bluuebunny/crossref_metadata_2025 huggingface dataset:
Bubble distributions and dynamics: The expansion‐coalescence equation
Authors: Lovejoy S. , Gaonac'h H. , Schertzer D.
Published: November 2004

As magma rises from depth, it forms bubbles by nucleation, followed by diffusion‐decompressive expansion. Expansion induces shearing, and shearing in turn causes coalescence. As the bubbles grow larger, coalescence gradually becomes more efficient and can be dominant. Coalescence first as a binary (bubble‐bubble) and later as a (possibly singular percolating) multibody process may thus be central to eruption dynamics. Here we consider a binary coalescence model governed by the Smoluchowski or coalescence/coagulation equation. The introduction of decompressive expansion is theoretically straightforward and yields the nonlinear partial integrodifferential expansion‐coalescence equation; we argue that this is a good model for bubble‐bubble dynamics in a decompressing magma. We show that when the collision/interaction kernel has the same form over a wide range of interaction volumes (i.e., it is scaling), exact truncated power law solutions are possible irrespective of the expansion and the collision rate histories. This enables us to reduce the problem to a readily solvable linear ordinary differential equation whose solutions primarily depend on the total interaction integral. In this framework, we investigate the behavior of several eruption models. The validity of the expansion coalescence model is empirically supported by analysis of samples of pumice and lava. Theoretically, the suggested power laws are indeed stable and attractive under a wide range of conditions. We finally point out the effect of small perturbations and new ways to test the theory.
Correlation of seismotectonic variables and GPS strain measurements in western Turkey
Authors: Öncel Ali Osman , Wilson Tom
Published: November 2004

Interrelationships between the multifractal properties of epicenter distribution (Dq), the Gutenberg‐Richter b value, and GPS‐derived strain (shear and dilatation) for the time period extending from 1981 to 1998 are examined in the eastern Mediterranean and western Turkey area. This analysis was conducted in three tectonic subdivisions corresponding to regions dominated by shear, extensional, and compressional deformation. The region of shear coincides with the Northern Anatolian Fault Zone (NAFZ); the region of extension, with the Aegean back arc region; and the region of compression, to the Aegean subduction zone between the African and Anatolian plates. The median b value in the region of extension is larger than those observed in the regions of shear and compression and suggests that large magnitude earthquakes are less probable in this region than along the NAFZ and Aegean subduction zone. A significant correlation is observed between D2 and D15 with dilatation over the 10–40 km range in the back arc region of extension. A marginal correlation between GPS‐derived strain and b value is observed only in the region of compression where it correlates negatively with shear and positively with dilatation. A significant positive correlation is observed between D and b along the NAFZ. A clear relationship between D and b does not exist in regions deformed by extension and compression. The relationship between D and b along the NAFZ may also be an indicator of increased seismic hazard since the positive relationship between D and b is a recent characteristic of the NAFZ observed only in the 20‐year period preceding the Izmit earthquake.
Remarks on the discrete Alfvén wave spectrum induced by the Hall current
Authors: Ito Atsushi , Hirose Akira , Mahajan Swadesh M. , Ohsaki Shuichi
Published: November 2004

It is shown that the discrete Alfvén wave induced by the Hall current [S. Ohsaki and S. M. Mahajan, Phys. Plasmas 11, 898 (2004)] is equivalent to the kinetic Alfvén wave (KAW). The KAW is, thus, accessible in a fluid description. It is further shown that the dispersion relation for the Hall magnetohydrodynamic waves can be reproduced from kinetic theory only if the ion temperature is negligible compared with the electron temperature.
Nonintegrability of nonhomogeneous nonlinear lattices
Authors: Yoshimura Kazuyuki , Umeno Ken
Published: November 2004

We study the integrability of nonlinear lattices with nonhomogeneous polynomial potentials. We prove a nonintegrability theorem for these dynamical systems.
Modeling ultrafast laser-driven ionization dynamics with Monte Carlo collisional particle-in-cell simulations
Authors: Kemp Andreas J. , Pfund Robert E. W. , Meyer-ter-Vehn Jürgen
Published: November 2004

Ionization dynamics of cold dense matter induced by ultrashort (<100 fs) laser pulses is studied for intensities at the onset of the relativistic regime by one-dimensional kinetic simulations. As a model we use a particle-in-cell code that includes field and electron collisional ionization, as well as elastic binary Coulomb collisions. As examples for the different ionization mechanisms, we give the spatial and temporal evolution of laser-induced ionization dynamics in helium gas and solid boron targets. Special attention is paid to the quasi-static electric fields at the rear surface of laser-irradiated targets that are important for laser-ion acceleration.
Operon prediction without a training set
Authors: Westover B. P. , Buhler J. D. , Sonnenburg J. L. , Gordon J. I.
Published: November 2004

Motivation: Annotation of operons in a bacterial genome is an important step in determining an organism's transcriptional regulatory program. While extensive studies of operon structure have been carried out in a few species such as Escherichia coli, fewer resources exist to inform operon prediction in newly sequenced genomes. In particular, many extant operon finders require a large body of training examples to learn the properties of operons in the target organism. For newly sequenced genomes, such examples are generally not available; moreover, a model of operons trained on one species may not reflect the properties of other, distantly related organisms. We encountered these issues in the course of predicting operons in the genome of Bacteroides thetaiotaomicron (B.theta), a common anaerobe that is a prominent component of the normal adult human intestinal microbial community.

