bluuebunny/crossref_metadata_2025 huggingface dataset:
Plasma effects on the location of the outermost magnetic surface
Authors: Boozer Allen H.
Published: September 2005

In both tokamaks and stellarators, the plasma boundary is often defined by the separatrix between magnetic field lines that form toroidal surfaces and field lines that strike solid objects such as the chamber walls or divertor plates. Extremely small magnetic perturbations can move this separatrix radially inward. However, in the presence of a plasma, this radial movement of the separatrix causes a complicated spatial distribution of plasma pressure, which gives rise to currents that can greatly modify or even shield out the magnetic perturbations. An estimate is given of the minimum magnetic perturbation that is required to break up toroidal magnetic surfaces while avoiding plasma shielding.
Photon-spin qubit-conversion based on Overhauser shift of Zeeman energies in quantum dots
Authors: Muto S. , Adachi S. , Yokoi T. , Sasakura H. , Suemune I.
Published: September 2005

We propose a method to realize the conversion of photon qubit and spin qubit using the effective magnetic field created by the nuclear polarization known as Overhauser field. We discuss the preliminary experiment on an InAlAs∕AlGaAs self-assembled quantum dot and also discuss the effects of electron-hole interaction on the conversion.
Ultrasound propagation in the micropores of track membranes
Authors: Gómez Álvarez-Arenas T. E. , González B. , Apel P. Yu. , Orelovitch O. L. , Mitrofanov A. V.
Published: September 2005

Air-coupled and high-frequency ultrasonic spectroscopy is used to study ultrasound transmission through track membranes (TMs). Observed behavior, anomalous compared to that observed for other membranes, suggests independent ultrasound propagation through the pores. This is proved experimentally by studying frequency dependence of some acoustical parameters and by closing the pore aperture at the TM surface. This changes boundary conditions so that such wave is inhibited. Ultrasound propagation in such small pores opens up a new way to investigate gas behavior under rarefied conditions and provides a new technique to characterize TMs.
Ferroelectric-like hysteresis loop in nonferroelectric systems
Authors: Pintilie L. , Alexe M.
Published: September 2005

A ferroelectric-like hysteresis loop is obtained for a nonferroelectric system consisting of two back-to-back metal-semiconductor Schottky contacts with a large concentration of traps distributed over a finite thickness near the electrodes. The presented results show that a simple hysteresis loop measured at a single frequency is not always solid evidence of ferroelectricity in low-dimensional structures such as thin films or nanoscale structures.
Raman spectroscopy of strain in subwavelength microelectronic devices
Authors: Bonera Emiliano , Fanciulli Marco , Mariani Marcello
Published: September 2005

The use of Raman spectroscopy to determine strain in microelectronic devices is intrinsically limited by optical diffraction. The critical issue is not the limited spatial resolution itself, but rather the averaging of inhomogeneously strained regions reducing the sensitivity significantly. To eliminate this effect, we took advantage of the near-field properties of an illuminated subwavelength periodic structure. As it is possible to restrict the investigated volume to the transistor channel only, the sensitivity increases significantly. The technique is advantaged by a very small pitch of the devices, and therefore can be also used in the future technological nodes.
Spin precession in a fractional quantum Hall state with spin-orbit coupling
Authors: Califano Marco , Chakraborty Tapash , Pietiläinen Pekka
Published: September 2005

In order to investigate the fundamental question of whether inter-electron interactions are important in planar electron-based spintronics devices, we have carried out a many-body study of the spin configuration in a quantum Hall state in the presence of Bychkov-Rashba-type spin-orbit interaction. We find that the spin orientation is position dependent and can be tuned via the applied electric field. We propose that this and other properties of such a system are ideally suited for exploitation in spin devices, and that the performance can be further optimized by a careful choice of the growth orientation.
Tungsten nanocrystals embedded in high-k materials for memory application
Authors: Samanta S. K. , Yoo Won Jong , Samudra Ganesh , Tok Eng Soon , Bera L. K. , Balasubramanian N.
Published: September 2005

The formation of tungsten nanocrystals (W-NCs) on atomic-layer-deposited HfAlO∕Al2O3 tunnel oxide was demonstrated for application in a memory device. It was found that the density and size distribution of W-NCs are not only controlled by the initial film thickness, annealing temperature, and time, but also by the metal∕tunnel oxide interface structure. Well-isolated W-NCs with an average diameter of 5 nm and a surface density of 5×1011cm−2 were obtained by applying a thin Al2O3 wetting layer onto HfAlO tunneling oxide. A large flatband voltage shift of 5.7 V was observed from capacitance–voltage measurement when a bias voltage up to ±4V was applied.
Complementary inverter based on interface doped pentacene
Authors: Ahles Marcus , Schmechel Roland , von Seggern Heinz
Published: September 2005

An organic complementary metal–oxide–semiconductor (O-CMOS) inverter is presented, which is based on a single pentacene layer acting both as n- and p-type organic semiconductors. The circuit consists of two spatially separated transistors realized by pairs of calcium and gold source and drain electrodes, respectively. The p transistor is obtained by utilizing the conventional pentacene∕SiO2 channel interface whereas the n transistor is realized by doping the SiO2 interface with traces of calcium prior to pentacene deposition. Both, n and p transistors work exclusively in unipolar mode within the range of the supply voltage of 60 V. The O-CMOS inverter works reliably with a gain in between 17 and 24, and the respective electron and hole mobilities were found around 0.1cm2V−1s−1. The circuit shows hysteresis, which can be explained by a gate voltage-dependent electron trapping in the n channel. Electron accumulation can also be realized by the application of a polymethylmethacrylate interlayer between SiO2 and pentacene.
Midinfrared intersubband absorption in lattice-matched AlInN∕GaN multiple quantum wells
Authors: Nicolay S. , Carlin J.-F. , Feltin E. , Butté R. , Mosca M. , Grandjean N. , Ilegems M. , Tchernycheva M. , Nevou L. , Julien F. H.
Published: September 2005

