bluuebunny/crossref_metadata_2025 huggingface dataset:
Decomposition of metabolic network into functional modules based on the global connectivity structure of reaction graph
Authors: Ma Hong-Wu , Zhao Xue-Ming , Yuan Ying-Jin , Zeng An-Ping
Published: March 2004

Motivation: Metabolic networks are organized in a modular, hierarchical manner. Methods for a rational decomposition of the metabolic network into relatively independent functional subsets are essential to better understand the modularity and organization principle of a large-scale, genome-wide network. Network decomposition is also necessary for functional analysis of metabolism by pathway analysis methods that are often hampered by the problem of combinatorial explosion due to the complexity of metabolic network. Decomposition methods proposed in literature are mainly based on the connection degree of metabolites. To obtain a more reasonable decomposition, the global connectivity structure of metabolic networks should be taken into account.

Results: In this work, we use a reaction graph representation of a metabolic network for the identification of its global connectivity structure and for decomposition. A bow-tie connectivity structure similar to that previously discovered for metabolite graph is found also to exist in the reaction graph. Based on this bow-tie structure, a new decomposition method is proposed, which uses a distance definition derived from the path length between two reactions. An hierarchical classification tree is first constructed from the distance matrix among the reactions in the giant strong component of the bow-tie structure. These reactions are then grouped into different subsets based on the hierarchical tree. Reactions in the IN and OUT subsets of the bow-tie structure are subsequently placed in the corresponding subsets according to a ‘majority rule’. Compared with the decomposition methods proposed in literature, ours is based on combined properties of the global network structure and local reaction connectivity rather than, primarily, on the connection degree of metabolites. The method is applied to decompose the metabolic network of Escherichia coli. Eleven subsets are obtained. More detailed investigations of the subsets show that reactions in the same subset are really functionally related. The rational decomposition of metabolic networks, and subsequent studies of the subsets, make it more amenable to understand the inherent organization and functionality of metabolic networks at the modular level.

Supplementary Information:  http://genome.gbf.de/bioinformatics/
Normalization of single-channel DNA array data by principal component analysis
Authors: Stoyanova Radka , Querec Troy D. , Brown Truman R. , Patriotis Christos
Published: March 2004

Motivation: Detailed comparison and analysis of the output of DNA gene expression arrays from multiple samples require global normalization of the measured individual gene intensities from the different hybridizations. This is needed for accounting for variations in array preparation and sample hybridization conditions.

Results: Here, we present a simple, robust and accurate procedure for the global normalization of datasets generated with single-channel DNA arrays based on principal component analysis. The procedure makes minimal assumptions about the data and performs well in cases where other standard procedures produced biased estimates. It is also insensitive to data transformation, filtering (thresholding) and pre-screening.
MITOPRED: a genome-scale method for prediction of nucleus-encoded mitochondrial proteins
Authors: Guda Chittibabu , Fahy Eoin , Subramaniam Shankar
Published: March 2004

Motivation: Currently available methods for the prediction of subcellular location of mitochondrial proteins rely largely on the presence of mitochondrial targeting signals in the protein sequences. However, a large fraction of mitochondrial proteins lack such signals, making those tools ineffective for genome-scale prediction of mitochondria-targeted proteins. Here, we propose a method for genome-scale prediction of nucleus-encoded mitochondrial proteins. The new method, MITOPRED, is based on the Pfam domain occurrence patterns and the amino acid compositional differences between mitochondrial and non-mitochondrial proteins.

Results: MITOPRED could predict mitochondrial proteins with 100% specificity at a 44% sensitivity rate and with 67% specificity at 99% sensitivity. Additionally, it was sufficiently robust to predict mitochondrial proteins across different eukaryotic species with similar accuracy. Based on Matthews correlation coefficient measure, the prediction performance of MITOPRED is clearly superior (0.73) to those of the two popular methods TargetP (0.51) and PSORT (0.53). Using this method, we predicted the nucleus-encoded mitochondrial proteins from six complete genomes (three invertebrate, two vertebrate and one plant species) and estimated the total number in each genome. In human, our method estimated the existence of 1362 mitochondrial proteins corresponding to 4.8% of the total proteome.

Availability: MITOPRED program is freely accessible at http://mitopred.sdsc.edu. Source code is available on request from the authors.

