bluuebunny/crossref_metadata_2025 huggingface dataset:
Information Technology and the Organization of Firms
Authors: Marschak Thomas
Published: July 2004

This paper seeks to understand why improved information technology (IT) might strengthen the case for decentralization, as recent empirical work suggests. We study a firm with a headquarters and two managers, each of whom gathers information about her changing local environment. The firm earns a gross profit that depends on actions taken as well as the current local environments. More information permits better actions, and information‐gathering costs drop as IT improves. When the firm is centralized, information‐gathering expenditures are first best, but after the firm decentralizes, each manager becomes a self‐interested player of a “sharing game” in which she collects a share of gross profit and bears the cost of her chosen information‐gathering activities. The firm's actions are determined by the information gathered at the equilibria of the game. As a result, the firm experiences a decentralization penalty, namely the change in net profit (gross profit minus informational costs) after decentralizing. If the penalty is small, then it is outweighed by the advantages of decentralizing—the vanishing of monitoring costs and perhaps the improved motivation of a decentralized manager's staff. To gather information a manager chooses (once and for all) a partitioning of her possible local environments and then searches to find the set in which her current environment lies. Our main measure of a manager's information cost is a technology parameter, θ, times the number of sets in her chosen partitioning. A second measure is θ times the partitioning's “Shannon content,” which may be interpreted as average search time when search is efficient. We ask whether improved IT, i.e., a drop in θ, indeed lowers the decentralization penalty. We obtain a strongly affirmative answer to this question for both cost measures in a class of examples and a mixed answer when we generalize so as to preserve some of the key properties of those examples. In a parallel manner we explore another conjecture suggested in the empirical literature, namely that better IT raises the coordination benefit, which we define as the increase in net profit when the firm bases its actions on pooled information, rather than letting each action variable depend on the information gathered by just one manager.
Patient characteristics and quality of life among a sample of Australian chronic pain clinic attendees
Authors: Kerr S. , Fairbrother G. , Crawford M. , Hogg M. , Fairbrother D. , Khor K.E.
Published: July 2004

Abstract

Background : Multidisciplinary chronic pain management programs have proliferated widely in recent decades. The clinical characteristics of patients attending these clinics are becoming the subject of increased research. Recent European data suggests that patients attending these clinics report very low quality of life.

Aims:  The present study profiles an Australian popu­lation in terms of demographics, clinical characteristics and quality of life, as measured by the Short Form 36 Quality of Life Questionnaire (SF‐36).

Methods:  Data were collected prospectively from consecutive patients presenting to a multidisciplinary chronic pain clinic at a major Sydney metropolitan teaching ­hospital. Cross‐sectional analysis of demographic and clin­ical characteristics and quality of life were then undertaken.

Results:  Descriptive analysis of demographics and clin­ical characteristics suggest a patient population group reporting significant pain severity and reduced quality of life. The comparison of SF‐36 domain scores between clinic patients and Australian norm values indicates a greatly reduced score on all SF‐36 domains for clinic patients. Pain clinic patients reported the most profound effect upon quality of life in the role physical, physical function and social function domains of the SF‐36. Stepwise multiple regression indicated impaired coping ability and depressive disability as the most significant correlates of low quality of life.

Conclusion:  Patients who attend chronic pain clinics are likely to report low quality of life with an inability to cope. These findings suggest that future intervention research should explore the impacts of behavioural and self‐management interventions. Psychological distress and ability to cope could be used as indices of improvement. (Intern Med J 2004; 34: 403−409)
Repetitive strain injuries: has the Australian epidemic burnt out?
Authors: Awerbuch M.
Published: July 2004

Abstract

In the 1980s Australia experienced an epidemic of medically certified claims for non‐specific arm symptoms described as repetitive strain injury. Although a number of factors were mooted as causal of the epidemic, no single factor emerged as a compelling putative candidate. The present paper discusses the results of research which was published only after the epidemic had waned. It provides possible insights into the rise and fall of repetitive strain injury. (Intern Med J 2004; 34: 416−419)
‘The more things change…’: Barriers to community services utilisation in Queensland
Authors: McDonald Catherine , Zetlin Di
Published: July 2004

Community services are central to the lives of many elderly Australians or Australians with disabilities if they are to remain in the community. Over the past two decades, significant advances have been made in policy and associated programs, significantly improving the standards of service delivery. This research reports on the perceptions of service providers in community services for people with disabilities in six communities in Queensland. It illustrates that significant barriers to service utilisation remain, despite reform. It suggests that continuous efforts to promote service access need to be built in at the program level.
Becoming a social work supervisor: A significant role transition
Authors: Cousins Carolyn
Published: July 2004

The move from social work practitioner to supervisor can cause a crisis in identity for some social workers, a point at which professional values, roles and commitment are questioned and re‐analysed. Yet it is an area in which there appears to be little written to assist the worker to normalise their fears and anxieties, or to assist with making the process smoother for both supervisor and supervisee. Drawing on practice experience in a range of health and community service settings, the present study will examine some of the feelings, issues, challenges and dilemmas faced by new social work supervisors. It will also explore the preparation of supervisors, the use of power in the supervisory relationship and the need for training. It will then discuss tips and conditions for improving the supervisory relationship.
Promotion of hair‐growth with Laminaria angustata extracts
Authors: Osawa Y. , Tamaki S. , Sawaki S. , Tomita Y.
Published: July 2004