We report the observation of midinfrared intersubband (ISB) absorption in nearly lattice-matched AlInN∕GaN multiple-quantum-wells. A clear absorption peak is observed around 3μm involving transitions from the conduction band ground state to the first excited state. In addition to ISB absorption, photoluminescence experiments were carried out on lattice- matched AlInN∕GaN single quantum wells in order to determine the spontaneous polarization discontinuity between GaN and Al0.82In0.18N compounds. The experimental value is in good agreement with theoretical predictions. Our results demonstrate that the AlInN∕GaN system is very promising to achieve crack-free and low dislocation density structures dedicated to intersubband devices in the 2–4μm wavelength range.
Large deep-ultraviolet photocurrent in metal-semiconductor-metal structures fabricated on as-grown boron-doped diamond
Authors: Alvarez Jose , Liao Meiyong , Koide Yasuo
Published: September 2005

Metal-semiconductor-metal planar devices have been fabricated on as-grown boron-doped homoepitaxial diamond thin films. They consist of two Schottky barriers connected back to back. The metal employed was a thin titanium (Ti) layer (5nm) followed by a gold (Au) cap layer (50nm), respectively. The structure shows a high ultraviolet photocurrent at 220nm, which is seven orders of magnitude higher than the reverse dark current (<1pA) for at least an applied voltage of ±0.4V. In addition, anomalous photoconductivity gain is observed. A plausible explanation could be the modification of the Schottky barrier under band-to-band illumination. The spectral photoresponse displays over six orders of magnitude discrimination between deep-ultraviolet (210nm) and visible light (630nm), and reveals a shoulder with an onset at 4.6eV.
Room-temperature operation of a nanoelectromechanical resonator embedded in a phase-locked loop
Authors: Kouh T. , Basarir O. , Ekinci K. L.
Published: September 2005

We describe the operation of a phase-locked loop (PLL) that tracks the high-frequency electromechanical resonance of a nanoscale beam resonator. The fundamental in-plane flexural resonance of the beam resonator embedded in the PLL is actuated electrostatically and detected optically. PLL operation is demonstrated by locking stably to the resonance frequency of the beam, and by tracking this resonance with high fidelity as the beam is mass loaded. Our analysis reproduces the observed locking behavior. Feedback control schemes for nanoelectromechanical resonators may offer prospects for miniature timekeeping devices and ultrasensitive sensors.
Electron emission from carbon nanotube-dispersed MgO layer
Authors: Heo J. N. , Lee J. H. , Jeong T. W. , Lee C. S. , Kim W. S. , Jin Y. W. , Kim J. M. , Yu SeGi , Yi W. K. , Park S. H. , Oh T. S. , Yoo J. B.
Published: September 2005

A simple secondary-electron-emission (SEE)-based source was fabricated using a carbon-nanotube-dispersed MgO precursor solution. This emission source exhibits a high SEE gain as over 103. In addition, the self-sustaining current was observed even after turning off the primary electron beam, then the emission current could be modulated by a small electric field variation (1.2–2V∕μm). The electron energy spectrum of the self-sustaining emission indicates that the major character of the emission is attributed to the field-enhanced SEE rather than direct field emission.
Patterning of ferroelectric nanodot arrays using a silicon nitride shadow mask
Authors: Shin Hyung-Joon , Choi Je Hyuk , Yang Hee Jun , Park Young Dae , Kuk Young , Kang Chi-Jung
Published: September 2005

We grew well-ordered arrays of ferroelectric Pb(Zr0.2Ti0.8)O3 (PZT) nanodots on a SrRuO3∕SrTiO3 substrate by pulsed laser deposition. A silicon nitride shadow mask with ordered holes was used for patterning of the PZT arrays. Each dot has a height of ∼15nm and a diameter of ∼120nm with a similar dome shape over a large area. The ferroelectric properties of individual PZT dots were investigated by piezoresponse force microscopy. A single dot could be polarized individually and the polarized state remained unrelaxed to ∼20min.
General relationship between contact stiffness, contact depth, and mechanical properties for indentation in linear viscoelastic solids using axisymmetric indenters of arbitrary profiles
Authors: Cheng Yang-Tse , Cheng Che-Min
Published: September 2005

We derive a relationship between the initial unloading slope, contact depth, and the instantaneous relaxation modulus for indentation in linear viscoelastic solids by a rigid indenter with an arbitrary axisymmetric smooth profile. Although the same expression is well known for indentation in elastic and in elastic-plastic solids, we show that it is also true for indentation in linear viscoelastic solids, provided that the unloading rate is sufficiently fast. Furthermore, the same expression holds true for both fast loading and unloading. These results should provide a sound basis for using the relationship for determining properties of viscoelastic solids using indentation techniques.
Highly efficient wide-angle transmission into uniform rod-type photonic crystals
Authors: White T. P. , de Sterke C. Martijn , McPhedran R. C. , Botten L. C.
Published: September 2005