Results: We describe an operon predictor designed to work without extensive training data. We rely on a small set of a priori assumptions about the properties of the genome being annotated that permit estimation of the probability that two adjacent genes lie in a common operon. Predictions integrate several sources of information, including intergenic distance, common functional annotation and a novel formulation of conserved gene order. We validate our predictor both on the known operons of E.coli and on the genome of B.theta, using expression data to evaluate our predictions in the latter.

Availability: The software is available online at http://www.cse.wustl.edu/~jbuhler/research/operons

Contact:  jbuhler@cse.wustl.edu
NetAcet: prediction of N-terminal acetylation sites
Authors: Kiemer Lars , Bendtsen Jannick Dyrløv , Blom Nikolaj
Published: November 2004

Summary: We present here a neural network based method for prediction of N-terminal acetylation—by far the most abundant post-translational modification in eukaryotes. The method was developed on a yeast dataset for N-acetyltransferase A (NatA) acetylation, which is the type of N-acetylation for which most examples are known and for which orthologs have been found in several eukaryotes. We obtain correlation coefficients close to 0.7 on yeast data and a sensitivity up to 74% on mammalian data, suggesting that the method is valid for eukaryotic NatA orthologs.

Availability: The NetAcet prediction method is available as a public web server at http://www.cbs.dtu.dk/services/NetAcet/

Contact:  nikob@cbs.dtu.dk

Supplementary information:  http://www.cbs.dtu.dk/services/NetAcet/
Detection of low level genomic alterations by comparative genomic hybridization based on cDNA micro-arrays
Authors: Bilke Sven , Chen Qing-Rong , Whiteford Craig C. , Khan Javed
Published: November 2004

Motivation: The accumulation of genomic alterations is an important process in tumor formation and progression. Comparative genomic hybridization performed on cDNA arrays (cDNA aCGH) is a common method to investigate the genomic alterations on a genome-wide scale. However, when detecting low-level DNA copy number changes this technology requires the use of noise reduction strategies due to a low signal to noise ratio.

Results: Currently a running average smoothing filter is the most frequently used noise reduction strategy. We analyzed this strategy theoretically and experimentally and found that it is not sensitive to very low level genomic alterations. The presence of systematic errors in the data is one of the main reasons for this failure. We developed a novel algorithm which efficiently reduces systematic noise and allows for the detection of low-level genomic alterations. The algorithm is based on comparison of the biological relevant data to data from so-called self–self hybridizations, additional experiments which contain no biological information but contain systematic errors. We find that with our algorithm the effective resolution for ±1 DNA copy number changes is about 2 Mb. For copy number changes larger than three the effective resolution is on the level of single genes.

Contact:  bilkes@mail.nih.gov
Bicarbonate-Induced Alkalinization of the Xylem Sap in Intact Maize Seedlings as Measured in Situ with a Novel Xylem pH Probe
Authors: Wegner Lars H. , Zimmermann Ulrich
Published: November 2004

In higher plants the pH of the xylem sap plays an important role in drought signaling, growth regulation, and plant nutrition. However, the interpretation of the data is very controversial. The main reason for this is that the xylem pH in intact plants was not directly accessible hitherto. We present here a novel, minimally-invasive probe based on the xylem pressure-potential probe (used for measuring directly xylem pressure and the electrical potential between root xylem sap and medium). Single-tipped, double-barreled capillaries were used, one barrel served as H+-selective electrode, whereas pressure and electrical potential were recorded by the other one. Upon insertion of the probe into the root xylem of maize (Zea mays) seedlings, pH values ranging between about 4.2 and 4.9 were monitored when the roots were immersed in standard nutrient solution. The pH did not respond to changes in light irradiation (up to 300 μmol m−2 s−1), but increased upon exposure of the root to 5 or 20 mm bicarbonate in the bath solution. Changes in pH could also be recorded in transpiring plants when the root was cut below the insertion point of the probe and placed in media with different pH. The data support the hypothesis of Mengel ([1994] Plant Soil 165: 275–283) that upon external supply with bicarbonate Fe is immobilized in the leaf apoplast via changes in xylem pH.
Apoplastic Binding of Aluminum Is Involved in Silicon-Induced Amelioration of Aluminum Toxicity in Maize
Authors: Wang Yunxia , Stass Angelika , Horst Walter J.
Published: November 2004

The alleviating effect of silicon (Si) supply on aluminum (Al) toxicity was suggested to be based on ex or in planta mechanisms. In our experiments with the Al-sensitive maize (Zea mays) cultivar Lixis, Si treatment but not Si pretreatment ameliorated Al-induced root injury as revealed by less root-growth inhibition and callose formation. Si treatment did not affect monomeric Al concentrations in the nutrient solution, suggesting an in planta effect of Si on Al resistance. A fractionated analysis of Si and Al in the 1-cm root apices revealed that more than 85% of the root-tip Al was bound in the cell wall. Al contents in the apoplastic sap, the symplastic sap, and the cell wall did not differ between −Si and +Si plants. Si did not affect the Al-induced exudation of organic acid anions and phenols from the root apices. However, Al treatment greatly enhanced Si accumulation in the cell wall fraction, reducing the mobility of apoplastic Al. From our data we conclude that Si treatment leads to the formation of hydroxyaluminumsilicates in the apoplast of the root apex, thus detoxifying Al.
Pharmacological Evidence That Multiple Phospholipid Signaling Pathways Link Rhizobium Nodulation Factor Perception in Medicago truncatula Root Hairs to Intracellular Responses, Including Ca2+ Spiking and Specific ENOD Gene Expression
Authors: Charron Dorothée , Pingret Jean-Luc , Chabaud Mireille , Journet Etienne-Pascal , Barker David G.
Published: November 2004

