensive in vivo interactions between bacterial signaling molecules and eukaryotic gene function occur, resulting in outcomes ranging from growth inhibition to death. cdiGMP functions through a mechanism that must be compensated by unhindered RNR activity or by functionally competent mitochondria. Mesh1p may be required for abrogating the damaging effects of ppGpp in human cells subjected to bacterial infection.

  6. Influence of Vitamin B Auxotrophy on Nitrogen Metabolism in Eukaryotic Phytoplankton

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    Erin M Bertrand

    2012-10-01

    Full Text Available While nitrogen availability is known to limit primary production in large parts of the ocean, vitamin starvation amongst eukaryotic phytoplankton is becoming increasingly recognized as an oceanographically relevant phenomenon. Cobalamin (B12 and thiamine (B1 auxotrophy are widespread throughout eukaryotic phytoplankton, with over 50% of cultured isolates requiring B12 and 20% requiring B1. The frequency of vitamin auxotrophy in harmful algal bloom species is even higher. Instances of colimitation between nitrogen and B vitamins have been observed in marine environments, and interactions between these nutrients have been shown to impact phytoplankton species composition. This review evaluates the potential for interactive effects of nitrogen and vitamin B12 and B1 starvation in eukaryotic phytoplankton. B12 plays essential roles in amino acid and one-carbon metabolism, while B1 is important for primary carbohydrate and amino acid metabolism and likely useful as an anti-oxidant. Here we will focus on three potential metabolic interconnections between vitamin, nitrogen and sulfur metabolism that may have ramifications for the role of vitamin and nitrogen scarcities in driving ocean productivity and species composition. These include: (1 B12, B1, and N starvation impacts on osmolyte and antioxidant production, (2 B12 and B1 starvation impacts on polyamine biosynthesis, and (3 influence of B12 and B1 starvation on the diatom urea cycle and amino acid recycling through impacts on the citric acid cycle. We evaluate evidence for these interconnections and identify oceanographic contexts in which each may impact rates of primary production and phytoplankton community composition. Major implications include that B12 and B1 deprivation may impair the ability of phytoplankton to recover from nitrogen starvation and that changes in vitamin and nitrogen availability may synergistically impact harmful algal bloom formation.

  7. How and why DNA barcodes underestimate the diversity of microbial eukaryotes.

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    Gwenael Piganeau

    Full Text Available BACKGROUND: Because many picoplanktonic eukaryotic species cannot currently be maintained in culture, direct sequencing of PCR-amplified 18S ribosomal gene DNA fragments from filtered sea-water has been successfully used to investigate the astounding diversity of these organisms. The recognition of many novel planktonic organisms is thus based solely on their 18S rDNA sequence. However, a species delimited by its 18S rDNA sequence might contain many cryptic species, which are highly differentiated in their protein coding sequences. PRINCIPAL FINDINGS: Here, we investigate the issue of species identification from one gene to the whole genome sequence. Using 52 whole genome DNA sequences, we estimated the global genetic divergence in protein coding genes between organisms from different lineages and compared this to their ribosomal gene sequence divergences. We show that this relationship between proteome divergence and 18S divergence is lineage dependent. Unicellular lineages have especially low 18S divergences relative to their protein sequence divergences, suggesting that 18S ribosomal genes are too conservative to assess planktonic eukaryotic diversity. We provide an explanation for this lineage dependency, which suggests that most species with large effective population sizes will show far less divergence in 18S than protein coding sequences. CONCLUSIONS: There is therefore a trade-off between using genes that are easy to amplify in all species, but which by their nature are highly conserved and underestimate the true number of species, and using genes that give a better description of the number of species, but which are more difficult to amplify. We have shown that this trade-off differs between unicellular and multicellular organisms as a likely consequence of differences in effective population sizes. We anticipate that biodiversity of microbial eukaryotic species is underestimated and that numerous "cryptic species" will become

  8. Construction and identification of eukaryotic expression vector of pcDNA3-UHRF1

    International Nuclear Information System (INIS)

    Li Xinli; Zhu Ran; Zhu Wei; Fan Saijun; Meng Qinghui

    2011-01-01

    Objective: To generate eukaryotic expression vector of pcDNA3-UHRF1(ubiquitin-like, containing PHD and RING finger domains 1, UHRF1) and testify its expression in breast cancer cells MDA-MB-231. Methods: A 2.3 kb cDNA fragment was amplified from the total RNA of the human breast cancer cells MCF-7 by the RT-PCR method and was cloned into the plasmid pcDNA3. The vector was identified by the double digestion with restriction enzymes Kpn I and Xho I and was sequenced. The cDNA of UHRF1 was transfected into human breast cancer cells MDA-MB-231 by Lipofactamin2000. The positive clones were selected by G418. The expression of the UHRF1 was detected by RT-PCR and Western blot analysis. Results: The recombinant eukaryotic expression vector pcDNA3-UHRF1 was digested with Kpn I and BamH I, and the electrophoresis of the digested products showed two fragments; 2.3kb fragment of UHRF1 and 5.4 kb fragment of pcDNA3, and the sequence inserted was identical to the published sequence. The MDA-MB-231 cells transfected with the pcDNA3-UHRF1 plasmid expressed a high level of the UHRF1 mRNA and protein. Conclusion: The recombinant eukaryotic cell expression vector of pcDNA3-UHRF1 is constructed successfully. The recombinant plasmid pcDNA3-UHRF1 can provide a very useful tool and lay an important foundation for the research on the function of UHRF1. (authors)

  9. Interplay of noncoding RNAs, mRNAs, and proteins during the growth of eukaryotic cells

    International Nuclear Information System (INIS)

    Zhdanov, V. P.

    2010-01-01

    Numerous biological functions of noncoding RNAs (ncRNAs) in eukaryotic cells are based primarily on their ability to pair with target mRNAs and then either to prevent translation or to result in rapid degradation of the mRNA-ncRNA complex. Using a general model describing this scenario, we show that ncRNAs may help to maintain constant mRNA and protein concentrations during the growth of cells. The possibility of observation of this effect on the global scale is briefly discussed.

  10. Microsatellites in the Eukaryotic DNA Mismatch Repair Genes as Modulators of Evolutionary Mutation Rate

    Science.gov (United States)

    Chang, Dong Kyung; Metzgar, David; Wills, Christopher; Boland, C. Richard

    2003-01-01

    All "minor" components of the human DNA mismatch repair (MMR) system-MSH3, MSH6, PMS2, and the recently discovered MLH3-contain mononucleotide microsatellites in their coding sequences. This intriguing finding contrasts with the situation found in the major components of the DNA MMR system-MSH2 and MLH1-and, in fact, most human genes. Although eukaryotic genomes are rich in microsatellites, non-triplet microsatellites are rare in coding regions. The recurring presence of exonal mononucleotide repeat sequences within a single family of human genes would therefore be considered exceptional.

  11. Evaluation by mass fragmentography of metabolic pathways of endogenous and exogenous compounds in eukaryote cell cultures

    International Nuclear Information System (INIS)

    Padieu, P.; Maume, B.F.

    1977-01-01

    Carbon-14 labelled compounds in cell cultures are used to establish the interconnections between different metabolic pathways as well as the competitive action of effectors on these different pathways. Analysis was performed by the GC-MS combination. Identification was carried out by comparison with the mass spectra of d9-TMS, 35 Cl-TMS and 37 Cl-TMS derivatizations of the culture extracts. Examples are given of the metabolic study of hormonal steroids and of safrale, a carcinogenic compound, by differentiated eukaryotic cells in cultures from the rat

  12. Heat degradation of eukaryotic and bacterial DNA: an experimental model for paleomicrobiology

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    Nguyen-Hieu Tung

    2012-09-01

    Full Text Available Abstract Background Theoretical models suggest that DNA degradation would sharply limit the PCR-based detection of both eukaryotic and prokaryotic DNA within ancient specimens. However, the relative extent of decay of eukaryote and prokaryote DNA over time is a matter of debate. In this study, the murine macrophage cell line J774, alone or infected with Mycobacterium smegmatis bacteria, were killed after exposure to 90°C dry heat for intervals ranging from 1 to 48 h in order to compare eukaryotic cells, extracellular bacteria and intracellular bacteria. The sizes of the resulting mycobacterial rpoB and murine rpb2 homologous gene fragments were then determined by real-time PCR and fluorescent probing. Findings The cycle threshold (Ct values of PCR-amplified DNA fragments from J774 cells and the M. smegmatis negative controls (without heat exposure varied from 26–33 for the J774 rpb2 gene fragments and from 24–29 for M. smegmatis rpoB fragments. After 90°C dry heat incubation for up to 48 h, the Ct values of test samples increased relative to those of the controls for each amplicon size. For each dry heat exposure time, the Ct values of the 146-149-bp fragments were lower than those of 746-747-bp fragments. During the 4- to 24-h dry heat incubation, the non-infected J774 cell DNA was degraded into 597-bp rpb2 fragments. After 48 h, however, only 450-bp rpb2 fragments of both non-infected and infected J774 cells could be amplified. In contrast, the 746-bp rpoB fragments of M. smegmatis DNA could be amplified after the 48-h dry heat exposure in all experiments. Infected and non-infected J774 cell DNA was degraded more rapidly than M. smegmatis DNA after dry heat exposure (ANOVA test, p  Conclusion In this study, mycobacterial DNA was more resistant to dry-heat stress than eukaryotic DNA. Therefore, the detection of large, experimental, ancient mycobacterial DNA fragments is a suitable approach for paleomicrobiological studies.

  13. A molecular timescale of eukaryote evolution and the rise of complex multicellular life

    Science.gov (United States)

    Hedges, S. Blair; Blair, Jaime E.; Venturi, Maria L.; Shoe, Jason L.

    2004-01-01

    BACKGROUND: The pattern and timing of the rise in complex multicellular life during Earth's history has not been established. Great disparity persists between the pattern suggested by the fossil record and that estimated by molecular clocks, especially for plants, animals, fungi, and the deepest branches of the eukaryote tree. Here, we used all available protein sequence data and molecular clock methods to place constraints on the increase in complexity through time. RESULTS: Our phylogenetic analyses revealed that (i) animals are more closely related to fungi than to plants, (ii) red algae are closer to plants than to animals or fungi, (iii) choanoflagellates are closer to animals than to fungi or plants, (iv) diplomonads, euglenozoans, and alveolates each are basal to plants+animals+fungi, and (v) diplomonads are basal to other eukaryotes (including alveolates and euglenozoans). Divergence times were estimated from global and local clock methods using 20-188 proteins per node, with data treated separately (multigene) and concatenated (supergene). Different time estimation methods yielded similar results (within 5%): vertebrate-arthropod (964 million years ago, Ma), Cnidaria-Bilateria (1,298 Ma), Porifera-Eumetozoa (1,351 Ma), Pyrenomycetes-Plectomycetes (551 Ma), Candida-Saccharomyces (723 Ma), Hemiascomycetes-filamentous Ascomycota (982 Ma), Basidiomycota-Ascomycota (968 Ma), Mucorales-Basidiomycota (947 Ma), Fungi-Animalia (1,513 Ma), mosses-vascular plants (707 Ma), Chlorophyta-Tracheophyta (968 Ma), Rhodophyta-Chlorophyta+Embryophyta (1,428 Ma), Plantae-Animalia (1,609 Ma), Alveolata-plants+animals+fungi (1,973 Ma), Euglenozoa-plants+animals+fungi (1,961 Ma), and Giardia-plants+animals+fungi (2,309 Ma). By extrapolation, mitochondria arose approximately 2300-1800 Ma and plastids arose 1600-1500 Ma. Estimates of the maximum number of cell types of common ancestors, combined with divergence times, showed an increase from two cell types at 2500 Ma to