Supplementary information: Training data sets are also available at http://mitopred.sdsc.edu
BLAST2GENE: a comprehensive conversion of BLAST output into independent genes and gene fragments
Authors: Suyama Mikita , Torrents David , Bork Peer
Published: March 2004

Summary: BLAST2GENE is a program that allows a detailed analysis of genomic regions containing completely or partially duplicated genes. From a BLAST (or BL2SEQ) comparison of a protein or nucleotide query sequence with any genomic region of interest, BLAST2GENE processes all high scoring pairwise alignments (HSPs) and provides the disposition of all independent copies along the genomic fragment. The results are provided in text and PostScript formats to allow an automatic and visual evaluation of the respective region.

Availability: The program is available upon request from the authors. A web server of BLAST2GENE is maintained at http://www.bork.embl.de/blast2gene
Time-dependent Inhibition of G Protein–coupled Receptor Signaling by Local Anesthetics
Authors: Hollmann Markus W. , Herroeder Susanne , Kurz Katrin S. , Hoenemann Christian W. , Struemper Danja , Hahnenkamp Klaus , Durieux Marcel E.
Published: March 2004

Background

Several beneficial effects of local anesthetics (LAs) were shown to be due to inhibition of G protein-coupled receptor signaling. Differences in exposure time might explain discrepancies in concentrations of LAs required to achieve these protective effects in vivo and in vitro (approximately 100-fold higher). Using Xenopus oocytes and human neutrophils, the authors studied time-dependent effects of LAs on G protein-coupled receptor signaling and characterized possible mechanisms and sites of action.

Methods

Measurement of agonist-induced Ca2+-activated Cl currents, using a two-electrode voltage clamp technique, and determination of superoxide anion production by cytochrome c assay were used to assess the effects of LAs on G protein-coupled receptor signaling in oocytes and primed and activated human neutrophils, respectively. Antisense knockdown of G alpha q protein and inhibition of various proteins within the signaling pathway served for defining mechanisms and sites of action more specifically.

Results

LAs attenuated G protein-coupled receptor signaling in both models in a time-dependent and reversible manner (lidocaine reduced lysophosphatidic acid signaling to 19 +/- 3% after 48 h and 25 +/- 2% after 6 h of control response in oocytes and human neutrophils, respectively). Whereas no effect was observed after extracellularly applied or intracellularly injected QX314, a lidocaine analog, using G alpha q-depleted oocytes, time-dependent inhibition also occurred after intracellular injection of QX314 into undepleted oocytes. Inhibition of phosphatases or protein kinases and agonist-independent G-protein stimulation, using guanosine 5'-O-3-thiotriphosphate or aluminum fluoride, did not affect time-dependent inhibition by LAs.

Conclusion

Inhibition of G protein-coupled receptor signaling by LAs was found to be time dependent and reversible. Critically requiring G alpha q-protein function, this effect is located downstream of guanosine diphosphate-guanosine triphosphate exchange and is not dependent on increased guanosine triphosphatase activity, phosphatases, or protein kinases.
Sevoflurane Inhibits Angiotensin II–induced, Protein Kinase C–mediated but Not Ca2+-elicited Contraction of Rat Aortic Smooth Muscle
Authors: Yu Jingui , Tokinaga Yasuyuki , Ogawa Koji , Iwahashi Shizue , Hatano Yoshio
Published: March 2004

Background

Whether volatile anesthetics attenuate angiotensin II-mediated vascular tone has not been determined. The current study was designed to investigate the effects of sevoflurane on the angiotensin II-stimulated, Ca2+- and protein kinase C (PKC)-mediated contraction of rat aortic smooth muscle.

Methods

The dose-dependent effects of sevoflurane on angiotensin II (10 m)-induced contraction, the increase in intracellular Ca2+ concentration, and PKC phosphorylation of rat aortic smooth muscle were measured using an isometric force transducer, a fluorometer, and Western blotting, respectively.

Results

Angiotensin II induced a transient increase in intracellular Ca2+ concentration, phosphorylation of Ca2+-dependent PKC (cPKC)-alpha, and consequently, a transient contraction of rat aortic smooth muscle. Phosphorylation of the Ca2+-independent PKC-epsilon was not detected. The angiotensin II-induced contraction was almost completely abolished by removing extracellular Ca2+ and was significantly inhibited by the selective cPKC inhibitor Gö 6976 (10 M) but was not inhibited by the nonselective PKC inhibitor Ro 31-8425 (10 M). Sevoflurane dose-dependently inhibited the angiotensin II-induced contraction, with reductions of 14.2 +/- 5.2% (P > 0.05), 26.7 +/- 8.9% (P < 0.05), and 38.5 +/- 12.8% (P < 0.01) (n = 10) in response to 1.7, 3.4, and 5.1% sevoflurane, respectively. The angiotensin II-elicited increase in intracellular Ca2+ concentration was not significantly influenced by 3.4, 5.1, or 8.5% sevoflurane. However, cPKC-alpha phosphorylation induced by angiotensin II was inhibited dose dependently by 1.7, 3.4, and 5.1% sevoflurane, with depressions of 20.5 +/- 14.2% (P > 0.05), 37.0 +/- 17.8% (P < 0.05), and 62.5 +/- 12.2% (P < 0.01) (n = 4), respectively.