Rhizobium nodulation (Nod) factors are specific lipochito-oligosaccharide signals essential for initiating in root hairs of the host legume developmental responses that are required for controlled entry of the microsymbiont. In this article, we focus on the Nod factor signal transduction pathway leading to specific and cell autonomous gene activation in Medicago truncatula cv Jemalong in a study making use of the Nod factor-inducible MtENOD11 gene. First, we show that pharmacological antagonists that interfere with intracellular ion channel and Ca2+ pump activities are efficient blockers of Nod factor-elicited pMtENOD11-β-glucuronidase (GUS) expression in root hairs of transgenic M. truncatula. These results indicate that intracellular Ca2+ release and recycling activities, essential for Ca2+ spiking, are also required for specific gene activation. Second, pharmacological effectors that inhibit phospholipase D and phosphoinositide-dependent phospholipase C activities are also able to block pMtENOD11-GUS activation, thus underlining a central role for multiple phospholipid signaling pathways in Nod factor signal transduction. Finally, pMtENOD11-GUS was introduced into all three Nod−/Myc−dmi M. truncatula mutant backgrounds, and gene expression was evaluated in response to the mastoparan peptide agonist Mas7. We found that Mas7 elicits root hair MtENOD11 expression in dmi1 and dmi2 mutants, but not in the dmi3 mutant, suggesting that the agonist acts downstream of DMI1/DMI2 and upstream of DMI3. In light of these results and the recently discovered identities of the DMI gene products, we propose an integrated cellular model for Nod factor signaling in legume root hairs in which phospholipids play a key role in linking the Nod factor perception apparatus to downstream components such as Ca2+ spiking and ENOD gene expression.
From imaging to knowledge
Authors: Hollingum Jack
Published: November 2004

This article summarises some of the presentations at a recent conference on the emerging technology of intelligent imaging.
VisionSensor™ ‐ a unique solution to particle size distribution analysis
Authors: Jakobsen Bo T.
Published: November 2004

Vision technology is gaining more and more acknowledgement in the process industry as an effective way for a company to ensure high quality of production. After an introduction to vision technology, we will give a deeper description of the VisionSensor™, which is uniquely designed for on‐line integration in real‐world industrial environments. The description of the VisionSensor™ will encompass where the system can be used, how it works, comparing its merits with alternative techniques and finally a description of two case stories from the sugar and fertiliser industries.
Optical pressure and temperature sensors for aerospace applications
Authors: Pinnock Robert A.
Published: November 2004

This paper gives a brief overview of requirements within the aerospace market sector for which optical sensors are of potential benefit, and goes on to describe sensors currently under development at Lucas Electrical and Electronic Systems which are primarily intended to meet some of these requirements. These sensors, based on the combination of silicon micromachining and optical sensing technologies, are sufficiently robust to provide the capability of directly measuring on‐engine parameters such as pressure and temperature. In association with FADEC‐mounted interface units, to which the sensor heads are coupled via optical fibre links, the sensors have the potential to provide measurement data for a number of aero‐engine control requirements.
Conoscopic probes are set to transform industrial metrology
Authors: Sirat Gabriel , Paz Freddy
Published: November 2004

Reports on developments at Optimet in the R&D Group, where a novel distance probe performs high accuracy distance measurement over broad length scales. Considers applications.
The application of reverse engineering in rapid product development
Authors: Bradley Colin
Published: November 2004

The process of capturing object form through surface data sampling and generating a CAD model of the part is termed reverse engineering because the process is the opposite of the normal design and manufacturing sequence. In the context of general manufacturing methods, reverse engineering is an important process for instances where a product initially exists as a designer’s model in a medium such as styling foam or modelling clay. The digitisation process can be achieved through spatial measurements taken manually by a co‐ordinate measuring machine (CMM). However, 3D computer vision systems are now being applied to the reverse engineering task owing to their ability to rapidly digitise the more intricate and free‐form surface patches that are common in many modern consumer items. New modelling software reduces the large number of data points into a more manageable number which can be translated into CAD drawings. The CAD drawings permit generation of CNC machine tool cutter paths or production of stereolithography “masters”.
Online focus control sensor for laser welding offers improved reliability without process intrusion
Authors: Hand Duncan P. , Peters Chris , Jones Julian
Published: November 2004

A robust and non‐intrusive focus control system for laser welding has been developed under a collaboration between GSI Lumonics (Rugby) and Heriot‐Watt University in Edinburgh, as part of the Laser Engineering for Manufacturing Applications Programme (LEMA), funded by EPSRC. It uses the light generated by the process, together with the chromatic aberrations of the focusing optics to determine the focal position of the laser relative to the workpiece. Successful closed loop operation has been demonstrated over a wide range of welding conditions, including different materials and laser powers.
Conductive polymer‐carbon black composites‐based sensor arrays for use in an electronic nose
Authors: Muñoz Beth C. , Steinthal Gregory , Sunshine Steven
Published: November 2004