  14. A molecular timescale of eukaryote evolution and the rise of complex multicellular life

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    Venturi Maria L

    2004-01-01

    Full Text Available Abstract Background The pattern and timing of the rise in complex multicellular life during Earth's history has not been established. Great disparity persists between the pattern suggested by the fossil record and that estimated by molecular clocks, especially for plants, animals, fungi, and the deepest branches of the eukaryote tree. Here, we used all available protein sequence data and molecular clock methods to place constraints on the increase in complexity through time. Results Our phylogenetic analyses revealed that (i animals are more closely related to fungi than to plants, (ii red algae are closer to plants than to animals or fungi, (iii choanoflagellates are closer to animals than to fungi or plants, (iv diplomonads, euglenozoans, and alveolates each are basal to plants+animals+fungi, and (v diplomonads are basal to other eukaryotes (including alveolates and euglenozoans. Divergence times were estimated from global and local clock methods using 20–188 proteins per node, with data treated separately (multigene and concatenated (supergene. Different time estimation methods yielded similar results (within 5%: vertebrate-arthropod (964 million years ago, Ma, Cnidaria-Bilateria (1,298 Ma, Porifera-Eumetozoa (1,351 Ma, Pyrenomycetes-Plectomycetes (551 Ma, Candida-Saccharomyces (723 Ma, Hemiascomycetes-filamentous Ascomycota (982 Ma, Basidiomycota-Ascomycota (968 Ma, Mucorales-Basidiomycota (947 Ma, Fungi-Animalia (1,513 Ma, mosses-vascular plants (707 Ma, Chlorophyta-Tracheophyta (968 Ma, Rhodophyta-Chlorophyta+Embryophyta (1,428 Ma, Plantae-Animalia (1,609 Ma, Alveolata-plants+animals+fungi (1,973 Ma, Euglenozoa-plants+animals+fungi (1,961 Ma, and Giardia-plants+animals+fungi (2,309 Ma. By extrapolation, mitochondria arose approximately 2300-1800 Ma and plastids arose 1600-1500 Ma. Estimates of the maximum number of cell types of common ancestors, combined with divergence times, showed an increase from two cell types at 2500 Ma to ~10

  15. Patterns of Transcript Abundance of Eukaryotic Biogeochemically-Relevant Genes in the Amazon River Plume.

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    Brian L Zielinski

    Full Text Available The Amazon River has the largest discharge of all rivers on Earth, and its complex plume system fuels a wide array of biogeochemical processes, across a large area of the western tropical North Atlantic. The plume thus stimulates microbial processes affecting carbon sequestration and nutrient cycles at a global scale. Chromosomal gene expression patterns of the 2.0 to 156 μm size-fraction eukaryotic microbial community were investigated in the Amazon River Plume, generating a robust dataset (more than 100 million mRNA sequences that depicts the metabolic capabilities and interactions among the eukaryotic microbes. Combining classical oceanographic field measurements with metatranscriptomics yielded characterization of the hydrographic conditions simultaneous with a quantification of transcriptional activity and identity of the community. We highlight the patterns of eukaryotic gene expression for 31 biogeochemically significant gene targets hypothesized to be valuable within forecasting models. An advantage to this targeted approach is that the database of reference sequences used to identify the target genes was selectively constructed and highly curated optimizing taxonomic coverage, throughput, and the accuracy of annotations. A coastal diatom bloom highly expressed nitrate transporters and carbonic anhydrase presumably to support high growth rates and enhance uptake of low levels of dissolved nitrate and CO2. Diatom-diazotroph association (DDA: diatoms with nitrogen fixing symbionts blooms were common when surface salinity was mesohaline and dissolved nitrate concentrations were below detection, and hence did not show evidence of nitrate utilization, suggesting they relied on ammonium transporters to aquire recently fixed nitrogen. These DDA blooms in the outer plume had rapid turnover of the photosystem D1 protein presumably caused by photodegradation under increased light penetration in clearer waters, and increased expression of silicon

  16. Regulation of eukaryotic elongation factor 1 alpha (eEF1A) by dynamic lysine methylation

    DEFF Research Database (Denmark)

    Jakobsson, Magnus E; Małecki, Jędrzej; Falnes, Pål Ø

    2018-01-01

    Lysine methylation is a frequent post-translational protein modification, which has been intensively studied in the case of histone proteins. Lysine methylations are also found on many non-histone proteins, and one prominent example is eukaryotic elongation factor 1 alpha (eEF1A). Besides its...... essential role in the protein synthesis machinery, a number of non-canonical functions have also been described for eEF1A, such as regulation of the actin cytoskeleton and the promotion of viral replication. The functional significance of the extensive lysine methylations on eEF1A, as well as the identity...

  17. Metatranscriptomics reveals the diversity of genes expressed by eukaryotes in forest soils.

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    Coralie Damon

    Full Text Available Eukaryotic organisms play essential roles in the biology and fertility of soils. For example the micro and mesofauna contribute to the fragmentation and homogenization of plant organic matter, while its hydrolysis is primarily performed by the fungi. To get a global picture of the activities carried out by soil eukaryotes we sequenced 2×10,000 cDNAs synthesized from polyadenylated mRNA directly extracted from soils sampled in beech (Fagus sylvatica and spruce (Picea abies forests. Taxonomic affiliation of both cDNAs and 18S rRNA sequences showed a dominance of sequences from fungi (up to 60% and metazoans while protists represented less than 12% of the 18S rRNA sequences. Sixty percent of cDNA sequences from beech forest soil and 52% from spruce forest soil had no homologs in the GenBank/EMBL/DDJB protein database. A Gene Ontology term was attributed to 39% and 31.5% of the spruce and beech soil sequences respectively. Altogether 2076 sequences were putative homologs to different enzyme classes participating to 129 KEGG pathways among which several were implicated in the utilisation of soil nutrients such as nitrogen (ammonium, amino acids, oligopeptides, sugars, phosphates and sulfate. Specific annotation of plant cell wall degrading enzymes identified enzymes active on major polymers (cellulose, hemicelluloses, pectin, lignin and glycoside hydrolases represented 0.5% (beech soil-0.8% (spruce soil of the cDNAs. Other sequences coding enzymes active on organic matter (extracellular proteases, lipases, a phytase, P450 monooxygenases were identified, thus underlining the biotechnological potential of eukaryotic metatranscriptomes. The phylogenetic affiliation of 12 full-length carbohydrate active enzymes showed that most of them were distantly related to sequences from known fungi. For example, a putative GH45 endocellulase was closely associated to molluscan sequences, while a GH7 cellobiohydrolase was closest to crustacean sequences, thus

  18. In vitro Assays for Eukaryotic Leading/Lagging Strand DNA Replication.

    Science.gov (United States)

    Schauer, Grant; Finkelstein, Jeff; O'Donnell, Mike

    2017-09-20

    The eukaryotic replisome is a multiprotein complex that duplicates DNA. The replisome is sculpted to couple continuous leading strand synthesis with discontinuous lagging strand synthesis, primarily carried out by DNA polymerases ε and δ, respectively, along with helicases, polymerase α-primase, DNA sliding clamps, clamp loaders and many other proteins. We have previously established the mechanisms by which the polymerases ε and δ are targeted to their 'correct' strands, as well as quality control mechanisms that evict polymerases when they associate with an 'incorrect' strand. Here, we provide a practical guide to differentially assay leading and lagging strand replication in vitro using pure proteins.

  19. Strong growth for Queensland mining

    Energy Technology Data Exchange (ETDEWEB)

    1990-10-01

    The Queensland mining industry experienced strong growth during 1989-90 as shown in the latest statistics released by the Department of Resource Industries. The total value of Queensland mineral and energy production rose to a new record of $5.1 billion, an increase of 16.5% on 1988-89 production. A major contributing factor was a 20.9 percent increase in the value of coal production. While the quantity of coal produced rose only 1.1 percent, the substantial increase in the value of coal production is attributable to higher coal prices negotiated for export contracts. In Australian dollar terms coal, gold, lead, zinc and crude oil on average experienced higher international prices than in the previous year. Only copper and silver prices declined. 3 tabs.

  20. Strong moduli stabilization and phenomenology

    CERN Document Server

    Dudas, Emilian; Mambrini, Yann; Mustafayev, Azar; Olive, Keith A

    2013-01-01

    We describe the resulting phenomenology of string theory/supergravity models with strong moduli stabilization. The KL model with F-term uplifting, is one such example. Models of this type predict universal scalar masses equal to the gravitino mass. In contrast, A-terms receive highly suppressed gravity mediated contributions. Under certain conditions, the same conclusion is valid for gaugino masses, which like A-terms, are then determined by anomalies. In such models, we are forced to relatively large gravitino masses (30-1000 TeV). We compute the low energy spectrum as a function of m_{3/2}. We see that the Higgs masses naturally takes values between 125-130 GeV. The lower limit is obtained from the requirement of chargino masses greater than 104 GeV, while the upper limit is determined by the relic density of dark matter (wino-like).

  1. Strongly interacting W's and Z's

    International Nuclear Information System (INIS)

    Gaillard, M.K.

    1984-01-01

    The study focussed primarily on the dynamics of a strongly interacting W, Z(SIW) sector, with the aim of sharpening predictions for total W, Z yield and W, Z multiplicities expected from WW fusion for various scenarios. Specific issues raised in the context of the general problem of modeling SIW included the specificity of the technicolor (or, equivalently, QCD) model, whether or not a composite scalar model can be evaded, and whether the standard model necessarily implies an I = J = O state (≅ Higgs particle) that is relatively ''light'' (M ≤ hundreds of TeV). The consensus on the last issue was that existing arguments are inconclusive. While the author shall briefly address compositeness and alternatives to the technicolor model, quantitative estimates will be of necessity based on technicolor or an extrapolation of pion data

  2. Uniquely Strongly Clean Group Rings

    Institute of Scientific and Technical Information of China (English)

    WANG XIU-LAN

    2012-01-01

    A ring R is called clean if every element is the sum of an idempotent and a unit,and R is called uniquely strongly clean (USC for short) if every element is uniquely the sum of an idempotent and a unit that commute.In this article,some conditions on a ring R and a group G such that RG is clean are given.It is also shown that if G is a locally finite group,then the group ring RG is USC if and only if R is USC,and G is a 2-group.The left uniquely exchange group ring,as a middle ring of the uniquely clean ring and the USC ring,does not possess this property,and so does the uniquely exchange group ring.

  3. Electrophoresis in strong electric fields.

    Science.gov (United States)

    Barany, Sandor

    2009-01-01

    Two kinds of non-linear electrophoresis (ef) that can be detected in strong electric fields (several hundred V/cm) are considered. The first ("classical" non-linear ef) is due to the interaction of the outer field with field-induced ionic charges in the electric double layer (EDL) under conditions, when field-induced variations of electrolyte concentration remain to be small comparatively to its equilibrium value. According to the Shilov theory, the non-linear component of the electrophoretic velocity for dielectric particles is proportional to the cubic power of the applied field strength (cubic electrophoresis) and to the second power of the particles radius; it is independent of the zeta-potential but is determined by the surface conductivity of particles. The second one, the so-called "superfast electrophoresis" is connected with the interaction of a strong outer field with a secondary diffuse layer of counterions (space charge) that is induced outside the primary (classical) diffuse EDL by the external field itself because of concentration polarization. The Dukhin-Mishchuk theory of "superfast electrophoresis" predicts quadratic dependence of the electrophoretic velocity of unipolar (ionically or electronically) conducting particles on the external field gradient and linear dependence on the particle's size in strong electric fields. These are in sharp contrast to the laws of classical electrophoresis (no dependence of V(ef) on the particle's size and linear dependence on the electric field gradient). A new method to measure the ef velocity of particles in strong electric fields is developed that is based on separation of the effects of sedimentation and electrophoresis using videoimaging and a new flowcell and use of short electric pulses. To test the "classical" non-linear electrophoresis, we have measured the ef velocity of non-conducting polystyrene, aluminium-oxide and (semiconductor) graphite particles as well as Saccharomice cerevisiae yeast cells as a

  4. Evidence of pervasive biologically functional secondary structures within the genomes of eukaryotic single-stranded DNA viruses.

    Science.gov (United States)

    Muhire, Brejnev Muhizi; Golden, Michael; Murrell, Ben; Lefeuvre, Pierre; Lett, Jean-Michel; Gray, Alistair; Poon, Art Y F; Ngandu, Nobubelo Kwanele; Semegni, Yves; Tanov, Emil Pavlov; Monjane, Adérito Luis; Harkins, Gordon William; Varsani, Arvind; Shepherd, Dionne Natalie; Martin, Darren Patrick

    2014-02-01

    Single-stranded DNA (ssDNA) viruses have genomes that are potentially capable of forming complex secondary structures through Watson-Crick base pairing between their constituent nucleotides. A few of the structural elements formed by such base pairings are, in fact, known to have important functions during the replication of many ssDNA viruses. Unknown, however, are (i) whether numerous additional ssDNA virus genomic structural elements predicted to exist by computational DNA folding methods actually exist and (ii) whether those structures that do exist have any biological relevance. We therefore computationally inferred lists of the most evolutionarily conserved structures within a diverse selection of animal- and plant-infecting ssDNA viruses drawn from the families Circoviridae, Anelloviridae, Parvoviridae, Nanoviridae, and Geminiviridae and analyzed these for evidence of natural selection favoring the maintenance of these structures. While we find evidence that is consistent with purifying selection being stronger at nucleotide sites that are predicted to be base paired than at sites predicted to be unpaired, we also find strong associations between sites that are predicted to pair with one another and site pairs that are apparently coevolving in a complementary fashion. Collectively, these results indicate that natural selection actively preserves much of the pervasive secondary structure that is evident within eukaryote-infecting ssDNA virus genomes and, therefore, that much of this structure is biologically functional. Lastly, we provide examples of various highly conserved but completely uncharacterized structural elements that likely have important functions within some of the ssDNA virus genomes analyzed here.