We report on very high coupling efficiencies into rod-type photonic crystal (PC) slabs with unmodified interfaces. Three-dimensional finite difference time domain simulations show that reflected powers of less than 1.5% can be achieved for incident angles from 0° to more than 25°. We use this property to design a beam combiner exhibiting both high efficiency coupling and self-collimation. The results could be applied to the design of efficient superprisms and other dispersion-based PC devices.
Effect of boundary conditions on the classical skin depth and nonlocal behavior in inductively coupled plasmas
Authors: Rehman Aman-ur- , Pu Yi-Kang
Published: September 2005

When the finiteness of plasma geometry is taken into account, the expression for classical skin depth is different from the one obtained for an unbounded plasma (for both the planar and cylindrical geometries). This change in the expression of the classical skin depth also changes the nonlocality parameter, which is defined as the square of the ratio of the effective mean free path to the classical skin depth. It is concluded that it is the compactness of the geometry due to the metallic boundary condition (E=0) that impacts nonlocal heating (particularly in the low-frequency regime) rather than the shape of the geometry.
Dissecting the mechanical unfolding of ubiquitin
Authors: Irbäck Anders , Mitternacht Simon , Mohanty Sandipan
Published: September 2005

The unfolding behavior of ubiquitin under the influence of a stretching force recently was investigated experimentally by single-molecule constant-force methods. Many observed unfolding traces had a simple two-state character, whereas others showed clear evidence of intermediate states. Here, we use Monte Carlo simulations to investigate the force-induced unfolding of ubiquitin at the atomic level. In agreement with experimental data, we find that the unfolding process can occur either in a single step or through intermediate states. In addition to this randomness, we find that many quantities, such as the frequency of occurrence of intermediates, show a clear systematic dependence on the strength of the applied force. Despite this diversity, one common feature can be identified in the simulated unfolding events, which is the order in which the secondary-structure elements break. This order is the same in two- and three-state events and at the different forces studied. The observed order remains to be verified experimentally but appears physically reasonable.
Neuronal sorting protein-related receptor sorLA/LR11 regulates processing of the amyloid precursor protein
Authors: Andersen Olav M. , Reiche Juliane , Schmidt Vanessa , Gotthardt Michael , Spoelgen Robert , Behlke Joachim , von Arnim Christine A. F. , Breiderhoff Tilman , Jansen Pernille , Wu Xin , Bales Kelly R. , Cappai Roberto , Masters Colin L. , Gliemann Jørgen , Mufson Elliott J. , Hyman Bradley T. , Paul Steven M. , Nykjær Anders , Willnow Thomas E.
Published: September 2005

sorLA (sorting protein-related receptor) is a type-1 membrane protein of unknown function that is expressed in neurons. Its homology to sorting receptors that shuttle between the plasma membrane, endosomes, and the Golgi suggests a related function in neuronal trafficking processes. Because expression of sorLA is reduced in the brain of patients with Alzheimer's disease (AD), we tested involvement of this receptor in intracellular transport and processing of the amyloid precursor protein (APP) to the amyloid β-peptide (Aβ), the principal component of senile plaques. We demonstrate that sorLA interacts with APP
in vitro
and in living cells and that both proteins colocalize in endosomal and Golgi compartments. Overexpression of sorLA in neurons causes redistribution of APP to the Golgi and decreased processing to Aβ, whereas ablation of sorLA expression in knockout mice results in increased levels of Aβ in the brain similar to the situation in AD patients. Thus, sorLA acts as a sorting receptor that protects APP from processing into Aβ and thereby reduces the burden of amyloidogenic peptide formation. Consequently, reduced receptor expression in the human brain may increase Aβ production and plaque formation and promote spontaneous AD.
Dissection of the carbohydrate specificity of the broadly neutralizing anti-HIV-1 antibody 2G12
Authors: Calarese Daniel A. , Lee Hing-Ken , Huang Cheng-Yuan , Best Michael D. , Astronomo Rena D. , Stanfield Robyn L. , Katinger Hermann , Burton Dennis R. , Wong Chi-Huey , Wilson Ian A.
Published: September 2005

Human antibody 2G12 neutralizes a broad range of HIV-1 isolates. Hence, molecular characterization of its epitope, which corresponds to a conserved cluster of oligomannoses on the viral envelope glycoprotein gp120, is a high priority in HIV vaccine design. A prior crystal structure of 2G12 in complex with Man9GlcNAc2highlighted the central importance of the D1 arm in antibody binding. To characterize the specificity of 2G12 more precisely, we performed solution-phase ELISA, carbohydrate microarray analysis, and cocrystallized Fab 2G12 with four different oligomannose derivatives (Man4, Man5, Man7, and Man8) that compete with gp120 for binding to 2G12. Our combined studies reveal that 2G12 is capable of binding both the D1 and D3 arms of the Man9GlcNAc2moiety, which would provide more flexibility to make the required multivalent interactions between the antibody and the gp120 oligomannose cluster than thought previously. These results have important consequences for the design of immunogens to elicit 2G12-like neutralizing antibodies as a component of an HIV vaccine.
Cytoskeletal coordination during neuronal migration
Authors: Schaar Bruce T. , McConnell Susan K.
Published: September 2005