Polymer‐carbon black composites are a new class of chemical detecting sensors used in electronic noses. These composites are prepared by mixing carbon black and polymer in an appropriate solvent. The mixture is deposited on a substrate between two metal electrodes, whereby the solvent evaporates leaving a composite film. Arrays of these chemiresistors, made from a chemically diverse number of polymers and carbon black, swell reversibly, inducing a resistance change on exposure to chemical vapors. These arrays generate a pattern that is a unique fingerprint for the vapor being detected. With the aid of algorithms these patterns are processed and recognized. These arrays can detect and discriminate between a large number of chemical vapors.
Resident participation in the evaluation of external accessibility requirements in housing estates
Authors: Kane Gillian , Heaney George , McGreal Stanley
Published: November 2004

Accessibility is now a major issue in the design and maintenance of housing estates. This paper considers the reasons why this issue has emerged and the main obstacles to accessibility encountered by housing estate residents. Focusing on a mature estate in South Belfast, Northern Ireland, investigates the obstacles to accessibility experienced by the residents in order to produce a hierarchy of impacts. Residents were surveyed on a number of issues including location, roads and pavements, transportation, level change, wayfinding, safety and security, and public areas. It was discovered that obstacles relating to wayfinding created the greatest problems for residents; however, attitudes towards accessibility varied according to the age of the respondent. The study concludes that accessibility should be introduced estate‐by‐estate dependent on a resident profile in order to maximise resources and advocates the adoption of a co‐ordinated interagency approach.
Human resource strategies and organisational structures for managing gold‐collar workers
Authors: Holland Peter J. , Hecker Robert , Steen John
Published: November 2004

States that the rapid expansion of the information technology (IT) industry has brought about skill shortages in many advanced western economies. Examines human resource (HR) policies and organisational structures for developing and retaining IT staff through two case studies of organisations in the IT and telecommunications sector in Australia. Concludes that developing HR policies in order to recruit and retain staff, and linking these to appropriate organisational structure, is becoming of increased importance in order to encourage employees to remain with a company.
Interactive TV and Formula One: a strategic issue for engine suppliers
Authors: Dransfeld Henning , Jacobs Gabriel , Dowsland William
Published: November 2004

During the last two years, the first experiments in digital interactive TV have been carried out, and full digital services are due to become available in many European states and elsewhere in the world within the next few years. These services will create a range of novel marketing opportunities, and Formula One Grand Prix would appear to offer significant potential in this respect. This paper reviews recent developments in the digital TV marketplace, focuses on how these may impact on Formula One, and suggests that production‐car manufacturers who also supply engines for the sport should now seriously consider taking advantage of the emerging medium.
Preceding muscle activity influences motor unit discharge and rate of torque development during ballistic contractions in humans
Authors: Van Cutsem Michaël , Duchateau Jacques
Published: November 2004

To investigate the effect of initial conditions on the modulation of motor unit discharge during fast voluntary contractions, we compared ballistic isometric contractions of the ankle dorsiflexor muscles that were produced from either a resting state or superimposed on a sustained contraction. The torque of the dorsiflexors and the surface and intramuscular EMGs from the tibialis anterior were recorded. The results showed that the performance of a ballistic contraction from a sustained contraction (∼25% maximal voluntary contraction (MVC)) had a negative effect on the maximal rate of torque development. Although the electromechanical delay was shortened, the EMG activity during the ballistic contraction was less synchronized. These observations were associated with a significant decline in the average discharge rate of single motor units (89.8 ± 3.8 versus 115 ± 5.8 Hz) and in the percentage of units (6.2 versus 15.5% of the whole sample) that exhibited double discharges at brief intervals (= 5 ms). High‐threshold units that were not recruited during the sustained contraction displayed the same activation pattern, which indicates that the mechanisms responsible for the decline in discharge rate were not restricted to previously activated units, but appear to influence the entire motor unit pool. When a premotor silent period (SP) was observed at the transition from the sustained muscular activity to the ballistic contraction (19% of the trials), these adjustments in motor unit activity were not present, and the ballistic contractions were similar to those performed from a resting state. Together, these results indicate that initial conditions can influence the capacity for motor unit discharge rate and hence the performance of a fast voluntary contraction.
Exogenous nitric oxide inhibits sympathetically mediated vasoconstriction in human skin
Authors: Durand S. , Davis S. L. , Cui J. , Crandall C. G.
Published: November 2004