  5. The conformational stability and biophysical properties of the eukaryotic thioredoxins of Pisum sativum are not family-conserved.

    Directory of Open Access Journals (Sweden)

    David Aguado-Llera

    2011-02-01

    Full Text Available Thioredoxins (TRXs are ubiquitous proteins involved in redox processes. About forty genes encode TRX or TRX-related proteins in plants, grouped in different families according to their subcellular localization. For instance, the h-type TRXs are located in cytoplasm or mitochondria, whereas f-type TRXs have a plastidial origin, although both types of proteins have an eukaryotic origin as opposed to other TRXs. Herein, we study the conformational and the biophysical features of TRXh1, TRXh2 and TRXf from Pisum sativum. The modelled structures of the three proteins show the well-known TRX fold. While sharing similar pH-denaturations features, the chemical and thermal stabilities are different, being PsTRXh1 (Pisum sativum thioredoxin h1 the most stable isoform; moreover, the three proteins follow a three-state denaturation model, during the chemical-denaturations. These differences in the thermal- and chemical-denaturations result from changes, in a broad sense, of the several ASAs (accessible surface areas of the proteins. Thus, although a strong relationship can be found between the primary amino acid sequence and the structure among TRXs, that between the residue sequence and the conformational stability and biophysical properties is not. We discuss how these differences in the biophysical properties of TRXs determine their unique functions in pea, and we show how residues involved in the biophysical features described (pH-titrations, dimerizations and chemical-denaturations belong to regions involved in interaction with other proteins. Our results suggest that the sequence demands of protein-protein function are relatively rigid, with different protein-binding pockets (some in common for each of the three proteins, but the demands of structure and conformational stability per se (as long as there is a maintained core, are less so.

  6. Direct ubiquitin independent recognition and degradation of a folded protein by the eukaryotic proteasomes-origin of intrinsic degradation signals.

    Directory of Open Access Journals (Sweden)

    Amit Kumar Singh Gautam

    Full Text Available Eukaryotic 26S proteasomes are structurally organized to recognize, unfold and degrade globular proteins. However, all existing model substrates of the 26S proteasome in addition to ubiquitin or adaptor proteins require unstructured regions in the form of fusion tags for efficient degradation. We report for the first time that purified 26S proteasome can directly recognize and degrade apomyoglobin, a globular protein, in the absence of ubiquitin, extrinsic degradation tags or adaptor proteins. Despite a high affinity interaction, absence of a ligand and presence of only helices/loops that follow the degradation signal, apomyoglobin is degraded slowly by the proteasome. A short floppy F-helix exposed upon ligand removal and in conformational equilibrium with a disordered structure is mandatory for recognition and initiation of degradation. Holomyoglobin, in which the helix is buried, is neither recognized nor degraded. Exposure of the floppy F-helix seems to sensitize the proteasome and primes the substrate for degradation. Using peptide panning and competition experiments we speculate that initial encounters through the floppy helix and additional strong interactions with N-terminal helices anchors apomyoglobin to the proteasome. Stabilizing helical structure in the floppy F-helix slows down degradation. Destabilization of adjacent helices accelerates degradation. Unfolding seems to follow the mechanism of helix unraveling rather than global unfolding. Our findings while confirming the requirement for unstructured regions in degradation offers the following new insights: a origin and identification of an intrinsic degradation signal in the substrate, b identification of sequences in the native substrate that are likely to be responsible for direct interactions with the proteasome, and c identification of critical rate limiting steps like exposure of the intrinsic degron and destabilization of an unfolding intermediate that are presumably

  7. [Construction and functional identification of eukaryotic expression vector carrying Sprague-Dawley rat MSX-2 gene].

    Science.gov (United States)

    Yang, Xian-Xian; Zhang, Mei; Yan, Zhao-Wen; Zhang, Ru-Hong; Mu, Xiong-Zheng

    2008-01-01

    To construct a high effective eukaryotic expressing plasmid PcDNA 3.1-MSX-2 encoding Sprague-Dawley rat MSX-2 gene for the further study of MSX-2 gene function. The full length SD rat MSX-2 gene was amplified by PCR, and the full length DNA was inserted in the PMD1 8-T vector. It was isolated by restriction enzyme digest with BamHI and Xhol, then ligated into the cloning site of the PcDNA3.1 expression plasmid. The positive recombinant was identified by PCR analysis, restriction endonudease analysis and sequence analysis. Expression of RNA and protein was detected by RT-PCR and Western blot analysis in PcDNA3.1-MSX-2 transfected HEK293 cells. Sequence analysis and restriction endonudease analysis of PcDNA3.1-MSX-2 demonstrated that the position and size of MSX-2 cDNA insertion were consistent with the design. RT-PCR and Western blot analysis showed specific expression of mRNA and protein of MSX-2 in the transfected HEK293 cells. The high effective eukaryotic expression plasmid PcDNA3.1-MSX-2 encoding Sprague-Dawley Rat MSX-2 gene which is related to craniofacial development can be successfully reconstructed. It may serve as the basis for the further study of MSX-2 gene function.

  8. The origin and evolution of the sexes: Novel insights from a distant eukaryotic linage.

    Science.gov (United States)

    Mignerot, Laure; Coelho, Susana M

    2016-01-01

    Sexual reproduction is an extraordinarily widespread phenomenon that assures the production of new genetic combinations in nearly all eukaryotic lineages. Although the core features of sexual reproduction (meiosis and syngamy) are highly conserved, the control mechanisms that determine whether an individual is male or female are remarkably labile across eukaryotes. In genetically controlled sexual systems, gender is determined by sex chromosomes, which have emerged independently and repeatedly during evolution. Sex chromosomes have been studied in only a handful of classical model organism, and empirical knowledge on the origin and evolution of the sexes is still surprisingly incomplete. With the advent of new generation sequencing, the taxonomic breadth of model systems has been rapidly expanding, bringing new ideas and fresh views on this fundamental aspect of biology. This mini-review provides a quick state of the art of how the remarkable richness of the sexual characteristics of the brown algae is helping to increase our knowledge about the evolution of sex determination. Copyright © 2016 Académie des sciences. Published by Elsevier SAS. All rights reserved.

  9. Eukaryotic tRNAs fingerprint invertebrates vis-à-vis vertebrates.

    Science.gov (United States)

    Mitra, Sanga; Das, Pijush; Samadder, Arpa; Das, Smarajit; Betai, Rupal; Chakrabarti, Jayprokas

    2015-01-01

    During translation, aminoacyl-tRNA synthetases recognize the identities of the tRNAs to charge them with their respective amino acids. The conserved identities of 58,244 eukaryotic tRNAs of 24 invertebrates and 45 vertebrates in genomic tRNA database were analyzed and their novel features extracted. The internal promoter sequences, namely, A-Box and B-Box, were investigated and evidence gathered that the intervention of optional nucleotides at 17a and 17b correlated with the optimal length of the A-Box. The presence of canonical transcription terminator sequences at the immediate vicinity of tRNA genes was ventured. Even though non-canonical introns had been reported in red alga, green alga, and nucleomorph so far, fairly motivating evidence of their existence emerged in tRNA genes of other eukaryotes. Non-canonical introns were seen to interfere with the internal promoters in two cases, questioning their transcription fidelity. In a first of its kind, phylogenetic constructs based on tRNA molecules delineated and built the trees of the vast and diverse invertebrates and vertebrates. Finally, two tRNA models representing the invertebrates and the vertebrates were drawn, by isolating the dominant consensus in the positional fluctuations of nucleotide compositions.

  10. Deep sequencing of subseafloor eukaryotic rRNA reveals active Fungi across marine subsurface provinces.

    Directory of Open Access Journals (Sweden)

    William Orsi

    Full Text Available The deep marine subsurface is a vast habitat for microbial life where cells may live on geologic timescales. Because DNA in sediments may be preserved on long timescales, ribosomal RNA (rRNA is suggested to be a proxy for the active fraction of a microbial community in the subsurface. During an investigation of eukaryotic 18S rRNA by amplicon pyrosequencing, unique profiles of Fungi were found across a range of marine subsurface provinces including ridge flanks, continental margins, and abyssal plains. Subseafloor fungal populations exhibit statistically significant correlations with total organic carbon (TOC, nitrate, sulfide, and dissolved inorganic carbon (DIC. These correlations are supported by terminal restriction length polymorphism (TRFLP analyses of fungal rRNA. Geochemical correlations with fungal pyrosequencing and TRFLP data from this geographically broad sample set suggests environmental selection of active Fungi in the marine subsurface. Within the same dataset, ancient rRNA signatures were recovered from plants and diatoms in marine sediments ranging from 0.03 to 2.7 million years old, suggesting that rRNA from some eukaryotic taxa may be much more stable than previously considered in the marine subsurface.

  11. Effects of Saponins against Clinical E. coli Strains and Eukaryotic Cell Line

    Science.gov (United States)

    Arabski, Michał; Węgierek-Ciuk, Aneta; Czerwonka, Grzegorz; Lankoff, Anna; Kaca, Wiesław

    2012-01-01

    Saponins are detergent-like substances showing antibacterial as well as anticancer potential. In this study, the effects of saponins from Quillaja saponaria were analyzed against prokaryotic and eukaryotic cells. Multidrug-resistant clinical E. coli strains were isolated from human urine. As eukaryotic cells, the CHO-K1 cell lines were applied. Antibacterial effect of ampicillin, streptomycin, and ciprofloxacin in the presence of saponins was measured by cultivation methods. Properties of saponins against CHO-K1 cells were measured by the MTT test, hemolysis assay and flow cytometry. Saponin from Quillaja saponaria has a cytotoxic effect at concentrations higher than 25 μg/mL and in the range of 12–50 μg/mL significantly increases the level of early apoptotic cells. Saponin at dose of 12 μg/mL enhances the six E. coli strains growth. We postulate that saponins increase the influx of nutrients from the medium into E. coli cells. Saponins do not have synergetic effects on antibacterial action of tested antibiotics. In contrary, in the presence of saponins and antibiotics, more CFU/mL E. coli cells were observed. This effect was similar to saponins action alone towards E. coli cells. In conclusion, saponins was cytotoxic against CHO-K1 cells, whereas against E. coli cells this effect was not observed. PMID:22500084

  12. The genome of the polar eukaryotic microalga Coccomyxa subellipsoidea reveals traits of cold adaptation

    Energy Technology Data Exchange (ETDEWEB)

    Blanc, Guillaume; Agarkova, Irina; Grimwood, Jane; Kuo, Alan; Brueggeman, Andrew; Dunigan, David D.; Gurnon, James; Ladunga, Istvan; Lindquist, Erika; Lucas, Susan; Pangilinan, Jasmyn; Proschold, Thomas; Salamov, Asaf; Schmutz, Jeremy; Weeks, Donald; Tamada, Takashi; Lomsadze, Alexandre; Borodovsky, Mark; Claverie, Jean-Michel; Grigoriev, Igor V.; Van Etten, James L.

    2012-02-13

    Background Little is known about the mechanisms of adaptation of life to the extreme environmental conditions encountered in polar regions. Here we present the genome sequence of a unicellular green alga from the division chlorophyta, Coccomyxa subellipsoidea C-169, which we will hereafter refer to as C-169. This is the first eukaryotic microorganism from a polar environment to have its genome sequenced. Results The 48.8 Mb genome contained in 20 chromosomes exhibits significant synteny conservation with the chromosomes of its relatives Chlorella variabilis and Chlamydomonas reinhardtii. The order of the genes is highly reshuffled within synteny blocks, suggesting that intra-chromosomal rearrangements were more prevalent than inter-chromosomal rearrangements. Remarkably, Zepp retrotransposons occur in clusters of nested elements with strictly one cluster per chromosome probably residing at the centromere. Several protein families overrepresented in C. subellipsoidae include proteins involved in lipid metabolism, transporters, cellulose synthases and short alcohol dehydrogenases. Conversely, C-169 lacks proteins that exist in all other sequenced chlorophytes, including components of the glycosyl phosphatidyl inositol anchoring system, pyruvate phosphate dikinase and the photosystem 1 reaction center subunit N (PsaN). Conclusions We suggest that some of these gene losses and gains could have contributed to adaptation to low temperatures. Comparison of these genomic features with the adaptive strategies of psychrophilic microbes suggests that prokaryotes and eukaryotes followed comparable evolutionary routes to adapt to cold environments.