The mechanism of hair growth has been clarifying itself by the improvement in the culture techniques for hair follicle cells. Recently stem cells inducing the anagen phase in the hair follicle cycle have been discovered in the bulge region of the outer root sheath (ORS). To find growth‐promoting agents for the ORS cells, we evaluated the effect of various botanical extracts on the growth of cultured human hair follicles. We found that Laminaria angustata extract increased the ORS cell growth. Further, hair‐growth in the shaved skin of C3H mice was also promoted by the topical application of the extract.
Justice and Union Participation: An Extension and Test of Mobilization Theory
Authors: Johnson Nancy Brown , Jarley Paul
Published: July 2004

Heeding John Kelly's call to reorient the field of industrial relations towards the study of injustice, we employ mobilization, social exchange and organizational justice theories to specify and test a model of union participation. Using individual‐level survey data from public‐sector union members to test our model, we find that differences in worker perceptions of workplace injustice and union justice explain slightly more variation in members’ union participation than more traditional measures of job satisfaction and union instrumentality perceptions. The implications of our analysis for researchers and union practitioners are also discussed.
Space for cognition: gesture and second language learning
Authors: McCafferty Steven G.
Published: July 2004

This paper explores the possibility that L2 learners utilize gesture, like speech, for intrapersonal problem solving. Kita (2000) argued that the “default” organization of information when speaking is analytic but that the use of representational gestures, specifically iconic and abstract deictic gestures, can constitute a spatio‐motoric mode of thinking, that is, that we deploy “gesturing for thinking”. Thus, Kita believes that gestures are actional as opposed to representational and form as part of the physical environment in conjunction with the cognitive system. In applying this perspective to second language learning, it is argued in the current study that, because of an inability to fully command discourse in the L2, at times learners utilize this mode of thinking. There is also speculation that the use of gestures in the form of beats in relation to prosodic features, notably intonation and syllable structure, may be a form of enaction, that is, internalizing the L2 through embodied processes.
How chronic inflammation can affect the brain and support the development of Alzheimer's disease in old age: the role of microglia and astrocytes
Authors: Blasko Imrich , Stampfer‐Kountchev Michaela , Robatscher Peter , Veerhuis Robert , Eikelenboom Piet , Grubeck‐Loebenstein Beatrix
Published: July 2004

A huge amount of evidence has implicated amyloid beta (Aβ) peptides and other derivatives of the amyloid precursor protein (βAPP) as central to the pathogenesis of Alzheimer's disease (AD). It is also widely recognized that age is the most important risk factor for AD and that the innate immune system plays a role in the development of neurodegeneration. Little is known, however, about the molecular mechanisms that underlie age‐related changes of innate immunity and how they affect brain pathology. Aging is characteristically accompanied by a shift within innate immunity towards a pro‐inflammatory status. Pro‐inflammatory mediators such as tumour necrosis factor‐α or interleukin‐1β can then in combination with interferon‐γ be toxic on neurons and affect the metabolism of βAPP such that increased concentrations of amyloidogenic peptides are produced by neuronal cells as well as by astrocytes. A disturbed balance between the production and the degradation of Aβ can trigger chronic inflammatory processes in microglial cells and astrocytes and thus initiate a vicious circle. This leads to a perpetuation of the disease.
Stress responses and innate immunity: aging as a contributory factor
Authors: Butcher Stephen K. , Lord Janet M.
Published: July 2004

Evolutionary pressure has selected individuals with traits that allow them to survive to reproduction, without consideration of the consequences for the post‐child rearing years and old age. In the 21st century, society is populated increasingly by the elderly and with the falling birth rate and improved health care this trend is set to continue for the foreseeable future. To minimize the potential burden on health services one would hope that ‘growing old gracefully’ should also mean ‘growing old healthily’. However, for too many the aging process is accompanied by increasing physical and mental frailty producing an elevated risk of physical and psychological stress in old age. Stress is a potent modulator of immune function, which in youth can be compensated for by the presence of an optimal immune response. In the elderly the immune response is blunted as a result of the decline in several components of the immune system (immune senescence) and a shifting to a chronic pro‐inflammatory status (the so‐called ‘inflamm‐aging’ effect). We discuss here what is known of the effects of both stress and aging upon the innate immune system, focusing in particular upon the age‐related alterations in the hypopituitary–adrenal axis. We propose a double hit model for age and stress in which the age‐related increase in the cortisol/sulphated dehydroepiandrosterone ratio synergizes with elevated cortisol during stress to reduce immunity in the elderly significantly.
NK and NKT cell functions in immunosenescence
Authors: Mocchegiani Eugenio , Malavolta Marco
Published: July 2004