Two experiments were performed to identify whether nitric oxide (NO) inhibits sympathetically mediated vasoconstriction in human skin. In eight subjects increasing doses of sodium nitroprusside (SNP; 8.4 × 10−6–8.4 × 10−3m) were administered via intradermal microdialysis. At each dose of SNP, cutaneous vasoconstrictor responsiveness was assessed during a 3 min whole‐body cold stress. The relative reduction in forearm cutaneous vascular conductance (CVC) during the cold stress was significantly attenuated for SNP doses greater than 8.4 × 10−4m (control: 63.0 ± 4.1%, SNP 8.4 × 10−6m: 57.1 ± 4.7%, SNP 8.4 × 10−5m: 57.0 ± 3.6%, SNP 8.4 × 10−4m: 44.5 ± 5.4% and SNP 8.4 × 10−3m: 28.8 ± 7.9%). The second experiment was performed to identify whether this response was due to NO attenuating sympathetically mediated vasoconstriction or due to a non‐specific effect of an elevated CVC secondary to SNP administration. In seven subjects forearm CVC during a whole‐body cold stress was assessed at two sites: at a site dilated via microdialysis administration of SNP and at a site dilated with isoproterenol (ISO). CVC was not different between sites prior to (SNP: 0.42 ± 0.11; ISO: 0.46 ± 0.11 AU mmHg−1 (AU, arbitrary units), P > 0.05) or following drug infusion (SNP: 1.36 ± 0.21; ISO: 1.27 ± 0.23 AU mmHg−1, P > 0.05). The reduction in CVC during the subsequent cold stress was significantly less at the SNP site (38.1 ± 6.2%) relative to the ISO site (65.0 ± 5.5%; P= 0.007). These data suggest NO is capable of inhibiting sympathetically mediated vasoconstriction in the cutaneous vasculature.
The kainate receptor subunit GluR6 mediates metabotropic regulation of the slow and medium AHP currents in mouse hippocampal neurones
Authors: Fisahn André , Heinemann Stephen F. , McBain Chris J.
Published: November 2004

Kainate receptors (KARs) play an important role in synaptic physiology, plasticity and pathological phenomena such as epilepsy. However, the physiological implications for single cells and neuronal networks of the distinct expression patterns of KAR subunits are unknown. One intriguing effect of KAR activation is a long‐term change to intrinsic neuronal excitability and neuronal firing patterns, such as single‐spike and spike‐burst firing. In this study, we describe the role of kainate receptor subunits in the metabotropic regulation of the slow and medium afterhyperpolarization (AHP) currents (IsAHP, ImAHP). Using whole‐cell patch‐clamp recordings from CA3 pyramidal cells of wild‐type (WT) and KAR knockout mice, we show that the kainate‐induced decrease of IsAHP and ImAHP amplitude is protein‐kinase‐C‐dependent and absent in GluR6−/− but not GluR5−/− pyramidal neurones. Our findings suggest that activation of GluR6‐containing KARs modulates AHP amplitude, and influences the firing frequency of pyramidal neurones.
A billiard-theoretic approach to elementary one-dimensional elastic collisions
Authors: Redner S.
Published: November 2004

A simple relation is developed between the elastic collisions of freely moving particles in one dimension and a corresponding billiard system. For two particles with masses m1 and m2 on the half-line x>0 that approach an elastic barrier at x=0, the corresponding billiard system is an infinite wedge. The collision history of the two particles can be easily inferred from the corresponding billiard trajectory. This connection explains the classic demonstrations of the “dime on the superball” and the “baseball on the basketball” that are a staple in elementary physics courses. It also is shown that three elastic particles on an infinite line and three particles on a finite ring correspond, respectively, to the motion of a billiard ball in an infinite wedge and on a triangular billiard table. It is shown how to determine the angles of these two sets in terms of the particle masses.
Simplified derivation of the Hawking–Unruh temperature for an accelerated observer in vacuum
Authors: Alsing Paul M. , Milonni Peter W.
Published: November 2004

A detector undergoing uniform acceleration a in a vacuum field responds as though it were immersed in thermal radiation of temperature T=ℏa/2πkc. An intuitive derivation of this result is given for a scalar field in one spatial dimension. The approach is extended to the case where the field detected by the accelerated observer is a spin 1/2 Dirac field.
Challenges to Faraday’s flux rule
Authors: Munley Frank
Published: November 2004

Faraday’s law (or flux rule) is beautiful in its simplicity, but difficulties are often encountered when applying it to specific situations, particularly those where points making contact to extended conductors move over finite time intervals. These difficulties have led some to challenge the generality of the flux rule. The challenges are usually coupled with the claim that the Lorentz force law is general, even though proofs have been given of the equivalence of the two for calculating instantaneous emfs in well-defined filamentary circuits. I review a rule for applying Faraday’s law, which says that the circuit at any instant must be fixed in a conducting material and must change continuously. The rule still leaves several choices for choosing the circuit. To explicate the rule, it will be applied to several challenges, including one by Feynman.
Nef Associates with p21-Activated Kinase 2 in a p21-GTPase-Dependent Dynamic Activation Complex within Lipid Rafts
Authors: Pulkkinen Kati , Renkema G. Herma , Kirchhoff Frank , Saksela Kalle
Published: November 2004

We have previously reported that Nef specifically interacts with a small but highly active subpopulation of p21-activated kinase 2 (PAK2). Here we show that this is due to a transient association of Nef with a PAK2 activation complex within a detergent-insoluble membrane compartment containing the lipid raft marker GM1. The low abundance of this Nef-associated kinase (NAK) complex was found to be due to an autoregulatory mechanism. Although activation of PAK2 was required for assembly of the NAK complex, catalytic activity of PAK2 also promoted dissociation of this complex. Testing different constitutively active PAK2 mutants indicated that the conformation associated with p21-mediated activation rather than kinase activity per se was required for PAK2 to become NAK. Although association with PAK2 is one of the most conserved properties of Nef, we found that the ability to stimulate PAK2 activity differed markedly among divergent Nef alleles, suggesting that PAK2 association and activation are distinct functions of Nef. However, mutations introduced into the p21-binding domain of PAK2 revealed that p21-GTPases are involved in both of these Nef functions and, in addition to promoting PAK2 activation, also help to physically stabilize the NAK complex.
A Cyclin-Binding Motif within the Amino-Terminal Homology Domain of EBNA3C Binds Cyclin A and Modulates Cyclin A-Dependent Kinase Activity in Epstein-Barr Virus-Infected Cells
Authors: Knight Jason S. , Sharma Nikhil , Kalman Danielle E. , Robertson Erle S.
Published: November 2004