  13. Variation in recombination frequency and distribution across eukaryotes: patterns and processes

    Science.gov (United States)

    Feulner, Philine G. D.; Johnston, Susan E.; Santure, Anna W.; Smadja, Carole M.

    2017-01-01

    Recombination, the exchange of DNA between maternal and paternal chromosomes during meiosis, is an essential feature of sexual reproduction in nearly all multicellular organisms. While the role of recombination in the evolution of sex has received theoretical and empirical attention, less is known about how recombination rate itself evolves and what influence this has on evolutionary processes within sexually reproducing organisms. Here, we explore the patterns of, and processes governing recombination in eukaryotes. We summarize patterns of variation, integrating current knowledge with an analysis of linkage map data in 353 organisms. We then discuss proximate and ultimate processes governing recombination rate variation and consider how these influence evolutionary processes. Genome-wide recombination rates (cM/Mb) can vary more than tenfold across eukaryotes, and there is large variation in the distribution of recombination events across closely related taxa, populations and individuals. We discuss how variation in rate and distribution relates to genome architecture, genetic and epigenetic mechanisms, sex, environmental perturbations and variable selective pressures. There has been great progress in determining the molecular mechanisms governing recombination, and with the continued development of new modelling and empirical approaches, there is now also great opportunity to further our understanding of how and why recombination rate varies. This article is part of the themed issue ‘Evolutionary causes and consequences of recombination rate variation in sexual organisms’. PMID:29109219

  14. Novel core promoter elements and a cognate transcription factor in the divergent unicellular eukaryote Trichomonas vaginalis.

    Science.gov (United States)

    Smith, Alias J; Chudnovsky, Lorissa; Simoes-Barbosa, Augusto; Delgadillo-Correa, Maria G; Jonsson, Zophonias O; Wohlschlegel, James A; Johnson, Patricia J

    2011-04-01

    A highly conserved DNA initiator (Inr) element has been the only core promoter element described in the divergent unicellular eukaryote Trichomonas vaginalis, although genome analyses reveal that only ∼75% of protein-coding genes appear to contain an Inr. In search of another core promoter element(s), a nonredundant database containing 5' untranslated regions of expressed T. vaginalis genes was searched for overrepresented DNA motifs and known eukaryotic core promoter elements. In addition to identifying the Inr, two elements that lack sequence similarity to the known protein-coding gene core promoter, motif 3 (M3) and motif 5 (M5), were identified. Mutational and functional analyses demonstrate that both are novel core promoter elements. M3 [(A/G/T)(A/G)C(G/C)G(T/C)T(T/A/G)] resembles a Myb recognition element (MRE) and is bound specifically by a unique protein with a Myb-like DNA binding domain. The M5 element (CCTTT) overlaps the transcription start site and replaces the Inr as an alternative, gene-specific initiator element. Transcription specifically initiates at the second cytosine within M5, in contrast to characteristic initiation by RNA polymerase II at an adenosine. In promoters that combine M3 with either M5 or Inr, transcription initiation is regulated by the M3 motif.

  15. Novel Core Promoter Elements and a Cognate Transcription Factor in the Divergent Unicellular Eukaryote Trichomonas vaginalis▿

    Science.gov (United States)

    Smith, Alias J.; Chudnovsky, Lorissa; Simoes-Barbosa, Augusto; Delgadillo-Correa, Maria G.; Jonsson, Zophonias O.; Wohlschlegel, James A.; Johnson, Patricia J.

    2011-01-01

    A highly conserved DNA initiator (Inr) element has been the only core promoter element described in the divergent unicellular eukaryote Trichomonas vaginalis, although genome analyses reveal that only ∼75% of protein-coding genes appear to contain an Inr. In search of another core promoter element(s), a nonredundant database containing 5′ untranslated regions of expressed T. vaginalis genes was searched for overrepresented DNA motifs and known eukaryotic core promoter elements. In addition to identifying the Inr, two elements that lack sequence similarity to the known protein-coding gene core promoter, motif 3 (M3) and motif 5 (M5), were identified. Mutational and functional analyses demonstrate that both are novel core promoter elements. M3 [(A/G/T)(A/G)C(G/C)G(T/C)T(T/A/G)] resembles a Myb recognition element (MRE) and is bound specifically by a unique protein with a Myb-like DNA binding domain. The M5 element (CCTTT) overlaps the transcription start site and replaces the Inr as an alternative, gene-specific initiator element. Transcription specifically initiates at the second cytosine within M5, in contrast to characteristic initiation by RNA polymerase II at an adenosine. In promoters that combine M3 with either M5 or Inr, transcription initiation is regulated by the M3 motif. PMID:21245378

  16. Similarities and Differences in the Glycosylation Mechanisms in Prokaryotes and Eukaryotes

    Directory of Open Access Journals (Sweden)

    Anne Dell

    2010-01-01

    Full Text Available Recent years have witnessed a rapid growth in the number and diversity of prokaryotic proteins shown to carry N- and/or O-glycans, with protein glycosylation now considered as fundamental to the biology of these organisms as it is in eukaryotic systems. This article overviews the major glycosylation pathways that are known to exist in eukarya, bacteria and archaea. These are (i oligosaccharyltransferase (OST-mediated N-glycosylation which is abundant in eukarya and archaea, but is restricted to a limited range of bacteria; (ii stepwise cytoplasmic N-glycosylation that has so far only been confirmed in the bacterial domain; (iii OST-mediated O-glycosylation which appears to be characteristic of bacteria; and (iv stepwise O-glycosylation which is common in eukarya and bacteria. A key aim of the review is to integrate information from the three domains of life in order to highlight commonalities in glycosylation processes. We show how the OST-mediated N- and O-glycosylation pathways share cytoplasmic assembly of lipid-linked oligosaccharides, flipping across the ER/periplasmic/cytoplasmic membranes, and transferring “en bloc” to the protein acceptor. Moreover these hallmarks are mirrored in lipopolysaccharide biosynthesis. Like in eukaryotes, stepwise O-glycosylation occurs on diverse bacterial proteins including flagellins, adhesins, autotransporters and lipoproteins, with O-glycosylation chain extension often coupled with secretory mechanisms.

  17. Biotransformation of mercury in pH-stat cultures of eukaryotic freshwater algae.

    Science.gov (United States)

    Kelly, David J A; Budd, Kenneth; Lefebvre, Daniel D

    2007-01-01

    Eukaryotic algae were studied to determine their ability to biotransform Hg(II) under aerated and pH controlled conditions. All algae converted Hg(II) into beta-HgS and Hg(0) to varying degrees. When Hg(II) was administered as HgCl(2) to the algae, biotransformation by species of Chlorophyceae (Selenastrum minutum and Chlorella fusca var. fusca) was initiated with beta-HgS synthesis (K (1/2) of hours) and concomitant Hg degrees evolution occurred in the first hour. Hg degrees synthesis was impeded by the formation of beta-HgS and this inhibition was released in C. fusca var. fusca when cellular thiols were oxidized by the addition of dimethylfumarate (DMF). The diatom, Navicula pelliculosa (Bacillariophyceae), converted a substantially greater proportion of the applied Hg(II) into Hg(0), whereas the thermophilic alga, Galdieria sulphuraria (Cyanidiophyceae), rapidly biotransformed as much as 90% of applied Hg(II) into beta-HgS (K (1/2) approximately 20 min). This thermophile was also able to generate Hg(0) even after all exogenously applied HgCl(2) had been biotransformed. The results suggest that beta-HgS may be the major dietary mercurial for grazers of contaminated eukaryotic algae.

  18. A Common Ca2+-Driven Interdomain Module Governs Eukaryotic NCX Regulation

    Science.gov (United States)

    Giladi, Moshe; Sasson, Yehezkel; Fang, Xianyang; Hiller, Reuben; Buki, Tal; Wang, Yun-Xing; Hirsch, Joel A.; Khananshvili, Daniel

    2012-01-01

    Na+/Ca2+ exchanger (NCX) proteins mediate Ca2+-fluxes across the cell membrane to maintain Ca2+ homeostasis in many cell types. Eukaryotic NCX contains Ca2+-binding regulatory domains, CBD1 and CBD2. Ca2+ binding to a primary sensor (Ca3-Ca4 sites) on CBD1 activates mammalian NCXs, whereas CALX, a Drosophila NCX ortholog, displays an inhibitory response to regulatory Ca2+. To further elucidate the underlying regulatory mechanisms, we determined the 2.7 Å crystal structure of mammalian CBD12-E454K, a two-domain construct that retains wild-type properties. In conjunction with stopped-flow kinetics and SAXS (small-angle X-ray scattering) analyses of CBD12 mutants, we show that Ca2+ binding to Ca3-Ca4 sites tethers the domains via a network of interdomain salt-bridges. This Ca2+-driven interdomain switch controls slow dissociation of “occluded” Ca2+ from the primary sensor and thus dictates Ca2+ sensing dynamics. In the Ca2+-bound conformation, the interdomain angle of CBD12 is very similar in NCX and CALX, meaning that the interdomain distances cannot account for regulatory diversity in NCX and CALX. Since the two-domain interface is nearly identical among eukaryotic NCXs, including CALX, we suggest that the Ca2+-driven interdomain switch described here represents a general mechanism for initial conduction of regulatory signals in NCX variants. PMID:22768191

  19. EKPD: a hierarchical database of eukaryotic protein kinases and protein phosphatases.

    Science.gov (United States)

    Wang, Yongbo; Liu, Zexian; Cheng, Han; Gao, Tianshun; Pan, Zhicheng; Yang, Qing; Guo, Anyuan; Xue, Yu

    2014-01-01

    We present here EKPD (http://ekpd.biocuckoo.org), a hierarchical database of eukaryotic protein kinases (PKs) and protein phosphatases (PPs), the key molecules responsible for the reversible phosphorylation of proteins that are involved in almost all aspects of biological processes. As extensive experimental and computational efforts have been carried out to identify PKs and PPs, an integrative resource with detailed classification and annotation information would be of great value for both experimentalists and computational biologists. In this work, we first collected 1855 PKs and 347 PPs from the scientific literature and various public databases. Based on previously established rationales, we classified all of the known PKs and PPs into a hierarchical structure with three levels, i.e. group, family and individual PK/PP. There are 10 groups with 149 families for the PKs and 10 groups with 33 families for the PPs. We constructed 139 and 27 Hidden Markov Model profiles for PK and PP families, respectively. Then we systematically characterized ∼50,000 PKs and >10,000 PPs in eukaryotes. In addition, >500 PKs and >400 PPs were computationally identified by ortholog search. Finally, the online service of the EKPD database was implemented in PHP + MySQL + JavaScript.

  20. Effects of Saponins against Clinical E. coli Strains and Eukaryotic Cell Line

    Directory of Open Access Journals (Sweden)

    Michał Arabski

    2012-01-01

    Full Text Available Saponins are detergent-like substances showing antibacterial as well as anticancer potential. In this study, the effects of saponins from Quillaja saponaria were analyzed against prokaryotic and eukaryotic cells. Multidrug-resistant clinical E. coli strains were isolated from human urine. As eukaryotic cells, the CHO-K1 cell lines were applied. Antibacterial effect of ampicillin, streptomycin, and ciprofloxacin in the presence of saponins was measured by cultivation methods. Properties of saponins against CHO-K1 cells were measured by the MTT test, hemolysis assay and flow cytometry. Saponin from Quillaja saponaria has a cytotoxic effect at concentrations higher than 25 μg/mL and in the range of 12–50 μg/mL significantly increases the level of early apoptotic cells. Saponin at dose of 12 μg/mL enhances the six E. coli strains growth. We postulate that saponins increase the influx of nutrients from the medium into E. coli cells. Saponins do not have synergetic effects on antibacterial action of tested antibiotics. In contrary, in the presence of saponins and antibiotics, more CFU/mL E. coli cells were observed. This effect was similar to saponins action alone towards E. coli cells. In conclusion, saponins was cytotoxic against CHO-K1 cells, whereas against E. coli cells this effect was not observed.