Discoveries from human and mouse genetics have identified cytoskeletal and signaling proteins that are essential for neuronal migration in the developing brain. To provide a meaningful context for these studies, we took an unbiased approach of correlative electron microscopy of neurons migrating through a three-dimensional matrix, and characterized the cytoskeletal events that occur as migrating neurons initiate saltatory forward movements of the cell nucleus. The formation of a cytoplasmic dilation in the proximal leading process precedes nuclear translocation. Cell nuclei translocate into these dilations in saltatory movements. Time-lapse imaging and pharmacological perturbation suggest that nucleokinesis requires stepwise or hierarchical interactions between microtubules, myosin II, and cell adhesion. We hypothesize that these interactions couple leading process extension to nuclear translocation during neuronal migration.
Satellite-observed photosynthetic trends across boreal North America associated with climate and fire disturbance
Authors: Goetz Scott J. , Bunn Andrew G. , Fiske Gregory J. , Houghton R. A.
Published: September 2005

We analyzed trends in a time series of photosynthetic activity across boreal North America over 22 years (1981 through 2003). Nearly 15% of the region displayed significant trends, of which just over half involved temperature-related increases in growing season length and photosynthetic intensity, mostly in tundra. In contrast, forest areas unaffected by fire during the study period declined in photosynthetic activity and showed no systematic change in growing season length. Stochastic changes across the time series were predominantly associated with a frequent and increasing fire disturbance regime. These trends have implications for the direction of feedbacks to the climate system and emphasize the importance of longer term synoptic observations of arctic and boreal biomes.
Eukaryotic translation initiation factor 4GI and p97 promote cellular internal ribosome entry sequence-driven translation
Authors: Hundsdoerfer Patrick , Thoma Christian , Hentze Matthias W.
Published: September 2005

Numerous cellular mRNAs encoding proteins critical during cell stress, apoptosis, and the cell cycle seem to be translated by means of internal ribosome entry sequences (IRES) when cap-dependent translation is compromised. The underlying molecular mechanisms are largely unknown. Using a HeLa-based cell-free translation system that mirrors the function of cellular IRESs
in vitro
, we recently demonstrated that translation from the c-
myc
IRES continues after proteolytic cleavage of eukaryotic translation initiation factor (eIF) 4G. To address the role of eIF4G in cellular IRES-driven translation directly, we immunodepleted eIF4GI from the HeLa cell translation extracts. After efficient depletion of eIF4GI (>90%), both cap-dependent and c-
myc
IRES-dependent translations are diminished to residual levels (<5%). In striking contrast to cap-dependent translation, c-
myc
IRES-dependent translation is fully restored by addition of the conserved middle fragment of eIF4GI, harboring the eIF3- and eIF4A-binding sites. p97, an eIF4G-related protein that has been described both as an inhibitor of translation and as a modulator of apoptosis, not only suffices to also rescue c-
myc
IRES-driven (but not cap-dependent) translation, but it even superinduces IRES-mediated translation 3-fold compared with nondepleted extracts. Interestingly, both p97 and the middle fragment of eIF4GI also rescue translation driven by proapoptotic (p97) and antiapoptotic [X-linked inhibitor of apoptosis (XIAP) and cellular inhibitor of apoptosis 1 (c-IAP1)] IRESs, reflecting a broader role of these polypeptides in cellular IRES-mediated translation and indicating their importance in apoptosis.
Correction of chromosomal mutation and random integration in embryonic stem cells with helper-dependent adenoviral vectors
Authors: Ohbayashi Fumi , Balamotis Michael A. , Kishimoto Atsuhiro , Aizawa Emi , Diaz Arturo , Hasty Paul , Graham Frank L. , Caskey C. Thomas , Mitani Kohnosuke
Published: September 2005

For gene therapy of inherited diseases, targeted integration/gene repair through homologous recombination (HR) between exogenous and chromosomal DNA would be an ideal strategy to avoid potentially serious problems of random integration such as cellular transformation and gene silencing. Efficient sequence-specific modification of chromosomes by HR would also advance both biological studies and therapeutic applications of a variety of stem cells. Toward these goals, we developed an improved strategy of adenoviral vector (AdV)-mediated HR and examined its ability to correct an insertional mutation in the hypoxanthine phosphoribosyl transferase (
Hprt
) locus in male mouse ES cells. The efficiency of HR was compared between four types of AdVs that contained various lengths of homologies at the
Hprt
locus and with various multiplicities of infections. The frequency of HR with helper-dependent AdVs (HD AdVs) with an 18.6-kb homology reached 0.2% per transduced cell at a multiplicity of infection of 10 genomes per cell. Detection of random integration at DNA levels by PCR revealed extremely high efficiency of 5% per cell. We also isolated and characterized chromosomal sites where HD AdVs integrated in a random manner. In contrast to retroviral, lentiviral, and adeno-associated viral vectors, which tend to integrate into genes, the integration sites of AdV was distributed randomly inside and outside genes. These findings suggest that HR mediated by HD AdVs is efficient and relatively safe and might be a new viable option for
ex vivo
gene therapy as well as a tool for chromosomal manipulation of a variety of stem cells.
Histone Deacetylase 6 Regulates Human Immunodeficiency Virus Type 1 Infection
Authors: Valenzuela-Fernández Agustín , Álvarez Susana , Gordon-Alonso Mónica , Barrero Marta , Ursa Ángeles , Cabrero J. Román , Fernández Gerónimo , Naranjo-Suárez Salvador , Yáñez-Mo Maria , Serrador Juan M. , Muñoz-Fernández M. Ángeles , Sánchez-Madrid Francisco
Published: September 2005