Immunosenescence is defined as the state of dysregulated immune function that contributes to the increased susceptibility to infection, cancer and autoimmune diseases observed in old organisms, including humans. However, dysregulations in the immune functions are normally counterbalanced by continuous adaptation of the body to the deteriorations that occur over time. These adaptive changes are likely to occur in healthy human centenarians. Both innate (natural) and adaptive (acquired) immune responses decline with advancing age. Natural killer (NK) and natural killer T (NKT) cells represent the best model to describe innate and adaptive immune response in aging. NK and NKT cell cytotoxicity decreases in aging as well as interferon‐γ (IFN‐γ) production by both activated cell types. Their innate and acquired immune responses are preserved in very old age. However, NKT cells bearing T‐cell receptor (TCR) γδ also display an increased cytotoxicity and IFN‐γ production in very old age. This fact suggests that NKT cells bearing TCRγδ are more involved in maintaining innate and adaptive immune response in aging leading to successful aging. The role played by the neuroendocrine–immune network and by nutritional factors, such as zinc, in maintaining NK and NKT cell functions in aging is discussed.
ALL THINGS CONSIDERED, TAXES DRIVE THE JANUARY EFFECT
Authors: Chen Honghui , Singal Vijay
Published: July 2004

The multitude of explanations for the January effect leaves the reader confused about its primary cause(s): is it tax‐loss selling, window dressing, information, bid‐ask bounce, or a combination of these causes? The confusion arises, in part, because evidence has generally been presented in support of a particular hypothesis though the same evidence may be consistent with another hypothesis. Furthermore, prior work does not adequately control for the bid‐ask bounce. In this article we try to disentangle different explanations of the January effect and identify its primary cause. We find that tax‐related selling is the most important cause, overshadowing other explanations.
TRADING COSTS AND QUOTE CLUSTERING ON THE NYSE AND NASDAQ AFTER DECIMALIZATION
Authors: Chung Kee H. , Van Ness Bonnie F. , Van Ness Robert A.
Published: July 2004

We examine execution costs and quote clustering on the New York Stock Exchange (NYSE) and NASDAQ using 517 matching pairs of stocks after decimalization. We find that the mean spread of NASDAQ stocks is greater than the mean spread of NYSE stocks when spreads are equally weighted across stocks, and the difference is greater for smaller stocks. In contrast, the mean NASDAQ spread is narrower than the mean NYSE spread when spreads are volume weighted, and the difference is statistically significant for large stocks. Both NYSE and NASDAQ stocks exhibit high degrees of quote clustering on nickels and dimes, and quote clustering has a significant effect on spreads in both markets.
Dental traumatology: essential diagnosis and treatment planning
Authors: Bakland Leif K. , Andreasen Jens Ove
Published: July 2004

Traumatic dental injuries are for the most part unanticipated events that, if not managed appropriately, can have serious consequences for the patient. The purpose of this review is to describe the current concepts in establishing diagnosis descriptive of specific traumatic entities, and to delineate recommended treatment approaches for these injuries based on available evidence.
Interappointment pain: mechanisms, diagnosis, and treatment
Authors: Siqueira J. F. , Barnett F.
Published: July 2004

Knowledge on the causes of and the mechanisms behind interappointment pain in endodontics is of utmost importance for the clinician to properly prevent or manage this undesirable condition. The causative factors of interappointment pain encompass mechanical, chemical, and/or microbial injury to the pulp or periradicular tissues, which are induced or exacerbated during root canal treatment. Microorganisms can participate in causation of interappointment pain in the following situations: apical extrusion of debris; incomplete instrumentation leading to changes in the endodontic microbiota or in environmental conditions; and secondary intraradicular infections. Interappointment pain is almost exclusively due to the development of acute inflammation at the periradicular tissues in response to an increase in the intensity of injury coming from the root canal system. When an interappointment emergency occurs, proper diagnosis and active treatment are required for the clinician to succeed in solving the problem. This review focuses on the mechanisms of interappointment pain, with special emphasis placed on the causative agents and the host response to injury that can precipitate pain. In addition, diagnostic measures and treatment approaches to manage interappointment pain are also discussed.
Advanced radiographic techniques for the detection of lesions in bone
Authors: Cotti Elisabetta , Campisi Girolamo
Published: July 2004

Radiographs in endodontics are of importance for the study and management of periapical lesions. With the development of advanced systems in traditional radiology, new and more accurate imaging techniques are constantly under investigation. Computerized tomography, magnetic resonance and real‐time echotomography have been introduced in recent years to the field of endodontics: they may have advantages over conventional techniques for the amount of detailed information they can provide on specific cases. This paper reviews these new imaging techniques with respect to their possible role in the diagnosis and management of periapical lesions.
Relativistic orbits with computer algebra
Authors: Wang Frank Y.-H.
Published: July 2004

We use computer algebra to derive the equations of motion from the Lagrangian, solve the equations of motion numerically, and plot the numerical solutions, to discover the difference between trajectories of a particle under gravity based on Newtonian theory and general relativity with the Schwarzschild and the Kerr metric.
A novel electrospray-based ion mobility spectrometer
Authors: Bathgate Ben , Cheong Eric C. S. , Backhouse Christopher J.
Published: July 2004