  1. Resilience of freshwater communities of small microbial eukaryotes undergoing severe drought events

    Directory of Open Access Journals (Sweden)

    Marianne eSimon

    2016-05-01

    Full Text Available Small and shallow aquatic ecosystems such as ponds and streams constitute a significant proportion of continental surface waters, especially in temperate zones. In comparison with bigger lakes and rivers, they harbor higher biodiversity but they also exhibit reduced buffering capacity face to environmental shifts, such that climate global change can affect them in a more drastic way. For instance, many temperate areas are predicted to undergo droughts with increasing frequency in the near future, which may lead to the temporal desiccation of streams and ponds. In this work, we monitored temporal dynamics of planktonic communities of microbial eukaryotes (cell size range 0.2-5 µm in one brook and one pond that experienced recurrent droughts from 1 to 5 consecutive months during a temporal survey carried out monthly for two years based on high-throughput 18S rDNA metabarcoding. During drought-induced desiccation events, protist communities present in the remaining dry sediment, though highly diverse, differed radically from their planktonic counterparts. However, after water refill, the aquatic protist assemblages recovered their original structure within a month. This rapid recovery indicates that these eukaryotic communities are resilient to droughts, most likely via the entrance in dormancy. This property is essential for the long-term survival and functional stability of small freshwater ecosystems.

  2. [Eukaryotic Expression and Immunogenic Research of Recombination Ebola Virus Membrane Protein Gp-Fc].

    Science.gov (United States)

    Zhang, Xiaoguang; Yang, Ren; Wang, Jiao; Wang, Xuan; Hou, Mieling; An, Lina; Zhu, Ying; Cao, Yuxi; Zeng, Yi

    2016-01-01

    We used 293 cells to express the recombinant membrane protein of the Ebola virus. Then, the immunogenicity of the recombinant protein was studied by immunized BALB/c mice. According to the codon use frequency of humans, the gene encoding the extracellular domain of the Ebola virus membrane protein was optimized, synthesized, and inserted into the eukaryotic expression plasmid pXG-Fc to construct the human IgG Fc and Ebola GP fusion protein expression plasmid pXG-modGP-Fc. To achieve expression, the fusion protein expression vector was transfected into high-density 293 cells using transient transfection technology. The recombinant protein was purified by protein A affinity chromatography. BALB/c mice were immunized with the purified fusion protein, and serum antibody titers evaluated by an indirect enzyme-linked immunosorbent assay (ELISA). Purification and analyses of the protein revealed that the eukaryotic expression vector could express the recombinant protein GP-Fc effectively, and that the recombinant protein in the supernatant of the cell culture was present as a dimer. After immunization with the purified recombinant protein, a high titer of antigen-specific IgG could be detected in the serum of immunized mice by indirect ELISA, showing that the recombinant protein had good immunogenicity. These data suggest that we obtained a recombinant protein with good immunogenicity. Our study is the basis for development of a vaccine against the Ebola virus and for screening of monoclonal antibodies.

  3. Protein N-myristoylation in Escherichia coli: Reconstitution of a eukaryotic protein modification in bacteria

    International Nuclear Information System (INIS)

    Duronio, R.J.; Jackson-Machelski, E.; Heuckeroth, R.O.; Gordon, J.I.; Olins, P.O.; Devine, C.S.; Yonemoto, W.; Slice, L.W.; Taylor, S.S.

    1990-01-01

    Protein N-myristoylation refers to the covalent attachment of a myristoyl group (C14:0), via amide linkage, to the NH 2 -terminal glycine residue of certain cellular and viral proteins. Myristoyl-CoA:protein N-myristoyltransferase (NMT) catalyzes this cotranslational modification. The authors have developed a system for studying the substrate requirements and biological effects of protein N-myristoylation as well as NMT structure-activity relationships. Expression of the yeast NMT1 gene in Escherichia coli, a bacterium that has no endogenous NMT activity, results in production of the intact 53-kDa NMT polypeptide as well as a truncated polypeptide derived from proteolytic removal of its NH 2 -terminal 39 amino acids. By using a dual plasmid system, N-myristoylation of a mammalian protein was reconstituted in E. coli by simultaneous expression of the yeast NMT1 gene and a murine cDNA encoding the catalytic (C) subunit of cAMP-dependent protein kinase (PK-A). A major advantage of the bacterial system over eukaryotic systems is the absence of endogenous NMT and substrates, providing a more straightforward way of preparing myristoylated, analog-substituted, and nonmyristoylated forms of a given protein for comparison of their structural and functional properties. The experimental system may prove useful for recapitulating other eukaryotic protein modifications in E. coli so that structure-activity relationships of modifying enzymes and their substrates can be more readily assessed

  4. Extracellular DNA amplicon sequencing reveals high levels of benthic eukaryotic diversity in the central Red Sea

    KAUST Repository

    Pearman, John K.

    2015-11-01

    The present study aims to characterize the benthic eukaryotic biodiversity patterns at a coarse taxonomic level in three areas of the central Red Sea (a lagoon, an offshore area in Thuwal and a shallow coastal area near Jeddah) based on extracellular DNA. High-throughput amplicon sequencing targeting the V9 region of the 18S rRNA gene was undertaken for 32 sediment samples. High levels of alpha-diversity were detected with 16,089 operational taxonomic units (OTUs) being identified. The majority of the OTUs were assigned to Metazoa (29.2%), Alveolata (22.4%) and Stramenopiles (17.8%). Stramenopiles (Diatomea) and Alveolata (Ciliophora) were frequent in a lagoon and in shallower coastal stations, whereas metazoans (Arthropoda: Maxillopoda) were dominant in deeper offshore stations. Only 24.6% of total OTUs were shared among all areas. Beta-diversity was generally lower between the lagoon and Jeddah (nearshore) than between either of those and the offshore area, suggesting a nearshore–offshore biodiversity gradient. The current approach allowed for a broad-range of benthic eukaryotic biodiversity to be analysed with significantly less labour than would be required by other traditional taxonomic approaches. Our findings suggest that next generation sequencing techniques have the potential to provide a fast and standardised screening of benthic biodiversity at large spatial and temporal scales.

  5. Evolutionary Inference across Eukaryotes Identifies Specific Pressures Favoring Mitochondrial Gene Retention.

    Science.gov (United States)

    Johnston, Iain G; Williams, Ben P

    2016-02-24

    Since their endosymbiotic origin, mitochondria have lost most of their genes. Although many selective mechanisms underlying the evolution of mitochondrial genomes have been proposed, a data-driven exploration of these hypotheses is lacking, and a quantitatively supported consensus remains absent. We developed HyperTraPS, a methodology coupling stochastic modeling with Bayesian inference, to identify the ordering of evolutionary events and suggest their causes. Using 2015 complete mitochondrial genomes, we inferred evolutionary trajectories of mtDNA gene loss across the eukaryotic tree of life. We find that proteins comprising the structural cores of the electron transport chain are preferentially encoded within mitochondrial genomes across eukaryotes. A combination of high GC content and high protein hydrophobicity is required to explain patterns of mtDNA gene retention; a model that accounts for these selective pressures can also predict the success of artificial gene transfer experiments in vivo. This work provides a general method for data-driven inference of the ordering of evolutionary and progressive events, here identifying the distinct features shaping mitochondrial genomes of present-day species. Copyright © 2016 Elsevier Inc. All rights reserved.

  6. Structure of the prolyl-tRNA synthetase from the eukaryotic pathogen Giardia lamblia

    Energy Technology Data Exchange (ETDEWEB)

    Larson, Eric T.; Kim, Jessica E.; Napuli, Alberto J.; Verlinde, Christophe L. M. J.; Fan, Erkang; Zucker, Frank H.; Van Voorhis, Wesley C.; Buckner, Frederick S.; Hol, Wim G. J.; Merritt, Ethan A., E-mail: merritt@u.washington.edu [Medical Structural Genomics of Pathogenic Protozoa, (United States); University of Washington, Seattle, WA 98195 (United States)

    2012-09-01

    The structure of Giardia prolyl-tRNA synthetase cocrystallized with proline and ATP shows evidence for half-of-the-sites activity, leading to a corresponding mixture of reaction substrates and product (prolyl-AMP) in the two active sites of the dimer. The genome of the human intestinal parasite Giardia lamblia contains only a single aminoacyl-tRNA synthetase gene for each amino acid. The Giardia prolyl-tRNA synthetase gene product was originally misidentified as a dual-specificity Pro/Cys enzyme, in part owing to its unexpectedly high off-target activation of cysteine, but is now believed to be a normal representative of the class of archaeal/eukaryotic prolyl-tRNA synthetases. The 2.2 Å resolution crystal structure of the G. lamblia enzyme presented here is thus the first structure determination of a prolyl-tRNA synthetase from a eukaryote. The relative occupancies of substrate (proline) and product (prolyl-AMP) in the active site are consistent with half-of-the-sites reactivity, as is the observed biphasic thermal denaturation curve for the protein in the presence of proline and MgATP. However, no corresponding induced asymmetry is evident in the structure of the protein. No thermal stabilization is observed in the presence of cysteine and ATP. The implied low affinity for the off-target activation product cysteinyl-AMP suggests that translational fidelity in Giardia is aided by the rapid release of misactivated cysteine.

  7. Lokiarchaea are close relatives of Euryarchaeota, not bridging the gap between prokaryotes and eukaryotes

    Science.gov (United States)

    Forterre, Patrick

    2017-01-01

    The eocyte hypothesis, in which Eukarya emerged from within Archaea, has been boosted by the description of a new candidate archaeal phylum, “Lokiarchaeota”, from metagenomic data. Eukarya branch within Lokiarchaeota in a tree reconstructed from the concatenation of 36 universal proteins. However, individual phylogenies revealed that lokiarchaeal proteins sequences have different evolutionary histories. The individual markers phylogenies revealed at least two subsets of proteins, either supporting the Woese or the Eocyte tree of life. Strikingly, removal of a single protein, the elongation factor EF2, is sufficient to break the Eukaryotes-Lokiarchaea affiliation. Our analysis suggests that the three lokiarchaeal EF2 proteins have a chimeric organization that could be due to contamination and/or homologous recombination with patches of eukaryotic sequences. A robust phylogenetic analysis of RNA polymerases with a new dataset indicates that Lokiarchaeota and related phyla of the Asgard superphylum are sister group to Euryarchaeota, not to Eukarya, and supports the monophyly of Archaea with their rooting in the branch leading to Thaumarchaeota. PMID:28604769

  8. Characterization and Evolution of the Cell Cycle-Associated Mob Domain-Containing Proteins in Eukaryotes

    Directory of Open Access Journals (Sweden)

    Nicola Vitulo

    2007-01-01

    Full Text Available The MOB family includes a group of cell cycle-associated proteins highly conserved throughout eukaryotes, whose founding members are implicated in mitotic exit and co-ordination of cell cycle progression with cell polarity and morphogenesis. Here we report the characterization and evolution of the MOB domain-containing proteins as inferred from the 43 eukaryotic genomes so far sequenced. We show that genes for Mob-like proteins are present in at least 41 of these genomes, confi rming the universal distribution of this protein family and suggesting its prominent biological function. The phylogenetic analysis reveals fi ve distinct MOB domain classes, showing a progressive expansion of this family from unicellular to multicellular organisms, reaching the highest number in mammals. Plant Mob genes appear to have evolved from a single ancestor, most likely after the loss of one or more genes during the early stage of Viridiplantae evolutionary history. Three of the Mob classes are widespread among most of the analyzed organisms. The possible biological and molecular function of Mob proteins and their role in conserved signaling pathways related to cell proliferation, cell death and cell polarity are also presented and critically discussed.