Efficient human immunodeficiency virus (HIV)-1 infection depends on multiple interactions between the viral gp41/gp120 envelope (Env) proteins and cell surface receptors. However, cytoskeleton-associated proteins that modify membrane dynamics may also regulate the formation of the HIV-mediated fusion pore and hence viral infection. Because the effects of HDAC6-tubulin deacetylase on cortical α-tubulin regulate cell migration and immune synapse organization, we explored the possible role of HDAC6 in HIV-1-envelope-mediated cell fusion and infection. The binding of the gp120 protein to CD4+-permissive cells increased the level of acetylated α-tubulin in a CD4-dependent manner. Furthermore, overexpression of active HDAC6 inhibited the acetylation of α-tubulin, and remarkably, prevented HIV-1 envelope-dependent cell fusion and infection without affecting the expression and codistribution of HIV-1 receptors. In contrast, knockdown of HDAC6 expression or inhibition of its tubulin deacetylase activity strongly enhanced HIV-1 infection and syncytia formation. These results demonstrate that HDAC6 plays a significant role in regulating HIV-1 infection and Env-mediated syncytia formation.
Role of the C Terminus of Mec1 Checkpoint Kinase in Its Localization to Sites of DNA Damage
Authors: Nakada Daisuke , Hirano Yukinori , Tanaka Yuya , Sugimoto Katsunori
Published: September 2005

The large protein kinases, ataxia-telangiectasia mutated (ATM) and ATM-Rad3-related (ATR), coordinate the cellular response to DNA damage. In budding yeast, ATR homologue Mec1 plays a central role in DNA damage signaling. Mec1 interacts physically with Ddc2 and functions in the form of the Mec1–Ddc2 complex. To identify proteins interacting with the Mec1–Ddc2 complex, we performed a modified two-hybrid screen and isolated RFA1 and RFA2, genes that encode subunits of replication protein A (RPA). Using the two-hybrid system, we found that the extreme C-terminal region of Mec1 is critical for RPA binding. The C-terminal substitution mutation does not affect the Mec1–Ddc2 complex formation, but it does impair the interaction of Mec1 and Ddc2 with RPA as well as their association with DNA lesions. The C-terminal mutation also decreases Mec1 kinase activity. However, the Mec1 kinase-defect by itself does not perturb Mec1 association with sites of DNA damage. We also found that Mec1 and Ddc2 associate with sites of DNA damage in an interdependent manner. Our findings support the model in which Mec1 and Ddc2 localize to sites of DNA damage by interacting with RPA in the form of the Mec1–Ddc2 complex.
Oscillatory ROP GTPase Activation Leads the Oscillatory Polarized Growth of Pollen Tubes
Authors: Hwang Jae-Ung , Gu Ying , Lee Yong-Jik , Yang Zhenbiao
Published: September 2005

Oscillation regulates a wide variety of processes ranging from chemotaxis in Dictyostelium through segmentation in vertebrate development to circadian rhythms. Most studies on the molecular mechanisms underlying oscillation have focused on processes requiring a rhythmic change in gene expression, which usually exhibit a periodicity of >10 min. Mechanisms that control oscillation with shorter periods (<10 min), presumably independent of gene expression changes, are poorly understood. Oscillatory pollen tube tip growth provides an excellent model to investigate such mechanisms. It is well established that ROP1, a Rho-like GTPase from plants, plays an essential role in polarized tip growth in pollen tubes. In this article, we demonstrate that tip-localized ROP1 GTPase activity oscillates in the same frequency with growth oscillation, and leads growth both spatially and temporally. Tip growth requires the coordinate action of two ROP1 downstream pathways that promote the accumulation of tip-localized Ca2+and actin microfilaments (F-actin), respectively. We show that the ROP1 activity oscillates in a similar phase with the apical F-actin but apparently ahead of tip-localized Ca2+. Furthermore, our observations support the hypothesis that the oscillation of tip-localized ROP activity and ROP-dependent tip growth in pollen tubes is modulated by the two temporally coordinated downstream pathways, an early F-actin assembly pathway and a delayed Ca2+gradient-forming pathway. To our knowledge, our report is the first to demonstrate the oscillation of Rho GTPase signaling, which may be a common mechanism underlying the oscillation of actin-dependent processes such as polar growth, cell movement, and chemotaxis.
Nuclear Actin Extends, with No Contraction in Sight
Authors: Pederson Thoru , Aebi Ueli
Published: September 2005

Within the past two years, actin has been implicated in eukaryotic gene transcription by all three classes of RNA polymerase. Moreover, within just the past year, actin has been identified as a constituent of filaments attached to the nuclear pore complexes and extending into the nucleus. This review summarizes these and other very recent advances in the nuclear actin field and emphasizes the key present issues. On the one hand, we are confronted with a body of evidence for a role of actin in gene transcription but with no known structural basis; on the other hand, there is now evidence for polymeric actin—not likely in the classical F-actin conformation—in the nuclear periphery with no known function. In addition, numerous proteins that interact with either G- or F-actin are increasingly being detected in the nucleus, suggesting that both monomeric and oligomeric or polymeric forms of actin are at play and raising the possibility that the equilibrium between them, perhaps differentially regulated at various intranuclear sites, may be a major determinant of nuclear function.
Roles of Replication Fork-interacting and Chk1-activating Domains from Claspin in a DNA Replication Checkpoint Response
Authors: Lee Joon , Gold Daniel A. , Shevchenko Anna , Shevchenko Andrej , Dunphy William G.
Published: September 2005