The Epstein-Barr virus (EBV) nuclear antigen 3C (EBNA3C) is a virus-encoded latent antigen essential for primary B-cell transformation. In this report we demonstrate that although the carboxy terminus of EBNA3C predominantly regulates cyclin A-dependent kinase activity, the region of greatest affinity for cyclin A lies within the EBNA3 amino-terminal homology domain of EBNA3C. Detailed mapping studies employing both in vitro binding assays and coimmunoprecipitation experiments implicated a small region of EBNA3C, amino acids 130 to 159 within the EBNA3 homology domain, as having the greatest affinity for cyclin A. The EBNA3 homology domain has the highest degree of amino acid similarity (approximately 30%) between the EBNA3 proteins, and, indeed, EBNA3B, but not EBNA3A, showed binding activity with cyclin A. We also show that EBNA3C binds to the α1 helix of the highly conserved mammalian cyclin box, with cyclin A amino acids 206 to 226 required for strong binding to EBNA3C amino acids 130 to 159. Interestingly, EBNA3C also bound human cyclins D1 and E in vitro, although the affinity was approximately 30% of that seen for cyclin A. Previously it was demonstrated that full-length EBNA3C rescues p27-mediated suppression of cyclin A-dependent kinase activity (J. S. Knight and E. S. Robertson, J. Virol.
78:
1981-1991, 2004). It was also demonstrated that the carboxy terminus of EBNA3C recapitulates this phenotype. Surprisingly, the amino terminus of EBNA3C with the highest affinity for cyclin A was unable to rescue p27 suppression of kinase activity and actually downregulates cyclin A activity when introduced into EBV-infected cells. The data presented here suggests that the amino terminus of EBNA3C may play an important role in recruiting cyclin A complexes, while the carboxy terminus of EBNA3C is necessary for the functional modulation of cyclin A complex kinase activity.
Requirement of Sur2 for Efficient Replication of Mouse Adenovirus Type 1
Authors: Fang Lei , Stevens Jennitte L. , Berk Arnold J. , Spindler Katherine R.
Published: November 2004

Mouse adenovirus type 1 (MAV-1) early region 1A (E1A) encodes a virulence gene in viral infection of mice. To broaden our understanding of the functions of E1A in MAV-1 pathogenesis, an unbiased experimental approach, glutathioneS-transferase (GST) pulldown, was used to screen for cellular proteins that interact with E1A protein. We identified mouse Sur2, a subunit of Mediator complex, as a protein that binds to MAV-1 E1A. The interaction between Sur2 and MAV-1 E1A was confirmed in virus-infected cells. Conserved region 3 (CR3) of MAV-1 E1A was mapped as the region required for Sur2-E1A interaction, as is the case for human adenovirus E1A. Although it has been proposed that human adenovirus E1A recruits the Mediator complex to transactivate transcription of viral early genes, Sur2 function in adenovirus replication has not been directly tested previously. Studies on the functions of Sur2 with mouse embryonic fibroblasts (MEFs) showed that there was a multiplicity-dependent growth defect of MAV-1 in Sur2−/−MEFs compared to Sur2+/+MEFs. Comparison of the viral DNA and viral mRNA levels in Sur2+/+and Sur2−/−MEFs confirmed that Sur2 was important for efficient viral replication. The viral replication defects in Sur2−/−MEFs appeared to be due at least in part to a defect in viral early gene transcription.
Cyclooxygenase-1 and -2 Are Required for Production of Infectious Pseudorabies Virus
Authors: Ray Neelanjana , Bisher Margaret E. , Enquist L. W.
Published: November 2004

We have recently shown that cyclooxygenase-2 (COX-2) transcription is markedly induced after herpes simplex virus type 1 and pseudorabies virus (PRV) infections of rat embryonic fibroblast (REF) cells (N. Ray and L. W. Enquist, J. Virol.78:3489-3501, 2004). For this study, we investigated the role of cyclooxygenase induction in the replication and growth of PRV. We demonstrate here a concordant increase in COX-2 mRNA and protein levels after the infection of REF cells. Inhibitors blocking the activity of cyclooxygenases caused a dramatic reduction in PRV growth. Viral growth could be restored if prostaglandin E2, the final product of COX-2 activity, was added simultaneously with the COX inhibitors. Immediate-early protein IE180, major capsid protein VP5, and glycoprotein expression were slightly reduced in the presence of COX-2 inhibitors, but expression of the early protein EP0 was not affected by COX inhibition. Viral DNA replication was marginally reduced in the presence of a COX-1/2 inhibitor, but there was no defect in viral DNA cleavage. Electron microscopy analysis revealed an increased number of unusual empty capsid structures in the nuclei of cells infected with PRV in the presence of a COX-1/2 inhibitor. These capsid structures shared some characteristics with procapsids but had a novel appearance by negative staining. Our data establish a role for COX-1 and COX-2 in facilitating the efficient growth and replication of PRV in primary cells.
Characterization of the Outer Domain of the gp120 Glycoprotein from Human Immunodeficiency Virus Type 1
Authors: Yang Xinzhen , Tomov Vesko , Kurteva Svetla , Wang Liping , Ren Xinping , Gorny Miroslaw K. , Zolla-Pazner Susan , Sodroski Joseph
Published: November 2004