  9. DNA binding polarity, dimerization, and ATPase ring remodeling in the CMG helicase of the eukaryotic replisome

    Science.gov (United States)

    Costa, Alessandro; Renault, Ludovic; Swuec, Paolo; Petojevic, Tatjana; Pesavento, James J; Ilves, Ivar; MacLellan-Gibson, Kirsty; Fleck, Roland A; Botchan, Michael R; Berger, James M

    2014-01-01

    The Cdc45/Mcm2-7/GINS (CMG) helicase separates DNA strands during replication in eukaryotes. How the CMG is assembled and engages DNA substrates remains unclear. Using electron microscopy, we have determined the structure of the CMG in the presence of ATPγS and a DNA duplex bearing a 3′ single-stranded tail. The structure shows that the MCM subunits of the CMG bind preferentially to single-stranded DNA, establishes the polarity by which DNA enters into the Mcm2-7 pore, and explains how Cdc45 helps prevent DNA from dissociating from the helicase. The Mcm2-7 subcomplex forms a cracked-ring, right-handed spiral when DNA and nucleotide are bound, revealing unexpected congruencies between the CMG and both bacterial DnaB helicases and the AAA+ motor of the eukaryotic proteasome. The existence of a subpopulation of dimeric CMGs establishes the subunit register of Mcm2-7 double hexamers and together with the spiral form highlights how Mcm2-7 transitions through different conformational and assembly states as it matures into a functional helicase. DOI: http://dx.doi.org/10.7554/eLife.03273.001 PMID:25117490

  10. Calcium sensors of ciliary outer arm dynein: functions and phylogenetic considerations for eukaryotic evolution.

    Science.gov (United States)

    Inaba, Kazuo

    2015-01-01

    The motility of eukaryotic cilia and flagella is modulated in response to several extracellular stimuli. Ca(2+) is the most critical intracellular factor for these changes in motility, directly acting on the axonemes and altering flagellar asymmetry. Calaxin is an opisthokont-specific neuronal calcium sensor protein first described in the sperm of the ascidian Ciona intestinalis. It binds to a heavy chain of two-headed outer arm dynein in a Ca(2+)-dependent manner and regulates 'asymmetric' wave propagation at high concentrations of Ca(2+). A Ca(2+)-binding subunit of outer arm dynein in Chlamydomonas reinhardtii, the light chain 4 (LC4), which is a Ca(2+)-sensor phylogenetically different from calaxin, shows Ca(2+)-dependent binding to a heavy chain of three-headed outer arm dynein. However, LC4 appears to participate in 'symmetric' wave propagation at high concentrations of Ca(2+). LC4-type dynein light chain is present in bikonts, except for some subclasses of the Excavata. Thus, flagellar asymmetry-symmetry conversion in response to Ca(2+) concentration represents a 'mirror image' relationship between Ciona and Chlamydomonas. Phylogenetic analyses indicate the duplication, divergence, and loss of heavy chain and Ca(2+)-sensors of outer arm dynein among excavate species. These features imply a divergence point with respect to Ca(2+)-dependent regulation of outer arm dynein in cilia and flagella during the evolution of eukaryotic supergroups.

  11. Reconstitution of a eukaryotic replisome reveals suppression mechanisms that define leading/lagging strand operation

    Science.gov (United States)

    Georgescu, Roxana E; Schauer, Grant D; Yao, Nina Y; Langston, Lance D; Yurieva, Olga; Zhang, Dan; Finkelstein, Jeff; O'Donnell, Mike E

    2015-01-01

    We have reconstituted a eukaryotic leading/lagging strand replisome comprising 31 distinct polypeptides. This study identifies a process unprecedented in bacterial replisomes. While bacteria and phage simply recruit polymerases to the fork, we find that suppression mechanisms are used to position the distinct eukaryotic polymerases on their respective strands. Hence, Pol ε is active with CMG on the leading strand, but it is unable to function on the lagging strand, even when Pol δ is not present. Conversely, Pol δ-PCNA is the only enzyme capable of extending Okazaki fragments in the presence of Pols ε and α. We have shown earlier that Pol δ-PCNA is suppressed on the leading strand with CMG (Georgescu et al., 2014). We propose that CMG, the 11-subunit helicase, is responsible for one or both of these suppression mechanisms that spatially control polymerase occupancy at the fork. DOI: http://dx.doi.org/10.7554/eLife.04988.001 PMID:25871847

  12. Strong Ideal Convergence in Probabilistic Metric Spaces

    Indian Academy of Sciences (India)

    In the present paper we introduce the concepts of strongly ideal convergent sequence and strong ideal Cauchy sequence in a probabilistic metric (PM) space endowed with the strong topology, and establish some basic facts. Next, we define the strong ideal limit points and the strong ideal cluster points of a sequence in this ...

  13. Strong Statistical Convergence in Probabilistic Metric Spaces

    OpenAIRE

    Şençimen, Celaleddin; Pehlivan, Serpil

    2008-01-01

    In this article, we introduce the concepts of strongly statistically convergent sequence and strong statistically Cauchy sequence in a probabilistic metric (PM) space endowed with the strong topology, and establish some basic facts. Next, we define the strong statistical limit points and the strong statistical cluster points of a sequence in this space and investigate the relations between these concepts.

  14. John Strong - 1941-2006

    CERN Multimedia

    2006-01-01

    Our friend and colleague John Strong was cruelly taken from us by a brain tumour on 31 July, a few days before his 65th birthday. John started his career and obtained his PhD in a group from Westfield College, initially working on experiments at Rutherford Appleton Laboratory (RAL). From the early 1970s onwards, however, his research was focused on experiments in CERN, with several particularly notable contributions. The Omega spectrometer adopted a system John had originally developed for experiments at RAL using vidicon cameras (a type of television camera) to record the sparks in the spark chambers. This highly automated system allowed Omega to be used in a similar way to bubble chambers. He contributed to the success of NA1 and NA7, where he became heavily involved in the electronic trigger systems. In these experiments the Westfield group joined forces with Italian colleagues to measure the form factors of the pion and the kaon, and the lifetime of some of the newly discovered charm particles. Such h...

  15. Remnants of strong tidal interactions

    International Nuclear Information System (INIS)

    Mcglynn, T.A.

    1990-01-01

    This paper examines the properties of stellar systems that have recently undergone a strong tidal shock, i.e., a shock which removes a significant fraction of the particles in the system, and where the shocked system has a much smaller mass than the producer of the tidal field. N-body calculations of King models shocked in a variety of ways are performed, and the consequences of the shocks are investigated. The results confirm the prediction of Jaffe for shocked systems. Several models are also run where the tidal forces on the system are constant, simulating a circular orbit around a primary, and the development of tidal radii under these static conditions appears to be a mild process which does not dramatically affect material that is not stripped. The tidal radii are about twice as large as classical formulas would predict. Remnant density profiles are compared with a sample of elliptical galaxies, and the implications of the results for the development of stellar populations and galaxies are considered. 38 refs

  16. Strongly correlated perovskite fuel cells

    Science.gov (United States)

    Zhou, You; Guan, Xiaofei; Zhou, Hua; Ramadoss, Koushik; Adam, Suhare; Liu, Huajun; Lee, Sungsik; Shi, Jian; Tsuchiya, Masaru; Fong, Dillon D.; Ramanathan, Shriram

    2016-06-01

    Fuel cells convert chemical energy directly into electrical energy with high efficiencies and environmental benefits, as compared with traditional heat engines. Yttria-stabilized zirconia is perhaps the material with the most potential as an electrolyte in solid oxide fuel cells (SOFCs), owing to its stability and near-unity ionic transference number. Although there exist materials with superior ionic conductivity, they are often limited by their ability to suppress electronic leakage when exposed to the reducing environment at the fuel interface. Such electronic leakage reduces fuel cell power output and the associated chemo-mechanical stresses can also lead to catastrophic fracture of electrolyte membranes. Here we depart from traditional electrolyte design that relies on cation substitution to sustain ionic conduction. Instead, we use a perovskite nickelate as an electrolyte with high initial ionic and electronic conductivity. Since many such oxides are also correlated electron systems, we can suppress the electronic conduction through a filling-controlled Mott transition induced by spontaneous hydrogen incorporation. Using such a nickelate as the electrolyte in free-standing membrane geometry, we demonstrate a low-temperature micro-fabricated SOFC with high performance. The ionic conductivity of the nickelate perovskite is comparable to the best-performing solid electrolytes in the same temperature range, with a very low activation energy. The results present a design strategy for high-performance materials exhibiting emergent properties arising from strong electron correlations.

  17. Strong seismic ground motion propagation

    International Nuclear Information System (INIS)

    Seale, S.; Archuleta, R.; Pecker, A.; Bouchon, M.; Mohammadioun, G.; Murphy, A.; Mohammadioun, B.

    1988-10-01

    At the McGee Creek, California, site, 3-component strong-motion accelerometers are located at depths of 166 m, 35 m and 0 m. The surface material is glacial moraine, to a depth of 30.5 m, overlying homfels. Accelerations were recorded from two California earthquakes: Round Valley, M L 5.8, November 23, 1984, 18:08 UTC and Chalfant Valley, M L 6.4, July 21, 1986, 14:42 UTC. By separating out the SH components of acceleration, we were able to determine the orientations of the downhole instruments. By separating out the SV component of acceleration, we were able to determine the approximate angle of incidence of the signal at 166 m. A constant phase velocity Haskell-Thomson model was applied to generate synthetic SH seismograms at the surface using the accelerations recorded at 166 m. In the frequency band 0.0 - 10.0 Hz, we compared the filtered synthetic records to the filtered surface data. The onset of the SH pulse is clearly seen, as are the reflections from the interface at 30.5 m. The synthetic record closely matches the data in amplitude and phase. The fit between the synthetic accelerogram and the data shows that the seismic amplification at the surface is a result of the contrast of the impedances (shear stiffnesses) of the near surface materials

  18. The N-terminal region of eukaryotic translation initiation factor 5A signals to nuclear localization of the protein

    International Nuclear Information System (INIS)

    Parreiras-e-Silva, Lucas T.; Gomes, Marcelo D.; Oliveira, Eduardo B.; Costa-Neto, Claudio M.

    2007-01-01

    The eukaryotic translation initiation factor 5A (eIF5A) is a ubiquitous protein of eukaryotic and archaeal organisms which undergoes hypusination, a unique post-translational modification. We have generated a polyclonal antibody against murine eIF5A, which in immunocytochemical assays in B16-F10 cells revealed that the endogenous protein is preferentially localized to the nuclear region. We therefore analyzed possible structural features present in eIF5A proteins that could be responsible for that characteristic. Multiple sequence alignment analysis of eIF5A proteins from different eukaryotic and archaeal organisms showed that the former sequences have an extended N-terminal segment. We have then performed in silico prediction analyses and constructed different truncated forms of murine eIF5A to verify any possible role that the N-terminal extension might have in determining the subcellular localization of the eIF5A in eukaryotic organisms. Our results indicate that the N-terminal extension of the eukaryotic eIF5A contributes in signaling this protein to nuclear localization, despite of bearing no structural similarity with classical nuclear localization signals

  19. Prokaryotic and eukaryotic features observed on the secondary structures of Giardia SSU rRNAs and its phylogenetic implications.

    Science.gov (United States)

    Hwang, Ui Wook

    2007-04-01

    Phylogenetic position of a diplomonad protist Giardia, a principle cause of diarrhea, among eukaryotes has been vigorously debated so far. Through the comparisons of primary and secondary structures of SSU rRNAs of G. intestinalis, G. microti, G. ardeae, and G. muris, I found two major indel regions (a 6-nt indel and a 22-26-nt indel), which correspond to the helix 10 of the V2 region and helices E23-8 to E23-9 of the V4 region, respectively. As generally shown in eukaryotes, G. intestinalis and G. microti have commonly a relatively longer helix 10 (a 7-bp stem and a 4-nt loop), and also the eukaryote-specific helices E23-6 to E23-9. On the other hand, G. muris and G. ardeae have a shorter helix 10: a 2-bp stem and a 6-nt loop in G. ardeae and a 3-bp stem and a 6-nt loop in G. muris. In the V4, they have a single long helix (like the P23-1 helix in prokaryotes) instead of the helices E23-6 to E23-9. Among the four Giardia species, co-appearance of prokaryote- and eukaryote-typical features might be significant evidence to suggest that Giardia (Archezoa) is a living fossil showing an "intermediate stage" during the evolution from prokaryotes to eukaryotes.

  20. Strongly interacting photons and atoms

    International Nuclear Information System (INIS)

    Alge, W.

    1999-05-01

    This thesis contains the main results of the research topics I have pursued during the my PhD studies at the University of Innsbruck and partly in collaboration with the Institut d' Optique in Orsay, France. It is divided into three parts. The first and largest part discusses the possibility of using strong standing waves as a tool to cool and trap neutral atoms in optical cavities. This is very important in the field of nonlinear optics where several successful experiments with cold atoms in cavities have been performed recently. A discussion of the optical parametric oscillator in a regime where the nonlinearity dominates the evolution is the topic of the second part. We investigated mainly the statistical properties of the cavity output of the three interactive cavity modes. Very recently a system has been proposed which promises fantastic properties. It should exhibit a giant Kerr nonlinearity with negligible absorption thus leading to a photonic turnstile device based on cold atoms in cavity. We have shown that this model suffers from overly simplistic assumptions and developed several more comprehensive approaches to study the behavior of this system. Apart from the division into three parts of different contents the thesis is divided into publications, supplements and invisible stuff. The intention of the supplements is to reach researchers which work in related areas and provide them with more detailed information about the concepts and the numerical tools we used. It is written especially for diploma and PhD students to give them a chance to use the third part of our work which is actually the largest one. They consist of a large number of computer programs we wrote to investigate the behavior of the systems in parameter regions where no hope exists to solve the equations analytically. (author)

  1. Topics in strong Langmuir turbulence

    International Nuclear Information System (INIS)

    Skoric, M.M.