We present a design for a low-cost ion mobility spectrometer that can be built using the equipment on hand in many electronics-oriented undergraduate laboratories. The construction of this system is based upon the use of printed circuit boards and does not require the specialized drift and sheath gases, vacuum pumps, heater assemblies, high voltage pulsers, or precision pumps that are characteristic of the systems generally reported in the literature. We demonstrate the system in the separation of ions of methanol and water in air. Despite the low cost of this system it has a performance comparable to more complex systems, with a sensitivity of approximately 100 ppm for the protein cytochrome c. This system is suitable for use as an electronics or signal-processing project, or even a biotechnology demonstration.
Realization of an optical profiler: Introduction to scanning probe microscopy
Authors: Friedt J.-M.
Published: July 2004

Scanning probe microscopy is a widely used instrumentation method based on scanning a probe over a sample to map the spatial variation of a physical property over its surface. Here we present the steps required for the realization of a generic scanning probe microscope. The selected sensor is a CD reader optical head that monitors the distance to the focusing point of a lens by which an infrared laser is focused on the reflecting surface of the sample. We thus develop an optical profiler and illustrate practical examples by mapping the topography of a coin. The focus of this article is to describe a low cost profiler illustrating all the steps of the development, from the definition of the probe of the physical property to be monitored to the result on an actual sample.
Quality factors and conductances in Helmholtz resonators
Authors: Moloney Michael J.
Published: July 2004

Three glass bottles were investigated as Helmholtz resonators. The acoustical conductance C of each neck region was determined experimentally and compared to the calculated values, even though no conductance could be calculated for the flaring region of each bottle. The quality factor Q also was measured for 15 frequencies. The theoretical Q values were computed using the experimental C values and were found to be slightly higher than the measured Q values, probably due to the inability to calculate small wall losses in the flaring region beyond the neck.
Fermi and Bose pressures in statistical mechanics
Authors: Durand Loyal
Published: July 2004

I show how the pressure in Fermi and Bose systems, identified in standard discussions of quantum statistical mechanics by the use of thermodynamic analogies, can be derived directly in terms of the flux of momentum across a surface by using the quantum mechanical stress tensor. In this approach, which is analogous to classical kinetic theory, the pressure is naturally defined locally. The approach leads to a simple interpretation of the pressure in terms of the momentum flow encoded in the wave functions. The stress-tensor and thermodynamic approaches are related by an interesting application of boundary perturbation theory for quantum systems. I investigate the properties of quasi-continuous systems, the relations for Fermi and Bose pressures, shape-dependent effects and anisotropies, and the treatment of particles in external fields, and note several interesting problems for graduate courses in statistical mechanics.
The distribution of composite measurements: How to be certain of the uncertainties in what we measure
Authors: Silverman M. P. , Strange W. , Lipscombe T. C.
Published: July 2004

We derive expressions for the exact probability distribution functions and statistical moments of measurements represented as products and quotients of independent random variables, test these relations by means of the α and β branching decays of Bi212, and discuss the implications of our theory for the measurement of lipid-panel analytes in the assessment of the risk of coronary heart disease.
An experiment in nonlinear beam theory
Authors: Valiente A.
Published: July 2004

An undergraduate level experiment involving the nonlinear mechanics of deformable bodies is described. The phenomenon of interest is the large elastic sideways deflection of a column under compressive loading. The experimental arrangement and the measurement’s simple equipment are usually found in undergraduate laboratories. The analysis is more involved because the use of nonelementary functions is required. To circumvent this difficulty, the analysis can be done with a spreadsheet or a data analysis program that has been preprogrammed to compute the required integrals.
Generalized Fierz identities
Authors: Nieves José F. , Pal Palash B.
Published: July 2004

Low energy weak interaction calculations with fermions frequently involve a superposition of quartic products of Dirac spinors, in which the order of the spinors is not the same in all the contributing terms. A common trick that is used to bring them to a uniform ordering is the Fierz transformation. We show that the standard Fierz transformation quoted in textbooks is one element of a class of transformations that exist between fermion amplitudes that are written with different, arbitrary reorderings, of the spinors. The explicit form of all such transformation matrices is given, and in addition to the usual Lorentz scalar quartic products, we consider the pseudoscalar ones. Moreover, we include the cases in which some, or all, of the spinors in the original amplitude appear in the transformed amplitudes in their Lorentz-invariant complex conjugate form. Such manipulations and formulas are useful in practical calculations, as we show with some examples.
Quantum interference in a driven washboard potential
Authors: Thommen Quentin , Garreau Jean Claude , Zehnlé Véronique
Published: July 2004

The dynamics of a quantum particle in a one-dimensional, tilted (“washboard”) and time-dependent lattice is studied. The approach uses quantum mechanics at the undergraduate level and leads to analytical results that show a rich variety of dynamical behavior and illustrate the fundamental role of interference in quantum systems.
Solutions of relativistic Newton’s equations for nonconstant fields
Authors: Wooten R. E. , Macek J. H.
Published: July 2004