Conclusion

The current study indicates that Ca2+ and cPKC-alpha are involved in angiotensin II-induced vascular contraction. Sevoflurane dose-dependently inhibited the angiotensin II-stimulated, cPKC-mediated but not Ca2+-elicited contraction of rat aortic smooth muscle.
Antiallodynic Effects of Systemic and Intrathecal Morphine in the Spared Nerve Injury Model of Neuropathic Pain in Rats
Authors: Zhao Chengshui , Tall Jill M. , Meyer Richard A. , Raja Srinivasa N.
Published: March 2004

Background

The efficacy of opioids for neuropathic pain remains controversial. The effects of morphine on pain behavior were investigated in two animal models of neuropathic pain: the spared nerve injury (SNI) model and the spinal nerve ligation (SNL) model.

Methods

Nerve injuries were created in rats either by tight ligation and section of the left tibial and common peroneal nerves (SNI) or by unilateral ligation of L5 and L6 spinal nerves (SNL). Paw withdrawal threshold to mechanical stimuli was measured using the up-down method in the hairy and glabrous skin territories of the sural nerve for SNI rats or in the mid-plantar paw of SNL rats.

Results

Before SNI, the median paw withdrawal thresholds in hairy and glabrous skin were similar (26 g [25%, 75% quartiles: 26, 26 g]). The paw withdrawal threshold decreased after SNI in both hairy and glabrous skin (P < 0.001). Thirty days after the SNI, the threshold in hairy skin (0.3 g) was significantly lower than in glabrous skin (1.9 g; P < 0.001). In blinded experiments, both subcutaneous and intrathecal morphine (0.1-10 microg) dose-dependently attenuated mechanical allodynia induced by SNI measured in the hairy skin, an effect that was naloxone reversible. The ED50 for the intrathecal morphine was 0.52 microg (95% confidence interval, 0.31-0.90 microg). Morphine (1 microg intrathecal) attenuated SNI-induced mechanical allodynia in glabrous skin with potency similar to that in hairy skin. In SNL rats, morphine (30 microg intrathecal) almost completely reversed the SNL-induced mechanical allodynia.

Conclusions

(1) SNI-induced mechanical allodynia is characterized by a lower paw withdrawal threshold in hairy versus glabrous skin; (2) systemic and intrathecal morphine reverse SNI-induced mechanical allodynia in a dose-dependent fashion; and (3) intrathecal morphine also reverses SNL-induced mechanical allodynia. These results suggest that intrathecal opioids are likely to be effective in the treatment of neuropathic pain.
Dissociable Brain Activation Responses to 5-Hz Electrical Pain Stimulation
Authors: Alkire Michael T. , White Nathan S. , Hsieh Raymond , Haier Richard J.
Published: March 2004

Background

To elucidate neural correlates associated with processing of tonic aching pain, the authors used high-field (3-T) functional magnetic resonance imaging with a blocked parametric study design and characterized regional brain responses to electrical stimulation according to stimulus intensity-response functions.

Methods

Pain was induced in six male volunteers using a 5-Hz electrical stimulus applied to the right index finger. Scanning sequences involved different levels of stimulation corresponding to tingling sensation (P1), mild pain (P2), or high pain (P3). Common effects across subjects were sought using a conjunction analyses approach, as implemented in statistical parametric mapping (SPM-99).

Results

The contralateral posterior/mid insula and contralateral primary somatosensory cortex were most associated with encoding stimulus intensity because they showed a positive linear relation between blood oxygenation level-dependent signal responses and increasing stimulation intensity (P1 < P2 < P3). The contralateral secondary somatosensory cortex demonstrated a response function most consistent with a role in pain intensity encoding because it had no significant response during the innocuous condition (P1) but proportionally increased activity with increasingly painful stimulus intensities (0 < P2 < P3). Finally, a portion of the anterior cingulate cortex (area 24) and supplementary motor area 6 demonstrated a high pain-specific response (P3).