    1981-01-01

    This thesis discusses certain aspects of the turbulence of a fully ionised non-isothermal plasma dominated by the Langmuir mode. Some of the basic properties of strongly turbulent plasmas are reviewed. In particular, interest is focused on the state of Langmuir turbulence, that is the turbulence of a simple externally unmagnetized plasma. The problem of the existence and dynamics of Langmuir collapse is discussed, often met as a non-linear stage of the modulational instability in the framework of the Zakharov equations (i.e. simple time-averaged dynamical equations). Possible macroscopic consequences of such dynamical turbulent models are investigated. In order to study highly non-linear collapse dynamics in its advanced stage, a set of generalized Zakharov equations are derived. Going beyond the original approximation, the author includes the effects of higher electron non-linearities and a breakdown of slow-timescale quasi-neutrality. He investigates how these corrections may influence the collapse stabilisation. Recently, it has been realised that the modulational instability in a Langmuir plasma will be accompanied by the collisionless-generation of a slow-timescale magnetic field. Accordingly, a novel physical situation has emerged which is investigated in detail. The stability of monochromatic Langmuir waves in a self-magnetized Langmuir plasma, is discussed, and the existence of a novel magneto-modulational instability shown. The wave collapse dynamics is investigated and a physical interpretation of the basic results is given. A problem of the transient analysis of an interaction of time-dependent electromagnetic pulses with linear cold plasma media is investigated. (Auth.)

  2. Promoting Strong Written Communication Skills

    Science.gov (United States)

    Narayanan, M.

    2015-12-01

    The reason that an improvement in the quality of technical writing is still needed in the classroom is due to the fact that universities are facing challenging problems not only on the technological front but also on the socio-economic front. The universities are actively responding to the changes that are taking place in the global consumer marketplace. Obviously, there are numerous benefits of promoting strong written communication skills. They can be summarized into the following six categories. First, and perhaps the most important: The University achieves learner satisfaction. The learner has documented verbally, that the necessary knowledge has been successfully acquired. This results in learner loyalty that in turn will attract more qualified learners.Second, quality communication lowers the cost per pupil, consequently resulting in increased productivity backed by a stronger economic structure and forecast. Third, quality communications help to improve the cash flow and cash reserves of the university. Fourth, having high quality communication enables the university to justify the need for high costs of tuition and fees. Fifth, better quality in written communication skills result in attracting top-quality learners. This will lead to happier and satisfied learners, not to mention greater prosperity for the university as a whole. Sixth, quality written communication skills result in reduced complaints, thus meaning fewer hours spent on answering or correcting the situation. The University faculty and staff are thus able to devote more time on scholarly activities, meaningful research and productive community service. References Boyer, Ernest L. (1990). Scholarship reconsidered: Priorities of the Professorate.Princeton, NJ: Carnegie Foundation for the Advancement of Teaching. Hawkins, P., & Winter, J. (1997). Mastering change: Learning the lessons of the enterprise.London: Department for Education and Employment. Buzzel, Robert D., and Bradley T. Gale. (1987

  3. Waggawagga-CLI: A command-line tool for predicting stable single α-helices (SAH-domains, and the SAH-domain distribution across eukaryotes.

    Directory of Open Access Journals (Sweden)

    Dominic Simm

    Full Text Available Stable single-alpha helices (SAH-domains function as rigid connectors and constant force springs between structural domains, and can provide contact surfaces for protein-protein and protein-RNA interactions. SAH-domains mainly consist of charged amino acids and are monomeric and stable in polar solutions, characteristics which distinguish them from coiled-coil domains and intrinsically disordered regions. Although the number of reported SAH-domains is steadily increasing, genome-wide analyses of SAH-domains in eukaryotic genomes are still missing. Here, we present Waggawagga-CLI, a command-line tool for predicting and analysing SAH-domains in protein sequence datasets. Using Waggawagga-CLI we predicted SAH-domains in 24 datasets from eukaryotes across the tree of life. SAH-domains were predicted in 0.5 to 3.5% of the protein-coding content per species. SAH-domains are particularly present in longer proteins supporting their function as structural building block in multi-domain proteins. In human, SAH-domains are mainly used as alternative building blocks not being present in all transcripts of a gene. Gene ontology analysis showed that yeast proteins with SAH-domains are particular enriched in macromolecular complex subunit organization, cellular component biogenesis and RNA metabolic processes, and that they have a strong nuclear and ribonucleoprotein complex localization and function in ribosome and nucleic acid binding. Human proteins with SAH-domains have roles in all types of RNA processing and cytoskeleton organization, and are predicted to function in RNA binding, protein binding involved in cell and cell-cell adhesion, and cytoskeletal protein binding. Waggawagga-CLI allows the user to adjust the stabilizing and destabilizing contribution of amino acid interactions in i,i+3 and i,i+4 spacings, and provides extensive flexibility for user-designed analyses.

  4. Russia needs a strong counterpart

    International Nuclear Information System (INIS)

    Slovak, K.; Marcan, P.

    2008-01-01

    In this paper an interview with the head of OMV, Wolfgang Ruttenstorfer is published. There is extract from this interview: Q: There have been attempts to take over MOL for a quite long time. Do you think you can still succeed? Since the beginning we kept saying that this would not happen from one day to another. But it may take two to three years. But we are positive that it is justified. Q: Resistance from MOL and the Hungarian government is strong. We have tried to persuade the Hungarian government. We offered them a split company management. A part of the management would be in Budapest. We would locate the management of the largest division - the refinery, there. And of course only the best could be part of the management. We would not nominate people according to their nationality, it would not matter whether the person was Austrian, Hungarian or Slovak. We want a Central European company, not Hungarian, Romanian or Slovak company. Q: Would the transaction still be attractive if, because of pressure exercised by Brussels, you had to sell Slovnaft or your refinery in Szazhalobatta? We do not intend to sell any refineries. Q: Rumours are spreading that the Commission may ask you to sell a refinery? We do not want to speculate. Let us wait and see what happens. We do not want to sell refineries. Q: It is said that OMV is coordinating or at least consulting its attempts to acquire MOL with Gazprom. There are many rumours in Central Europe. But I can tell you this is not true. We are interested in this merger because we feel the increasing pressure exercised by Kazakhstan and Russia. We, of course, have a good relationship with Gazprom which we have had enjoyed for over forty years. As indeed Slovakia has. Q: A few weeks ago Austrian daily Wirtschaftsblatt published an article about Gazprom's interest in OMV shares. That is gossip that is more than ten years' old. Similarly to the rumours that Gazprom is a shareholder of MOL. There are no negotiations with Gazprom

  5. Heterogeneous Family of Cyclomodulins: Smart Weapons That Allow Bacteria to Hijack the Eukaryotic Cell Cycle and Promote Infections

    Directory of Open Access Journals (Sweden)

    Rachid A. El-Aouar Filho

    2017-05-01

    Full Text Available Some bacterial pathogens modulate signaling pathways of eukaryotic cells in order to subvert the host response for their own benefit, leading to successful colonization and invasion. Pathogenic bacteria produce multiple compounds that generate favorable conditions to their survival and growth during infection in eukaryotic hosts. Many bacterial toxins can alter the cell cycle progression of host cells, impairing essential cellular functions and impeding host cell division. This review summarizes current knowledge regarding cyclomodulins, a heterogeneous family of bacterial effectors that induce eukaryotic cell cycle alterations. We discuss the mechanisms of actions of cyclomodulins according to their biochemical properties, providing examples of various cyclomodulins such as cycle inhibiting factor, γ-glutamyltranspeptidase, cytolethal distending toxins, shiga toxin, subtilase toxin, anthrax toxin, cholera toxin, adenylate cyclase toxins, vacuolating cytotoxin, cytotoxic necrotizing factor, Panton-Valentine leukocidin, phenol soluble modulins, and mycolactone. Special attention is paid to the benefit provided by cyclomodulins to bacteria during colonization of the host.

  6. Wholly Rickettsia! Reconstructed Metabolic Profile of the Quintessential Bacterial Parasite of Eukaryotic Cells.

    Science.gov (United States)

    Driscoll, Timothy P; Verhoeve, Victoria I; Guillotte, Mark L; Lehman, Stephanie S; Rennoll, Sherri A; Beier-Sexton, Magda; Rahman, M Sayeedur; Azad, Abdu F; Gillespie, Joseph J

    2017-09-26

    Reductive genome evolution has purged many metabolic pathways from obligate intracellular Rickettsia ( Alphaproteobacteria ; Rickettsiaceae ). While some aspects of host-dependent rickettsial metabolism have been characterized, the array of host-acquired metabolites and their cognate transporters remains unknown. This dearth of information has thwarted efforts to obtain an axenic Rickettsia culture, a major impediment to conventional genetic approaches. Using phylogenomics and computational pathway analysis, we reconstructed the Rickettsia metabolic and transport network, identifying 51 host-acquired metabolites (only 21 previously characterized) needed to compensate for degraded biosynthesis pathways. In the absence of glycolysis and the pentose phosphate pathway, cell envelope glycoconjugates are synthesized from three imported host sugars, with a range of additional host-acquired metabolites fueling the tricarboxylic acid cycle. Fatty acid and glycerophospholipid pathways also initiate from host precursors, and import of both isoprenes and terpenoids is required for the synthesis of ubiquinone and the lipid carrier of lipid I and O-antigen. Unlike metabolite-provisioning bacterial symbionts of arthropods, rickettsiae cannot synthesize B vitamins or most other cofactors, accentuating their parasitic nature. Six biosynthesis pathways contain holes (missing enzymes); similar patterns in taxonomically diverse bacteria suggest alternative enzymes that await discovery. A paucity of characterized and predicted transporters emphasizes the knowledge gap concerning how rickettsiae import host metabolites, some of which are large and not known to be transported by bacteria. Collectively, our reconstructed metabolic network offers clues to how rickettsiae hijack host metabolic pathways. This blueprint for growth determinants is an important step toward the design of axenic media to rescue rickettsiae from the eukaryotic cell. IMPORTANCE A hallmark of obligate intracellular

  7. EuGI: a novel resource for studying genomic islands to facilitate horizontal gene transfer detection in eukaryotes.

    Science.gov (United States)

    Clasen, Frederick Johannes; Pierneef, Rian Ewald; Slippers, Bernard; Reva, Oleg

    2018-05-03

    Genomic islands (GIs) are inserts of foreign DNA that have potentially arisen through horizontal gene transfer (HGT). There are evidences that GIs can contribute significantly to the evolution of prokaryotes. The acquisition of GIs through HGT in eukaryotes has, however, been largely unexplored. In this study, the previously developed GI prediction tool, SeqWord Gene Island Sniffer (SWGIS), is modified to predict GIs in eukaryotic chromosomes. Artificial simulations are used to estimate ratios of predicting false positive and false negative GIs by inserting GIs into different test chromosomes and performing the SWGIS v2.0 algorithm. Using SWGIS v2.0, GIs are then identified in 36 fungal, 22 protozoan and 8 invertebrate genomes. SWGIS v2.0 predicts GIs in large eukaryotic chromosomes based on the atypical nucleotide composition of these regions. Averages for predicting false negative and false positive GIs were 20.1% and 11.01% respectively. A total of 10,550 GIs were identified in 66 eukaryotic species with 5299 of these GIs coding for at least one functional protein. The EuGI web-resource, freely accessible at http://eugi.bi.up.ac.za , was developed that allows browsing the database created from identified GIs and genes within GIs through an interactive and visual interface. SWGIS v2.0 along with the EuGI database, which houses GIs identified in 66 different eukaryotic species, and the EuGI web-resource, provide the first comprehensive resource for studying HGT in eukaryotes.

  8. Transcription factor IID in the Archaea: sequences in the Thermococcus celer genome would encode a product closely related to the TATA-binding protein of eukaryotes

    Science.gov (United States)

    Marsh, T. L.; Reich, C. I.; Whitelock, R. B.; Olsen, G. J.; Woese, C. R. (Principal Investigator)

    1994-01-01

    The first step in transcription initiation in eukaryotes is mediated by the TATA-binding protein, a subunit of the transcription factor IID complex. We have cloned and sequenced the gene for a presumptive homolog of this eukaryotic protein from Thermococcus celer, a member of the Archaea (formerly archaebacteria). The protein encoded by the archaeal gene is a tandem repeat of a conserved domain, corresponding to the repeated domain in its eukaryotic counterparts. Molecular phylogenetic analyses of the two halves of the repeat are consistent with the duplication occurring before the divergence of the archael and eukaryotic domains. In conjunction with previous observations of similarity in RNA polymerase subunit composition and sequences and the finding of a transcription factor IIB-like sequence in Pyrococcus woesei (a relative of T. celer) it appears that major features of the eukaryotic transcription apparatus were well-established before the origin of eukaryotic cellular organization. The divergence between the two halves of the archael protein is less than that between the halves of the individual eukaryotic sequences, indicating that the average rate of sequence change in the archael protein has been less than in its eukaryotic counterparts. To the extent that this lower rate applies to the genome as a whole, a clearer picture of the early genes (and gene families) that gave rise to present-day genomes is more apt to emerge from the study of sequences from the Archaea than from the corresponding sequences from eukaryotes.