Newton’s second law can be readily solved for many forces, but few situations can be solved for the relativistic form of Newton’s second law. The only problems directly solvable are those involving charged particles in constant electromagnetic fields. If the external field represents a light pulse, Dirac’s relativistic equation can be solved, as done by Volkov in 1935. Classical solutions based on Volkov’s work employ the Hamilton–Jacobi equations. We discuss the solution of this problem using Newton’s equations, thereby making the solution more accessible.
Resource Letter FNP-1: Frontiers of nuclear physics
Authors: Bertsch G. F.
Published: July 2004

This Resource Letter provides a bibliography of the current research activities in nuclear physics and also a guide for finding useful nuclear data. The major areas included are nuclear structure and reactions, symmetry tests, nuclear astrophysics, nuclear theory, high-density matter, and nuclear instrumentation.
A computational approach to teaching conservative chaos
Authors: Timberlake Todd
Published: July 2004

A computational approach to teaching conservative chaos that is suitable for undergraduates in an upper-level classical mechanics course is discussed. Some ways that computation can be used to facilitate the teaching of several important topics in conservative chaos are described.
Concept substitution: A teaching strategy for helping students disentangle related physics concepts
Authors: Grayson Diane J.
Published: July 2004

To be effective, physics teachers need both content knowledge and pedagogical content knowledge, which includes knowledge of student conceptions and effective teaching strategies. Although much information is available on student conceptual and reasoning difficulties in physics, much less information is available on how to remedy such difficulties. In this paper I describe a teaching strategy, concept substitution, which is useful when student difficulties arise from a failure to distinguish distinct but related physics concepts. By using the topic of electric circuits as the context, I show how this strategy enables the teacher to identify and build on students’ correct intuition, while enabling students to distinguish among related concepts. I also illustrate the complexity of the conceptual change process, including the presence of intermediate conceptions while the process is taking place.
Adaptive Mutations Producing Efficient Replication of Genotype 1a Hepatitis C Virus RNA in Normal Huh7 Cells
Authors: Yi MinKyung , Lemon Stanley M.
Published: July 2004

Despite recent successes in generating subgenomic RNA replicons derived from genotype 1b strains of hepatitis C virus (HCV) that replicate efficiently in cultured cells, it has proven difficult to generate efficiently replicating RNAs from any other genotype of HCV. This includes genotype 1a, even though it is closely related to genotype 1b. We show here that an important restriction to replication of the genotype 1a H77c strain RNA in normal Huh7 cells resides within the amino-terminal 75 residues of the NS3 protease. We identified adaptive mutations located within this NS3 domain and within NS4A, in close proximity to the essential protease cofactor sequence, that act cooperative to substantially enhance the replication of this genotype 1a RNA in Huh7 cells. These and additional adaptive mutations, identified through a series of iterative transfections and the selection of G418-resistant cell clones, form two groups associating with distinct nonstructural protein domains: the NS3/4A protease and NS5A. A combination of mutations from both groups led to robust replication of otherwise unmodified H77c genomic RNA that was readily detectable by northern analysis within 4 days of transfection into Huh7 cells. We speculate that these adaptive mutations favorably influence assembly of the replicase complex with host cell-specific proteins, or alternatively promote interactions of NS3/4A and/or NS5A with cellular proteins involved in host cell antiviral defenses.
Superinfection Exclusion in Duck Hepatitis B Virus Infection Is Mediated by the Large Surface Antigen
Authors: Walters Kathie-Anne , Joyce Michael A. , Addison William R. , Fischer Karl P. , Tyrrell D. Lorne J.
Published: July 2004

Superinfection exclusion is the phenomenon whereby a virus prevents the subsequent infection of an already infected host cell. The Pekin duck hepatitis B virus (DHBV) model was used to investigate superinfection exclusion in hepadnavirus infections. Superinfection exclusion was shown to occur both in vivo and in vitro with a genetically marked DHBV, DHBV-ClaI, which was unable to establish an infection in either DHBV-infected ducklings or DHBV-infected primary duck hepatocytes (PDHs). In addition, exclusion occurred in vivo even when the second virus had a replicative advantage. Superinfection exclusion appears to be restricted to DHBV, as adenovirus, herpes simplex virus type 1, and vesicular stomatitis virus were all capable of efficiently infecting DHBV-infected PDHs. Exclusion was dependent on gene expression by the original infecting virus, since UV-irradiated DHBV was unable to mediate the exclusion of DHBV-ClaI. Using recombinant adenoviruses expressing DHBV proteins, we determined that the large surface antigen mediated exclusion. The large surface antigen is known to cause down-regulation of a DHBV receptor, carboxypeptidase D (CPD). Receptor down-regulation is a mechanism of superinfection exclusion seen in other viral infections, and so it was investigated as a possible mechanism of DHBV-mediated exclusion. However, a mutant large surface antigen which did not down-regulate CPD was still capable of inhibiting DHBV infection of PDHs. In addition, exclusion of DHBV-ClaI did not correlate with a decrease in CPD levels. Finally, virus binding assays and confocal microscopy analysis of infected PDHs indicated that the block in infection occurs after internalization of the second virus. We suggest that superinfection exclusion may result from the role of the L surface antigen as a regulator of intracellular trafficking.
Assembly of
Turnip Yellow Mosaic Virus
Replication Complexes: Interaction between the Proteinase and Polymerase Domains of the Replication Proteins
Authors: Jakubiec Anna , Notaise Julien , Tournier Vincent , Héricourt François , Block Maryse A. , Drugeon Gabrièle , van Aelst Linda , Jupin Isabelle
Published: July 2004