Claspin is essential for the ATR-dependent activation of Chk1 in Xenopus egg extracts containing incompletely replicated DNA. Claspin associates with replication forks upon origin unwinding. We show that Claspin contains a replication fork-interacting domain (RFID, residues 265–605) that associates with Cdc45, DNA polymerase ϵ, replication protein A, and two replication factor C complexes on chromatin. The RFID contains two basic patches (BP1 and BP2) at amino acids 265–331 and 470–600, respectively. Deletion of either BP1 or BP2 compromises optimal binding of Claspin to chromatin. Absence of BP1 has no effect on the ability of Claspin to mediate activation of Chk1. By contrast, removal of BP2 causes a large reduction in the Chk1-activating potency of Claspin. We also find that Claspin contains a small Chk1-activating domain (residues 776–905) that does not bind stably to chromatin, but it is fully effective at high concentrations for mediating activation of Chk1. These results indicate that stable retention of Claspin on chromatin is not necessary for activation of Chk1. Instead, our findings suggest that only transient interaction of Claspin with replication forks potentiates its Chk1-activating function. Another implication of this work is that stable binding of Claspin to chromatin may play a role in other functions besides the activation of Chk1.
High-level resistance to oxidative stress in Lactococcus lactis conferred by Bacillus subtilis catalase KatE
Authors: Rochat T. , Miyoshi A. , Gratadoux J. J. , Duwat P. , Sourice S. , Azevedo V. , Langella P.
Published: September 2005

Lactococcus lactis, a lactic acid bacterium widely used for food fermentations, is often exposed to damaging stress conditions. In particular, oxidative stress leads to DNA, protein and membrane damages that can be lethal. AsL. lactishas no catalase, the impact of production of theBacillus subtilishaem catalase KatE on its oxidative stress resistance was tested. This cytoplasmic catalase was engineered for extracellular expression inL. lactiswith an optimization strategy based on fusion to the nisin-inducible promoter and a lactococcal signal peptide (SPUsp45). The production of KatE byL. lactisconferred an 800-fold increase in survival after 1 h exposure to 4 mM hydrogen peroxide, and a 160-fold greater survival in long-term (3 days) survival of aerated cultures in acydAmutant, which is unable to respire. The presence of KatE protected DNA from oxidative damage and limited its degradation after long-term aeration in acydA/recAmutant, defective in DNA repair.L. lactisis thus able to produce active catalase that can provide efficient antioxidant activity.
Ca2+ release from host intracellular stores and related signal transduction during Campylobacter jejuni 81-176 internalization into human intestinal cells
Authors: Hu Lan , Raybourne Richard B. , Kopecko Dennis J.
Published: September 2005

Campylobacter jejuniis the leading bacterial cause of human diarrhoeal disease in many parts of the world, including the USA. The ability ofC. jejunito invade the host intestinal epithelium is an important determinant of virulence. A common theme among pathogenic invasive micro-organisms is their ability to usurp the eukaryotic cell-signalling systems both to allow for invasion and to trigger disease pathogenesis. Ca2+is very important in a great variety of eukaryotic cell-signalling processes (e.g. calmodulin-activated enzymes, nuclear transcriptional upregulation, and cytoskeletal rearrangements). This study analyses the effects of Ca2+availability on invasion of human INT407 intestinal epithelial cells byC. jejunistrain 81-176. The ability ofC. jejunito invade INT407 cells was not blocked by chelation of any remaining extracellular Ca2+from host cells incubated in Ca2+-free, serum-free media. In contrast,C. jejuniinvasion was markedly reduced either by chelating host intracellular Ca2+with 1,2-bis-(2-)ethane-N,N,N′,N′-tetraacetic acid acetoxymethyl ester (BAPTA, AM) or by blocking the release of Ca2+from intracellular stores with dantrolene or U73122. Moreover, Bay K8644, a plasma-membrane Ca2+-channel agonist, was observed to stimulateC. jejuniinvasion, presumably by increasing host intracellular free Ca2+levels. Measurement of host-cell cytosolic Ca2+via spectrofluorimetry and fluorescence microscopy revealed an increase in Ca2+from 10 min post-infection. Monolayer pretreatment with either a calmodulin antagonist or a specific inhibitor of protein kinase C was found to cause a marked reduction inC. jejuniinvasion, suggesting roles for these Ca2+-activated modulators in signal-transduction events involved inC. jejuniinvasion. These results demonstrate thatC. jejuniinduces the mobilization of Ca2+from host intracellular stores, which is an essential step in the invasion of intestinal cells by this pathogen.
The artAB genes encode a putative ADP-ribosyltransferase toxin homologue associated with Salmonella enterica serovar Typhimurium DT104
Authors: Saitoh Mariko , Tanaka Kiyoshi , Nishimori Kei , Makino Sou-ichi , Kanno Toru , Ishihara Ryoko , Hatama Shinichi , Kitano Rie , Kishima Masato , Sameshima Toshiya , Akiba Masato , Nakazawa Muneo , Yokomizo Yuichi , Uchida Ikuo
Published: September 2005