Conclusions

The use of response function modeling, conjunction analysis, and high-field imaging reveals dissociable regional responses to a tonic aching electrical pain. Most specifically, the primary somatosensory cortex and insula seem to encode stimulus intensity information, whereas the secondary somatosensory cortex encodes pain intensity information. The cingulate findings are consistent with its proposed role in processing affective-motivational aspects of pain.
Identification of a virus trans-acting regulatory element on the latent DNA replication of Kaposi's sarcoma-associated herpesvirus
Authors: Lim Chunghun , Seo Taegun , Jung Jun , Choe Joonho
Published: March 2004

Latency-associated nuclear antigen 1 (LANA1) of Kaposi's sarcoma-associated herpesvirus (KSHV) plays a pivotal role in the maintenance of the virus genome in latently infected cells. LANA1 links virus genomes to host chromosomes via a C-terminal DNA-binding domain which interacts with the sequences located in terminal repeats (TRs) of the virus genome and via an N-terminal chromosome-binding sequence which associates with the host chromosomes, respectively. Recent data suggest that LANA1 also actively participates in the replication of KSHV TR-containing plasmid in the transient DNA replication assay. In this report, it was found that C33A and COS-1, but not NIH/3T3, cell lines are permissive for the transient replication of KSHV TR-containing plasmid. Using several LANA1-deletion mutants, the minimum domain of LANA1 required for replication activity was also determined. In addition, the N terminus of LANA1 inhibited the transient replication systems of KSHV and Epstein–Barr virus (EBV) in transiently transfected 293 and 293T cells, but the C terminus of LANA1 specifically inhibited the transient replication system of KSHV in other cell lines. Consistent with previous reports, these data further emphasize the functional importance of the N terminus of LANA1 on replication from the KSHV latent origin of DNA replication.
Herpes simplex virus type 1 infection of polarized epithelial cells requires microtubules and access to receptors present at cell–cell contact sites
Authors: Marozin Sabrina , Prank Ute , Sodeik Beate
Published: March 2004

Mucosal epithelia are invaded from the apical surface during a primary infection by herpes simplex virus type 1 (HSV-1). HSV-1 progeny virus, synthesized from latently infected peripheral neurons that innervate such epithelia, reinfects the epithelia most likely from the basolateral surface. The epithelial cell lines MDCK and Caco-2 can be inducedin vitroto differentiate into polarized cells with distinct apical and plasma membrane domains separated by tight junctions if they are cultured on porous membrane filters. Our data using these culture systems showed that highly polarized epithelial cells were not susceptible to apical HSV-1 infection. However, HSV-1 infected these cells if added from the basolateral surface or if a depletion of extracellular Ca2+had weakened the strength of the cell–cell contacts. Basolateral infection and apical infection after the Ca2+switch required an intact microtubule network for genome targeting to the nucleus. This system can be used to identify the microtubule motors that HSV-1 uses during virus entry in polarized epithelial cells.
Sequence analysis of the guanylyltransferase (VP3) of group A rotaviruses
Authors: Cook Jonathan P. , McCrae Malcolm A.
Published: March 2004

The RNA segment encoding the guanylyltransferase (VP3) from 12 group A rotavirus isolates has been sequenced following RT-PCR and molecular cloning of the full-length amplicons produced. Alignment of the derived amino acid sequences including those of the four VP3 sequences available from GenBank revealed two levels of sequence divergence. Virus isolates from humans showed greater than 94 % sequence identity, whereas those isolated from different mammalian species showed as low as 79 % sequence identity. The exceptions were avian virus isolates, which diverged ∼45 % from those of mammalian origin, and the human virus isolates DS1 and 69M, which showed much closer (over 90 %) identity to viruses of bovine origin, suggesting that these human isolates may have undergone recent reassortment events with a bovine virus. Analysis of the sequences for a putative enzymic active site has revealed that the KXTAMDXEXP and KXXGNNH motifs around amino acids 385 and 545, respectively, are conserved across both group A and C rotaviruses.
Hepatitis D virus RNA editing is inhibited by a GFP fusion protein containing a C-terminally deleted delta antigen
Authors: Shih Ko-Nien , Chuang Ya-Ting , Liu Hsuan , Lo Szecheng J.
Published: March 2004