Turnip yellow mosaic virus
(TYMV), a positive-strand RNA virus in the alphavirus-like supergroup, encodes two nonstructural replication proteins (140K and 66K), both of which are required for its RNA genome replication. The 140K protein contains domains indicative of methyltransferase, proteinase, and NTPase/helicase activities, while the 66K protein encompasses the RNA-dependent RNA polymerase domain. Recruitment of the 66K protein to the sites of viral replication, located at the periphery of chloroplasts, is dependent upon the expression of the 140K protein. Using antibodies raised against the 140K and 66K proteins and confocal microscopy, we report the colocalization of the TYMV replication proteins at the periphery of chloroplasts in transfected or infected cells. The replication proteins cofractionated in functional replication complexes or with purified chloroplast envelope membranes prepared from infected plants. Using a two-hybrid system and coimmunoprecipitation experiments, we also provide evidence for a physical interaction of the TYMV replication proteins. In contrast to what has been found for other members of the alphavirus-like supergroup, the interaction domains were mapped to the proteinase domain of the 140K protein and to a large region encompassing the core polymerase domain within the 66K protein. Coexpression and colocalization experiments confirmed that the helicase domain of the 140K protein is unnecessary for the proper recruitment of the 66K protein to the chloroplast envelope, while the proteinase domain appears to be essential for that process. These results support a novel model for the interaction of TYMV replication proteins and suggest that viruses in the alphavirus-like supergroup may have selected different pathways to assemble their replication complexes.
Targeting of Hepatitis C Virus Core Protein to Mitochondria through a Novel C-Terminal Localization Motif
Authors: Schwer Björn , Ren Shaotang , Pietschmann Thomas , Kartenbeck Jürgen , Kaehlcke Katrin , Bartenschlager Ralf , Yen T. S. Benedict , Ott Melanie
Published: July 2004

The hepatitis C virus (HCV) core protein represents the first 191 amino acids of the viral precursor polyprotein and is cotranslationally inserted into the membrane of the endoplasmic reticulum (ER). Processing at position 179 by a recently identified intramembrane signal peptide peptidase leads to the generation and potential cytosolic release of a 179-amino-acid matured form of the core protein. Using confocal microscopy, we observed that a fraction of the mature core protein colocalized with mitochondrial markers in core-expressing HeLa cells and in Huh-7 cells containing the full-length HCV replicon. Subcellular fractionation confirmed this observation and showed that the core protein associates with purified mitochondrial fractions devoid of ER contaminants. The core protein also fractionated with mitochondrion-associated membranes, a site of physical contact between the ER and mitochondria. Using immunoelectron microscopy and in vitro mitochondrial import assays, we showed that the core protein is located on the mitochondrial outer membrane. A stretch of 10 amino acids within the hydrophobic C terminus of the processed core protein conferred mitochondrial localization when it was fused to green fluorescent protein. The location of the core protein in the outer mitochondrial membrane suggests that it could modulate apoptosis or lipid transfer, both of which are associated with this subcellular compartment, during HCV infection.
Intracellular Processing of Human Herpesvirus 6 Glycoproteins Q1 and Q2 into Tetrameric Complexes Expressed on the Viral Envelope
Authors: Akkapaiboon Pilailuk , Mori Yasuko , Sadaoka Tomohiko , Yonemoto Sayoko , Yamanishi Koichi
Published: July 2004

Human herpesvirus 6 (HHV-6) glycoproteins H and L (gH and gL, respectively) and the 80-kDa form of glycoprotein Q (gQ-80K) form a heterotrimeric complex that is found on the viral envelope and that is a viral ligand for human CD46. Besides gQ-80K, the gQ gene encodes an additional product whose mature molecular mass is 37 kDa (gQ-37K) and which is derived from a different transcript. Therefore, we designated gQ-80K as gQ1 and gQ-37K as gQ2. We show here that gQ2 also interacts with the gH-gL-gQ1 complex in HHV-6-infected cells and in virions. To examine how these components interact in HHV-6-infected cells, we performed pulse-chase studies. The results demonstrated that gQ2-34K, which is endo-β-
N
-acetylglucosaminidase H sensitive and which is the precursor form of gQ2-37K, associates with gQ1-74K, which is the precursor form of gQ1-80K, within 30 min of the pulse period. After a 1-h chase, these precursor forms had associated with the gH-gL dimer. Interestingly, an anti-gH monoclonal antibody coimmunoprecipitated mainly gQ1-80K and gQ2-37K, with little gQ1-74K or gQ2-34K. These results indicate that although gQ2-34K and gQ1-74K interact in the endoplasmic reticulum, the gH-gL-gQ1-80K-gQ2-37K heterotetrameric complex arises in the post-endoplasmic reticulum compartment. The mature complex is subsequently incorporated into viral particles.
Functional Antagonism of Chemokine Receptor CCR1 Reduces Mortality in Acute Pneumovirus Infection In Vivo
Authors: Bonville Cynthia A. , Lau Vincent K. , DeLeon Jordana M. , Gao Ji-Liang , Easton Andrew J. , Rosenberg Helene F. , Domachowske Joseph B.
Published: July 2004