Many bacterial pathogens encode ADP-ribosyltransferase toxins. The authors identified an ADP-ribosyltransferase toxin homologue (ArtA, ArtB) inSalmonella entericaserovar Typhimurium (S. typhimurium) DT104. ArtA is most homologous to a putative pertussis-like toxin subunit present inSalmonella typhi(STY1890) andSalmonella paratyphiA (SPA1609), while ArtB shows homology to a hypothetical periplasmic protein ofS. typhi(STY1364) andS. paratyphiA (SPA1188), and a putative pertussis-like toxin subunit inS. typhi(STY1891) andS. paratyphiA (SPA1610). TheartAgene was detected from the phage particle fraction upon mitomycin C induction, and the flanking region ofartABcontains a prophage-like sequence, suggesting that these putative toxin genes reside within a prophage. Southern blotting analysis revealed thatartAis conserved in 12 confirmed DT104 strains and in four related strains which are not phage-typed but are classified into the same group as DT104 by both amplified-fragment length polymorphism and pulsed-field gel electrophoresis. Except for one strain, NCTC 73, all 13S. typhimuriumstrains which were classified into different groups from that of DT104 lacked theartAlocus. The results suggest that phage-mediated recombination has resulted in the acquisition ofartgenes inS. typhimuriumDT104 strains.
Identification of two lysophosphatidic acid acyltransferase genes with overlapping function in Pseudomonas fluorescens
Authors: Cullinane Méabh , Baysse Christine , Morrissey John P. , O'Gara Fergal
Published: September 2005

Phosphatidic acid (PA) is known to be a crucial phospholipid intermediate in cell membrane biosynthesis. InEscherichia coli, this molecule is produced from lysophosphatidic acid (LPA) by LPA acyltransferase (EC 2.3.1.51), encoded byplsC.E. colipossesses only one such LPA acyltransferase and aplsCmutant is non-permissive for growth at elevated temperatures. This study describes the identification and characterization of two genes fromPseudomonas fluorescensF113 that encode enzymes with LPA acyltransferase activity. One of the genes,hdtS, was previously described, whereaspatBis a novel gene. In addition, a putative lyso-ornithine lipid acyltransferase was also identified. All three proteins possess conserved acyltransferase domains and are homologous to PlsC and to LPA acyltransferases identified inNeisseria meningitidis. Functional analysis determined that both HdtS and PatB are functional LPA acyltransferases, as both complemented anE. coli plsCmutant. Mutants lacking each of the putative acyltransferases were constructed and analysed. Growth defects were observed forhdtSandpatBsingle mutants, and a doublehdtSpatBmutant could not be constructed. To determine precise roles in phospholipid synthesis, fatty acid methyl ester analysis was carried out. ThehdtSmutant displayed a profile consistent with a defect in LPA acyltransferase activity, whereas no such phenotype was observed in thepatBmutant, indicating thathdtSencodes the primary LPA acyltransferase in the cell. The presence of at least two genes specifying LPA acyltransferase activity may have implications for the function and survival ofP. fluorescensin diverse environments.
epsABCJ genes are involved in the biosynthesis of the exopolysaccharide mauran produced by Halomonas maura
Authors: Arco Yolanda , Llamas Inmaculada , Martínez-Checa Fernando , Argandoña Montserrat , Quesada Emilia , Moral Ana del
Published: September 2005

The moderately halophilic strainHalomonas mauraS-30 produces a high-molecular-mass acidic polymer (4·7×106 Da) composed of repeating units of mannose, galactose, glucose and glucuronic acid. This exopolysaccharide (EPS), known as mauran, has interesting functional properties that make it suitable for use in many industrial fields. Analysis of the flanking regions of a mini-Tn5insertion site in an EPS-deficient mutant ofH. maura, strain TK71, led to the identification of five ORFs (epsABCDJ), which form part of a gene cluster (eps) with the same structural organization as others involved in the biosynthesis of group 1 capsules and some EPSs. Conserved genetic features were found such as JUMPstart andopselements, which are characteristically located preceding the gene clusters for bacterial polysaccharides. On the basis of their amino-acid-sequence homologies, their putative hydropathy profiles and the effect of their mutations, it is predicted that EpsA (an exporter-protein homologue belonging to the OMA family) and EpsC (a chain-length-regulator homologue belonging to the PCP family) play a role in the assembly, polymerization and translocation of mauran. The possibility that mauran might be synthesized via a Wzy-like biosynthesis system, just as it is for many other polysaccharides, is also discussed. This hypothesis is supported by the fact that EpsJ is homologous with some members of the PST-exporter-protein family, which seems to function together with each OMA–PCP pair in polysaccharide transport in Gram-negative bacteria, transferring the assembled lipid-linked repeating units from the cytoplasmic membrane to the periplasmic space. Maximum induction of theepsgenes is reached during stationary phase in the presence of 5 % (w/v) marine salts.
Structural features, properties and regulation of the outer-membrane protein W (OmpW) of Vibrio cholerae
Authors: Nandi Bisweswar , Nandy Ranjan K. , Sarkar Amit , Ghose Asoke C.
Published: September 2005