The core of the gp120 glycoprotein from human immunodeficiency virus type 1 (HIV-1) is comprised of three major structural domains: the outer domain, the inner domain, and the bridging sheet. The outer domain is exposed on the HIV-1 envelope glycoprotein trimer and contains binding surfaces for neutralizing antibodies such as 2G12, immunoglobulin G1b12, and anti-V3 antibodies. We expressed the outer domain of HIV-1YU2gp120 as an independent protein, termed OD1. OD1 efficiently bound 2G12 and a large number of anti-V3 antibodies, indicating its structural integrity. Immunochemical studies with OD1 indicated that antibody responses against the outer domain of the HIV-1 gp120 envelope glycoprotein are rare in HIV-1-infected human sera that potently neutralize the virus. Surprisingly, such outer-domain-directed antibody responses are commonly elicited by immunization with recombinant monomeric gp120. Immunization with soluble, stabilized HIV-1 envelope glycoprotein trimers elicited antibody responses that more closely resembled those in the sera of HIV-1-infected individuals. These results underscore the qualitatively different humoral immune responses elicited during natural infection and after gp120 vaccination and help to explain the failure of gp120 as an effective vaccine.
Differential Rescue of Poliovirus RNA Replication Functions by Genetically Modified RNA Polymerase Precursors
Authors: Cornell Christopher T. , Brunner Jo Ellen , Semler Bert L.
Published: November 2004

We have previously described the RNA replication properties of poliovirus transcripts harboring chimeric RNA polymerase sequences representing suballelic exchanges between poliovirus type 1 (PV1) and coxsackievirus B3 (CVB3) utilizing an in vitro translation and RNA replication assay (C. Cornell, R. Perera, J. E. Brunner, and B. L. Semler, J. Virol.
78:
4397-4407, 2004). We showed that three of the seven chimeras were capable of RNA replication in vitro, although replication levels were greatly reduced compared to that of wild-type transcripts. Interestingly, one of the replication-competent transcripts displayed a strand-specific RNA synthesis defect suggesting (i) a differential replication complex assembly mechanism involving 3D and/or precursor molecules (i.e., 3CD) required for negative- versus positive-strand RNA synthesis or (ii) effect(s) on the ability of the 3D polymerase to form higher-ordered structures required for positive-strand RNA synthesis. In this study, we have attempted to rescue defective RNA replication in vitro by cotranslating nonstructural proteins from a transcript encoding a large precursor polyprotein (P3) to complement 3D polymerase and/or precursor polypeptide functions altered in each of the chimeric constructs. Utilization of a wild-type P3 construct revealed that all transcripts containing chimeric PV1/CVB3 polymerase sequences can be complemented in
trans
for both negative- and positive-strand RNA synthesis. Furthermore, data from experiments utilizing genetically modified forms of the P3 polyprotein, containing mutations within 3C or 3D sequences, strongly suggest the existence of different protein-protein and protein-RNA interactions required for positive- versus negative-strand RNA synthesis. These results, combined with data from in vitro RNA elongation assays, indicate that the delivery of active 3D RNA polymerase to replication complexes requires a series of macromolecular interactions that rely on the presence of specific 3D amino acid sequences.
Structural Protein Requirements in Equine Arteritis Virus Assembly
Authors: Wieringa Roeland , de Vries Antoine A. F. , van der Meulen Jannes , Godeke Gert-Jan , Onderwater Jos J. M. , van Tol Hans , Koerten Henk K. , Mommaas A. Mieke , Snijder Eric J. , Rottier Peter J. M.
Published: November 2004

Equine arteritis virus (EAV) is an enveloped, positive-stranded RNA virus belonging to the family
Arteriviridae
of the order
Nidovirales
. EAV particles contain seven structural proteins: the nucleocapsid protein N, the unglycosylated envelope proteins M and E, and the N-glycosylated membrane proteins GP
2b
(previously named G
S
), GP
3
, GP
4
, and GP
5
(previously named G
L
). Proteins N, M, and GP
5
are major virion components, E occurs in virus particles in intermediate amounts, and GP
4
, GP
3
, and GP
2b
are minor structural proteins. The M and GP
5
proteins occur in virus particles as disulfide-linked heterodimers while the GP
4
, GP
3
, and GP
2b
proteins are incorporated into virions as a heterotrimeric complex. Here, we studied the effect on virus assembly of inactivating the structural protein genes one by one in the context of a (full-length) EAV cDNA clone. It appeared that the three major structural proteins are essential for particle formation, while the other four virion proteins are dispensable. When one of the GP
2b
, GP
3
, or GP
4
proteins was missing, the incorporation of the remaining two minor envelope glycoproteins was completely blocked while that of the E protein was greatly reduced. The absence of E entirely prevented the incorporation of the GP
2b
, GP
3
, and GP
4
proteins into viral particles. EAV particles lacking GP
2b
, GP
3
, GP
4
, and E did not markedly differ from wild-type virions in buoyant density, major structural protein composition, electron microscopic appearance, and genomic RNA content. On the basis of these results, we propose a model for the EAV particle in which the GP
2b
/GP
3
/GP
4
heterotrimers are positioned, in association with a defined number of E molecules, above the vertices of the putatively icosahedral nucleocapsid.
Expression and Characterization of a Soluble Form of
Tomato Spotted Wilt Virus
Glycoprotein G
N
Authors: Whitfield Anna E. , Ullman Diane E. , German Thomas L.
Published: November 2004