We present an antiviral-immunomodulatory therapeutic strategy involving the chemokine receptor antagonist Met-RANTES, which yields significant survival in the setting of an otherwise fatal respiratory virus infection. In previous work, we demonstrated that infection with the natural rodent pathogen pneumonia virus of mice involves robust virus replication accompanied by cellular inflammation modulated by the CC chemokine macrophage inflammatory protein 1α (MIP-1α). We found that the antiviral agent ribavirin limited virus replication in vivo but had no impact on morbidity and mortality associated with this disease in the absence of immunomodulatory control. We show here that ribavirin reduces mortality, from 100% to 10 and 30%, respectively, in gene-deleted CCR1
−/−
mice and in wild-type mice treated with the small-molecule chemokine receptor antagonist, Met-RANTES. As MIP-1α-mediated inflammation is a common response to several distantly related respiratory virus pathogens, specific antiviral therapy in conjunction with blockade of the MIP-1α/CCR1 inflammatory cascade may ultimately prove to be a useful, generalized approach to severe respiratory virus infection and its pathological sequelae in human subjects.
Transport of African Swine Fever Virus from Assembly Sites to the Plasma Membrane Is Dependent on Microtubules and Conventional Kinesin
Authors: Jouvenet Nolwenn , Monaghan Paul , Way Michael , Wileman Thomas
Published: July 2004

African swine fever virus (ASFV) is a large DNA virus that assembles in perinuclear viral factories located close to the microtubule organizing center. In this study, we have investigated the mechanism by which ASFV reaches the cell surface from the site of assembly. Immunofluorescence microscopy revealed that at 16 h postinfection, mature virions were aligned along microtubules. Furthermore, virus movement to the cell periphery was inhibited when microtubules were depolymerized by nocodazole. In addition, ASFV infection resulted in the increased acetylation of microtubules as well as their protection against depolymerization by nocodazole. Immunofluorescence microscopy showed that conventional kinesin was recruited to virus factories and to a large fraction of virus particles in the cytoplasm. Consistent with a role for conventional kinesin during ASFV egress to the cell periphery, overexpression of the cargo-binding domain of the kinesin light chain severely inhibited the movement of particles to the plasma membrane. Based on our observations, we propose that ASFV is recognized as cargo by conventional kinesin and uses this plus-end microtubule motor to move from perinuclear assembly sites to the plasma membrane.
Compartmentalization of VP16 in Cells Infected with Recombinant Herpes Simplex Virus Expressing VP16-Green Fluorescent Protein Fusion Proteins
Authors: La Boissière Sylvie , Izeta Ander , Malcomber Sophie , O'Hare Peter
Published: July 2004

VP16 is an essential structural protein of herpes simplex virus. It plays important roles in immediate-early transcriptional regulation, in the modulation of the activities of other viral components, and in the pathway of assembly and egress of infectious virions. To gain further insight into the compartmentalization of this multifunctional protein we constructed and characterized recombinant viruses expressing VP16 linked to the green fluorescent protein (GFP). These viruses replicate with virtually normal kinetics and yields and incorporate the fusion protein into the virion, resulting in autofluorescent particles. De novo-synthesized VP16-GFP was first detected in a diffuse pattern within the nucleus. Nuclear VP16-GFP was progressively recruited to replication compartments, which coalesced into large globular domains. By 10 to 12 h after infection additional distinct foci containing VP16-GFP could be seen, almost exclusively located at the periphery of the replication compartments. At the same time pronounced accumulation was observed in the cytoplasm, first in a diffuse pattern and then accumulating in vesicle-like compartments which were concentrated in an asymmetric fashion reminiscent of the Golgi. Inhibition of DNA replication resulted in prolonged diffuse nuclear distribution with minimal cytoplasmic accumulation. Treatment with brefeldin disrupted the cytoplasm vesicular pattern, resulting in redistributed large foci. Time-lapse microscopy demonstrated various dynamic features of infection, including the active induction of very long cellular projections (up to 100 μM). Vesicular clusters containing VP16 were transported within projections to the termini, which developed bulbous ends and appeared to embed into the membranes of adjacent uninfected cells.
Coexpression of UL20p and gK Inhibits Cell-Cell Fusion Mediated by Herpes Simplex Virus Glycoproteins gD, gH-gL, and Wild-Type gB or an Endocytosis-Defective gB Mutant and Downmodulates Their Cell Surface Expression
Authors: Avitabile Elisa , Lombardi Giulia , Gianni Tatiana , Capri Miriam , Campadelli-Fiume Gabriella
Published: July 2004