The outer-membrane protein OmpW ofVibrio choleraewas studied with respect to its structure, functional properties and regulation of expression. On SDS-PAGE, the membrane-associated form of OmpW protein (solubilized by either 0·1 % or 2 % SDS at 25 °C) migrated as a monomer of 19 kDa that changed to 21 kDa on boiling. The protein was hyperexpressed inEscherichia coliin the histidine-tagged form and the purified His6-OmpW (heated or unheated) migrated as a 23 kDa protein on SDS-PAGE. Circular dichroism and Fourier-transform infrared spectroscopic analyses of the recombinant protein showed the presence ofβ-structures (∼40 %) with minor amounts (8–15 %) ofα-helix. These results were consistent with those obtained by computational analysis of the sequence data of the protein using the secondary structure prediction program Jnet. The recombinant protein did not exhibit any porin-like property in a liposome-swelling assay. An antiserum to the purified protein induced a moderate level (66·6 % and 33·3 % at 1 : 50 and 1 : 100 dilutions, respectively) of passive protection against live vibrio challenge in a suckling mouse model. OmpW-deficient mutants ofV. choleraestrains were generated by insertion mutagenesis. In a competitive assay in mice, the intestinal colonization activities of these mutants were found to be either only marginally diminished (for O1 strains) or 10-fold less (for an O139 strain) as compared to those of the corresponding wild-type strains. The OmpW protein was expressedin vivoas well asin vitroin liquid culture medium devoid of glucose. Interestingly, the glucose-dependent regulation of OmpW expression was less prominent in a ToxR−mutant ofV. cholerae. Further, the expression of OmpW protein was found to be dependent onin vitrocultural conditions such as temperature, salinity, and availability of nutrients or oxygen. These results suggest that the modulation of OmpW expression by environmental factors may be linked to the adaptive response of the organism under stress conditions.
Candida albicans IRS4 contributes to hyphal formation and virulence after the initial stages of disseminated candidiasis
Authors: Badrane Hassan , Cheng Shaoji , Nguyen M. Hong , Jia Hong Yan , Zhang Zongde , Weisner Nghe , Clancy Cornelius J.
Published: September 2005

Candida albicansis a common cause of mucosal and bloodstream infections. As a screening strategy to identify novel candidal virulence factors, sera recovered from HIV-infected patients with active oropharyngeal candidiasis (OPC) were previously used to probe aC. albicansgenomic expression library.IRS4was identified as a gene that encodes an immunogenic protein. In the present study, the presence ofIRS4transcripts was verified within OPC pseudomembranes recovered from patients. Having confirmed that the gene is expressed during human candidiasis, gene disruption strains were created and this implicatedIRS4in diverse processes, including hyphal formation on solid media and under embedded conditions, cell wall integrity and structure, and adherence to human epithelial cellsin vitro.IRS4disruption, however, did not influence hyphal formation or virulence in a murine model of OPC. Rather, the gene was found to be necessary for normal morphogenesis and full virulence during murine intravenously disseminated candidiasis (DC).IRS4's effects on hyphal formation and virulence during DC were not evident on the first day after intravenous inoculation, even though transcripts were detected within murine kidneys. After 4 days, however, anirs4null mutant strain was associated with attenuated mortality, diminished tissue burdens, less extensive infections, impairedC. albicanshyphal formation and decreased kidney damage. Taken together, these findings suggest thatIRS4makes distinct temporal-spatial contributions to the pathogenesis of candidiasis, which appear to vary between different tissue sites as well as within a given tissue over time.
A novel bacteriocin-like substance (BLIS) from a pathogenic strain of Vibrio harveyi
Authors: Prasad Sathish , Morris Peter C. , Hansen Rasmus , Meaden Philip G. , Austin Brian
Published: September 2005

Inter-strain and inter-species inhibition mediated by a bacteriocin-like inhibitory substance (BLIS) from a pathogenic Vibrio harveyi strain VIB 571 was demonstrated against four isolates of the same species, and one culture each of a Vibrio sp., Vibrio fischeri, Vibrio gazogenes and Vibrio parahaemolyticus. The crude BLIS, which was obtained by ammonium-sulphate precipitation of the cell-free supernatant of a 72 h broth culture of strain VIB 571, was inactivated by lipase, proteinase K, pepsin, trypsin, pronase E, SDS and incubation at ≥60 °C for 10 min. The activity was stable between pH 2–11 for at least 5 h. Anion-exchange chromatography, gel filtration, SDS-PAGE and two-dimensional gel electrophoresis revealed the presence of a single major peak, comprising a protein with a pI of ∼5·4 and a molecular mass of ∼32 kDa. The N-terminal amino acid sequence of the protein comprised Asp-Glu-Tyr-Ile-Ser-X-Asn-Lys-X-Ser-Ser-Ala-Asp-Ile (with X representing cysteine or modified amino acid residues). A similarity search based on the matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) generated peptide masses and the N-terminal sequence did not yield any significant matches.
Analysis of teichoic acid biosynthesis regulation reveals that the extracytoplasmic function sigma factor σ
M is induced by phosphate depletion in Bacillus subtilis W23
Authors: Minnig Kathrin , Lazarevic Vladimir , Soldo Blazenka , Mauël Catherine
Published: September 2005

The expression of the Bacillus subtilis W23 tar genes specifying the biosynthesis of the major wall teichoic acid, the poly(ribitol phosphate), was studied under phosphate limitation using lacZ reporter fusions. Three different regulation patterns can be deduced from these β-galactosidase activity data: (i) tarD and tarL gene expression is downregulated under phosphate starvation; (ii) tarA and, to a minor extent, tarB expression after an initial decrease unexpectedly increases; and (iii) tarO is not influenced by phosphate concentration. To dissect the tarA regulatory pattern, its two promoters were analysed under phosphate limitation: The P
tarA
-ext promoter is repressed under phosphate starvation by the PhoPR two-component system, whereas, under the same conditions, the P
tarA
-int promoter is upregulated by the action of an extracytoplasmic function (ECF) σ factor, σ
M. In contrast to strain 168, σ
M is activated in strain W23 in phosphate-depleted conditions, a phenomenon indirectly dependent on PhoPR, the two-component regulatory system responsible for the adaptation to phosphate starvation. These results provide further evidence for the role of σ
M in cell-wall stress response, and suggest that impairment of cell-wall structure is the signal activating this ECF σ factor.