Tomato spotted wilt virus
(TSWV), a member of the
Tospovirus
genus within the
Bunyaviridae
, is an economically important plant pathogen with a worldwide distribution. TSWV is transmitted to plants via thrips (Thysanoptera: Thripidae), which transmit the virus in a persistent propagative manner. The envelope glycoproteins, G
N
and G
C
, are critical for the infection of thrips, but they are not required for the initial infection of plants. Thus, it is assumed that the envelope glycoproteins play important roles in the entry of TSWV into the insect midgut, the first site of infection. To directly test the hypothesis that G
N
plays a role in TSWV acquisition by thrips, we expressed and purified a soluble, recombinant form of the G
N
protein (G
N
-S). The expression of G
N
-S allowed us to examine the function of G
N
in the absence of other viral proteins. We detected specific binding to thrips midguts when purified G
N
-S was fed to thrips in an in vivo binding assay. The TSWV nucleocapsid protein and human cytomegalovirus glycoprotein B did not bind to thrips midguts, indicating that the G
N
-S-thrips midgut interaction is specific. TSWV acquisition inhibition assays revealed that thrips that were concomitantly fed purified TSWV and G
N
-S had reduced amounts of virus in their midguts compared to thrips that were fed TSWV only. Our findings that G
N
-S binds to larval thrips guts and decreases TSWV acquisition provide evidence that G
N
may serve as a viral ligand that mediates the attachment of TSWV to receptors displayed on the epithelial cells of the thrips midgut.
Membrane Association of the RNA-Dependent RNA Polymerase Is Essential for Hepatitis C Virus RNA Replication
Authors: Moradpour Darius , Brass Volker , Bieck Elke , Friebe Peter , Gosert Rainer , Blum Hubert E. , Bartenschlager Ralf , Penin François , Lohmann Volker
Published: November 2004

The hepatitis C virus (HCV) RNA-dependent RNA polymerase (RdRp), represented by nonstructural protein 5B (NS5B), belongs to a class of integral membrane proteins termed tail-anchored proteins. Its membrane association is mediated by the C-terminal 21 amino acid residues, which are dispensable for RdRp activity in vitro. For this study, we investigated the role of this domain, termed the insertion sequence, in HCV RNA replication in cells. Based on a structural model and the amino acid conservation among different HCV isolates, we designed a panel of insertion sequence mutants and analyzed their membrane association and RNA replication. Subgenomic replicons with a duplication of an essential
cis
-acting replication element overlapping the sequence that encodes the C-terminal domain of NS5B were used to unequivocally distinguish RNA versus protein effects of these mutations. Our results demonstrate that the membrane association of the RdRp is essential for HCV RNA replication. Interestingly, certain amino acid substitutions within the insertion sequence abolished RNA replication without affecting membrane association, indicating that the C-terminal domain of NS5B has functions beyond serving as a membrane anchor and that it may be involved in critical intramembrane protein-protein interactions. These results have implications for the functional architecture of the HCV replication complex and provide new insights into the expanding spectrum of tail-anchored proteins.
Hepatitis Delta Virus Antigen Is Methylated at Arginine Residues, and Methylation Regulates Subcellular Localization and RNA Replication
Authors: Li Yi-Jia , Stallcup Michael R. , Lai Michael M. C.
Published: November 2004

Hepatitis delta virus (HDV) contains a circular RNA which encodes a single protein, hepatitis delta antigen (HDAg). HDAg exists in two forms, a small form (S-HDAg) and a large form (L-HDAg). S-HDAg can transactivate HDV RNA replication. Recent studies have shown that posttranslational modifications, such as phosphorylation and acetylation, of S-HDAg can modulate HDV RNA replication. Here we show that S-HDAg can be methylated by protein arginine methyltransferase (PRMT1) in vitro and in vivo. The major methylation site is at arginine-13 (R13), which is in the RGGR motif of an RNA-binding domain. The methylation of S-HDAg is essential for HDV RNA replication, especially for replication of the antigenomic RNA strand to form the genomic RNA strand. An R13A mutation in S-HDAg inhibited HDV RNA replication. The presence of a methylation inhibitor,
S
-adenosyl-homocysteine, also inhibited HDV RNA replication. We further found that the methylation of S-HDAg affected its subcellular localization. Methylation-defective HDAg lost the ability to form a speckled structure in the nucleus and also permeated into the cytoplasm. These results thus revealed a novel posttranslational modification of HDAg and indicated its importance for HDV RNA replication. This and other results further showed that, unlike replication of the HDV genomic RNA strand, replication of the antigenomic RNA strand requires multiple types of posttranslational modification, including the phosphorylation and methylation of HDAg.