Syncytium formation in cells that express herpes simplex virus glycoprotein B (gB), gD, gH, and gL is blocked by gK (E. Avitabile, G. Lombardi, and G. Campadelli-Fiume, J. Virol.
77:
6836-6844, 2003). Here, we report the results of two series of experiments. First, UL20 protein (UL20p) expression weakly inhibited cell-cell fusion. Coexpression of UL20p and gK drastically reduced fusion in a cell-line-dependent manner, with the highest inhibition in BHK cells. Singly expressed UL20p and gK localized at the endoplasmic reticulum and nuclear membranes. When they were coexpressed, both proteins relocalized to the Golgi apparatus. Remarkably, in cells that coexpressed UL20p and gK, the antifusion activity correlated with a downmodulation of gD, gB, gH, and gL cell surface expression. Second, gB
Δ867
has a partial deletion in the cytoplasmic tail that removed endocytosis motifs. Whereas wild-type (wt) gB was internalized in vesicles lined with the endosomal marker Rab5, gB
Δ867
was not internalized, exhibited enhanced cell surface expression, and was more efficient in mediating cell-cell fusion than wt gB. The antifusion activity of UL20p and gK was also exerted when gB
Δ867
replaced wt gB in the cell fusion assay. These studies show that the gB C tail carries a functional endocytosis motif(s) and that the removal of the motif correlated with increased gB surface expression and increased fusion activity. We conclude that cell-cell fusion in wt-virus-infected cells is negatively controlled by at least two mechanisms. The novel mechanism described here involves the concerted action of UL20p and gK and correlates with a moderate but consistent reduction in the cell surface expression of the fusion glycoproteins. This mechanism is independent of the one exerted through endocytosis-mediated downmodulation of gB from the plasma membrane.
Degenerate In Vitro Genetic Selection Reveals Mutations That Diminish Alfalfa Mosaic Virus RNA Replication without Affecting Coat Protein Binding
Authors: Rocheleau Gail , Petrillo Jessica , Guogas Laura , Gehrke Lee
Published: July 2004

The alfalfa mosaic virus (AMV) RNAs are infectious only in the presence of the viral coat protein; however, the mechanisms describing coat protein's role during replication are disputed. We reasoned that mechanistic details might be revealed by identifying RNA mutations in the 3′-terminal coat protein binding domain that increased or decreased RNA replication without affecting coat protein binding. Degenerate (doped) in vitro genetic selection, based on a pool of randomized 39-mers, was used to select 30 variant RNAs that bound coat protein with high affinity. AUGC sequences that are conserved among AMV and ilarvirus RNAs were among the invariant nucleotides in the selected RNAs. Five representative clones were analyzed in functional assays, revealing diminished viral RNA expression resulting from apparent defects in replication and/or translation. These data identify a set of mutations, including G-U wobble pairs and nucleotide mismatches in the 5′ hairpin, which affect viral RNA functions without significant impact on coat protein binding. Because the mutations associated with diminished function were scattered over the 3′-terminal nucleotides, we considered the possibility that RNA conformational changes rather than disruption of a precise motif might limit activity. Native polyacrylamide gel electrophoresis experiments showed that the 3′ RNA conformation was indeed altered by nucleotide substitutions. One interpretation of the data is that coat protein binding to the AUGC sequences determines the orientation of the 3′ hairpins relative to one another, while local structural features within these hairpins are also critical determinants of functional activity.
Cytokine-Mediated Downregulation of Coxsackievirus-Adenovirus Receptor in Endothelial Cells
Authors: Vincent Theresa , Pettersson Ralf F. , Crystal Ronald G. , Leopold Philip L.
Published: July 2004

Endothelial cells have the ability to change their complement of cell surface proteins in response to inflammatory cytokines. We hypothesized that the expression of the coxsackievirus-adenovirus receptor (CAR), a viral receptor and putative cell-cell adhesion molecule, may be altered during the response of endothelial cells to inflammation. To test this hypothesis, we evaluated CAR protein and mRNA levels in human umbilical vein endothelial cells after they were exposed to tumor necrosis factor alpha, gamma interferon, or a combination of the two cytokines. Flow cytometric and Western blot analyses indicated that cytokine treatment led to a synergistic decrease in CAR protein expression. A Western blot analysis showed that CAR levels decreased to 16% ± 4% or 1% ± 4% of the CAR protein levels in untreated cells with either 24 or 48 h of cytokine treatment, respectively. Quantitative reverse transcription-PCR demonstrated that the combination treatment caused CAR mRNA levels to decrease to 21% ± 12% or 5% ± 3% of the levels in untreated cells after a 24- or 48-h cytokine treatment, respectively. Reduced CAR expression led to a decrease in adenovirus (Ad) binding of 80% ± 3% (compared with untreated endothelial cells), with a subsequent decrease in Ad-mediated gene transfer that was dependent on the dose and duration of cytokine treatment but not on the dose of Ad. A similar decrease in CAR protein level and susceptibility to Ad infection was observed in human microvascular endothelial cells, while CAR expression on normal human bronchial epithelial cells or A549 lung epithelial cells was less affected by cytokine treatments. Taken together, the data demonstrate that inflammatory cytokines decrease CAR mRNA and protein expression with a concomitant decrease in Ad binding, reflecting the impact of cell physiology on the function of CAR and the potential effect of inflammation on the ability of Ad to transfer genes to endothelial cells.