  9. Modular architecture of eukaryotic RNase P and RNase MRP revealed by electron microscopy.

    Science.gov (United States)

    Hipp, Katharina; Galani, Kyriaki; Batisse, Claire; Prinz, Simone; Böttcher, Bettina

    2012-04-01

    Ribonuclease P (RNase P) and RNase MRP are closely related ribonucleoprotein enzymes, which process RNA substrates including tRNA precursors for RNase P and 5.8 S rRNA precursors, as well as some mRNAs, for RNase MRP. The structures of RNase P and RNase MRP have not yet been solved, so it is unclear how the proteins contribute to the structure of the complexes and how substrate specificity is determined. Using electron microscopy and image processing we show that eukaryotic RNase P and RNase MRP have a modular architecture, where proteins stabilize the RNA fold and contribute to cavities, channels and chambers between the modules. Such features are located at strategic positions for substrate recognition by shape and coordination of the cleaved-off sequence. These are also the sites of greatest difference between RNase P and RNase MRP, highlighting the importance of the adaptation of this region to the different substrates.

  10. Crystallization and preliminary X-ray analysis of eukaryotic initiation factor 4E from Pisum sativum

    International Nuclear Information System (INIS)

    Ashby, Jamie A.; Stevenson, Clare E. M.; Maule, Andrew J.; Lawson, David M.

    2009-01-01

    Crystals of N-terminally truncated eIF4E from pea were obtained and X-ray data were recorded in-house to a resolution of 2.2 Å. Crystals of an N-terminally truncated 20 kDa fragment of Pisum sativum eIF4E (ΔN-eIF4E) were grown by vapour diffusion. X-ray data were recorded to a resolution of 2.2 Å from a single crystal in-house. Indexing was consistent with primitive monoclinic symmetry and solvent-content estimations suggested that between four and nine copies of the eIF4E fragment were possible per crystallographic asymmetric unit. eIF4E is an essential component of the eukaryotic translation machinery and recent studies have shown that point mutations of plant eIF4Es can confer resistance to potyvirus infection

  11. A high-throughput screening assay for eukaryotic elongation factor 2 kinase inhibitors

    Directory of Open Access Journals (Sweden)

    Ting Xiao

    2016-10-01

    Full Text Available Eukaryotic elongation factor 2 kinase (eEF2K inhibitors may aid in the development of new therapeutic agents to combat cancer. Purified human eEF2K was obtained from an Escherichia coli expression system and a luminescence-based high-throughput screening (HTS assay was developed using MH-1 peptide as the substrate. The luminescent readouts correlated with the amount of adenosine triphosphate remaining in the kinase reaction. This method was applied to a large-scale screening campaign against a diverse compound library and subsequent confirmation studies. Nine initial hits showing inhibitory activities on eEF2K were identified from 56,000 synthetic compounds during the HTS campaign, of which, five were chosen to test their effects in cancer cell lines.

  12. Functional 5' UTR mRNA structures in eukaryotic translation regulation and how to find them.

    Science.gov (United States)

    Leppek, Kathrin; Das, Rhiju; Barna, Maria

    2018-03-01

    RNA molecules can fold into intricate shapes that can provide an additional layer of control of gene expression beyond that of their sequence. In this Review, we discuss the current mechanistic understanding of structures in 5' untranslated regions (UTRs) of eukaryotic mRNAs and the emerging methodologies used to explore them. These structures may regulate cap-dependent translation initiation through helicase-mediated remodelling of RNA structures and higher-order RNA interactions, as well as cap-independent translation initiation through internal ribosome entry sites (IRESs), mRNA modifications and other specialized translation pathways. We discuss known 5' UTR RNA structures and how new structure probing technologies coupled with prospective validation, particularly compensatory mutagenesis, are likely to identify classes of structured RNA elements that shape post-transcriptional control of gene expression and the development of multicellular organisms.

  13. Dynamic Architecture of Eukaryotic DNA Replication Forks In Vivo, Visualized by Electron Microscopy.

    Science.gov (United States)

    Zellweger, Ralph; Lopes, Massimo

    2018-01-01

    The DNA replication process can be heavily perturbed by several different conditions of genotoxic stress, particularly relevant for cancer onset and therapy. The combination of psoralen crosslinking and electron microscopy has proven instrumental to reveal the fine architecture of in vivo DNA replication intermediates and to uncover their remodeling upon specific conditions of genotoxic stress. The replication structures are stabilized in vivo (by psoralen crosslinking) prior to extraction and enrichment procedures, allowing their visualization at the transmission electron microscope. This chapter outlines the procedures required to visualize and interpret in vivo replication intermediates of eukaryotic genomic DNA, and includes an improved method for enrichment of replication intermediates, compared to previously used BND-cellulose columns.

  14. Structure of a Eukaryotic CLC Transporter Defines an Intermediate State in the Transport Cycle

    International Nuclear Information System (INIS)

    Feng, Liang; Campbell, Ernest B.; Hsiung, Yichun; MacKinnon, Roderick

    2010-01-01

    CLC proteins transport chloride (Cl - ) ions across cell membranes to control the electrical potential of muscle cells, transfer electrolytes across epithelia, and control the pH and electrolyte composition of intracellular organelles. Some members of this protein family are Cl - ion channels, whereas others are secondary active transporters that exchange Cl - ions and protons (H + ) with a 2:1 stoichiometry. We have determined the structure of a eukaryotic CLC transporter at 3.5 angstrom resolution. Cytoplasmic cystathionine beta-synthase (CBS) domains are strategically positioned to regulate the ion-transport pathway, and many disease-causing mutations in human CLCs reside on the CBS-transmembrane interface. Comparison with prokaryotic CLC shows that a gating glutamate residue changes conformation and suggests a basis for 2:1 Cl - /H + exchange and a simple mechanistic connection between CLC channels and transporters.

  15. Structure of a eukaryotic CLC transporter defines an intermediate state in the transport cycle

    Science.gov (United States)

    Feng, Liang; Campbell, Ernest B.; Hsiung, Yichun; MacKinnon, Roderick

    2011-01-01

    CLC proteins transport Cl− ions across cell membranes to control the electrical potential of muscle cells, transfer electrolytes across epithelia, and control the pH and electrolyte composition of intracellular organelles. Some members of this protein family are Cl− ion channels, while others are secondary active transporters that exchange Cl− ions and H+ with a 2:1 stoichiometry. We have determined the structure of a eukaryotic CLC transporter at 3.5 Å resolution. Cytoplasmic CBS domains are strategically positioned to regulate the ion transport pathway, and many disease-causing mutations in human CLCs reside on the CBS-transmembrane interface. Comparison with prokaryotic CLC shows that a gating glutamate changes conformation and suggests a basis for 2:1 Cl−/H+ exchange and a simple mechanistic connection between CLC channels and transporters. PMID:20929736

  16. Advanced method for high-throughput expression of mutated eukaryotic membrane proteins in Saccharomyces cerevisiae

    International Nuclear Information System (INIS)

    Ito, Keisuke; Sugawara, Taishi; Shiroishi, Mitsunori; Tokuda, Natsuko; Kurokawa, Azusa; Misaka, Takumi; Makyio, Hisayoshi; Yurugi-Kobayashi, Takami; Shimamura, Tatsuro; Nomura, Norimichi; Murata, Takeshi; Abe, Keiko; Iwata, So

    2008-01-01

    Crystallization of eukaryotic membrane proteins is a challenging, iterative process. The protein of interest is often modified in an attempt to improve crystallization and diffraction results. To accelerate this process, we took advantage of a GFP-fusion yeast expression system that uses PCR to direct homologous recombination and gene cloning. We explored the possibility of employing more than one PCR fragment to introduce various mutations in a single step, and found that when up to five PCR fragments were co-transformed into yeast, the recombination frequency was maintained as the number of fragments was increased. All transformants expressed the model membrane protein, while the resulting plasmid from each clone contained the designed mutations only. Thus, we have demonstrated a technique allowing the expression of mutant membrane proteins within 5 days, combining a GFP-fusion expression system and yeast homologous recombination

  17. Structure of Prokaryotic Polyamine Deacetylase Reveals Evolutionary Functional Relationships with Eukaryotic Histone Deacetylases

    Energy Technology Data Exchange (ETDEWEB)

    P Lombardi; H Angell; D Whittington; E Flynn; K Rajashankar; D Christianson

    2011-12-31

    Polyamines are a ubiquitous class of polycationic small molecules that can influence gene expression by binding to nucleic acids. Reversible polyamine acetylation regulates nucleic acid binding and is required for normal cell cycle progression and proliferation. Here, we report the structures of Mycoplana ramosa acetylpolyamine amidohydrolase (APAH) complexed with a transition state analogue and a hydroxamate inhibitor and an inactive mutant complexed with two acetylpolyamine substrates. The structure of APAH is the first of a histone deacetylase-like oligomer and reveals that an 18-residue insert in the L2 loop promotes dimerization and the formation of an 18 {angstrom} long 'L'-shaped active site tunnel at the dimer interface, accessible only to narrow and flexible substrates. The importance of dimerization for polyamine deacetylase function leads to the suggestion that a comparable dimeric or double-domain histone deacetylase could catalyze polyamine deacetylation reactions in eukaryotes.

  18. Tus-Ter as a tool to study site-specific DNA replication perturbation in eukaryotes

    DEFF Research Database (Denmark)

    Larsen, Nicolai B; Hickson, Ian D; Mankouri, Hocine W

    2014-01-01

    The high-affinity binding of the Tus protein to specific 21-bp sequences, called Ter, causes site-specific, and polar, DNA replication fork arrest in E coli. The Tus-Ter complex serves to coordinate DNA replication with chromosome segregation in this organism. A number of recent and ongoing studies...... have demonstrated that Tus-Ter can be used as a heterologous tool to generate site-specific perturbation of DNA replication when reconstituted in eukaryotes. Here, we review these recent findings and explore the molecular mechanism by which Tus-Ter mediates replication fork (RF) arrest in the budding...... yeast, S. cerevisiae. We propose that Tus-Ter is a versatile, genetically tractable, and regulatable RF blocking system that can be utilized for disrupting DNA replication in a diverse range of host cells....

  19. Non-AUG translation: a new start for protein synthesis in eukaryotes.

    Science.gov (United States)

    Kearse, Michael G; Wilusz, Jeremy E

    2017-09-01

    Although it was long thought that eukaryotic translation almost always initiates at an AUG start codon, recent advancements in ribosome footprint mapping have revealed that non-AUG start codons are used at an astonishing frequency. These non-AUG initiation events are not simply errors but instead are used to generate or regulate proteins with key cellular functions; for example, during development or stress. Misregulation of non-AUG initiation events contributes to multiple human diseases, including cancer and neurodegeneration, and modulation of non-AUG usage may represent a novel therapeutic strategy. It is thus becoming increasingly clear that start codon selection is regulated by many trans -acting initiation factors as well as sequence/structural elements within messenger RNAs and that non-AUG translation has a profound impact on cellular states. © 2017 Kearse and Wilusz; Published by Cold Spring Harbor Laboratory Press.

  20. Modulation of mutagenesis in eukaryotes by DNA replication fork dynamics and quality of nucleotide pools

    Science.gov (United States)

    Waisertreiger, Irina S.-R.; Liston, Victoria G.; Menezes, Miriam R.; Kim, Hyun-Min; Lobachev, Kirill S.; Stepchenkova, Elena I.; Tahirov, Tahir H.; Rogozin, Igor B.; Pavlov, Youri. I.

    2014-01-01

    The rate of mutations in eukaryotes depends on a plethora of factors and is not immediately derived from the fidelity of DNA polymerases (Pols). Replication of chromosomes containing the anti-parallel strands of duplex DNA occurs through the copying of leading and lagging strand templates by a trio of Pols α, δ and ε, with the assistance of Pol ζ and Y-family Pols at difficult DNA template structures or sites of DNA damage. The parameters of the synthesis at a given location are dictated by the quality and quantity of nucleotides in the pools, replication fork architecture, transcription status, regulation of Pol switches, and structure of chromatin. The result of these transactions is a subject of survey and editing by DNA repair. PMID:23055184