During its life cycle, hepatitis D virus (HDV) produces two forms of delta antigen (HDAg), small delta antigen (SDAg) and large delta antigen (LDAg), which differ in their C-terminal 19 amino acids. Host enzymes termed ADARs (adenosine deaminases that act on double-stranded RNA) are required for LDAg production. These enzymes change the stop codon (UAG) of SDAg to a tryptophan codon (UGG). However, the temporal and spatial regulation of HDV RNA editing is largely unknown. In this study, we constructed three GFP fusion proteins containing different lengths of SDAg and characterized their cellular localization and effects on HDV replication. One of these fusion proteins, designated D(1–88)-GFP, inhibited LDAg but not SDAg production, suggesting that D(1–88)-GFP inhibits HDV RNA editing. Two experiments further supported this supposition: (i) RT-PCR analysis combined with NcoI restriction enzyme digestion revealed that HDV RNA editing was reduced by 42 % in HeLa-D(1–88)-GFP when compared with HeLa cells; and (ii) the ratio of SDAg/LDAg production from the reporter RNAs was reduced in cells co-transfected with ADAR-expressing and reporter plasmids in the presence of D(1–88)-GFP. Double fluorescence microscopy found that D(1–88)-GFP was either associated with SC-35 or was adjacent to PML (premyelocytic leukaemia antigen) at nuclear speckles, but D(1–88)-GFP was not co-localized with ADAR, which was mainly located in the nucleolus. In situ hybridization showing co-localization of HDV RNA with D(1–88)-GFP at nuclear speckles suggested that HDV RNA editing might occur in the nuclear speckles and require other nuclear factor(s), in addition to ADAR.
Amino acid changes responsible for attenuation of virus neurovirulence in an infectious cDNA clone of the Oshima strain of Tick-borne encephalitis virus
Authors: Hayasaka Daisuke , Gritsun Tamara S. , Yoshii Kentarou , Ueki Tomotaka , Goto Akiko , Mizutani Tetsuya , Kariwa Hiroaki , Iwasaki Takuya , Gould Ernest A. , Takashima Ikuo
Published: March 2004

A stable full-length infectious cDNA clone of the Oshima strain of Tick-borne encephalitis virus (Far-Eastern subtype) was developed by a long high-fidelity RT-PCR and one-step cloning procedure. The infectious clone (O-IC) had four amino acid substitutions and produced smaller plaques when compared with the parent Oshima 5-10 strain. Using site-directed mutagenesis, the substitutions were reverted to restore the parent virus sequence (O-IC-pt). Although genetically identical, parent virus Oshima 5-10 and virus recovered from O-IC-pt demonstrated some biological differences that are possibly explained by the presence of quasispecies with differing virulence characteristics within the original virus population. These observations may have implications for vaccines based on modified infectious clones. It was also demonstrated that the amino acid substitution E-S40→P at position 40 in the envelope (E) glycoprotein was responsible for plaque size reduction, reduced infectious virus yields in cell culture and reduced mouse neurovirulence. Additionally, two amino acid substitutions in the non-structural (NS)5 protein (virus RNA-dependent RNA polymerase) NS5-V378→A and NS5-R674→K also contributed to attenuation of virulence in mice, but did not demonstrate a noticeable biological effect in baby hamster kidney cell culture. Comparative neurovirulence tests revealed how the accumulation of individual mutations (E-S40→P, NS5-V378→A and NS5-R674→K) can result in the attenuation of a virus.
Compatibility of the movement protein and the coat protein of cucumoviruses is required for cell-to-cell movement
Authors: Salánki Katalin , Gellért Ákos , Huppert Emese , Náray-Szabó Gábor , Balázs Ervin
Published: March 2004

For the cell-to-cell movement of cucumoviruses both the movement protein (MP) and the coat protein (CP) are required. These are not reversibly exchangeable betweenCucumber mosaic virus(CMV) andTomato aspermy virus(TAV). The MP of CMV is able to function with the TAV CP (chimera RT), but TAV MP is unable to promote the cell-to-cell movement in the presence of CMV CP (chimera TR). To gain further insight into the non-infectious nature of the TR recombinant, RNA 3 chimeras were constructed with recombinant MPs and CPs. The chimeric MP and one of the CP recombinants were infectious. The other recombinant CP enabled virus movement only after the introduction of two point mutations (Glu→Lys and Lys→Arg at aa 62 and 65, respectively). The mutations served to correct the CP surface electrostatic potential that was altered by the recombination. The infectivity of the TR virus on different test plants was restored by replacing the sequence encoding the C-terminal 29 aa of the MP with the corresponding sequence of the CMV MP gene or by exchanging the sequence encoding the C-terminal 15 aa of the CP with the same region of TAV. The analysis of the recombinant clones suggests a requirement for compatibility between the C-terminal 29 aa of the MP and the C-terminal two-thirds of the CP for cell-to-cell movement of cucumoviruses.
A human immunodeficiency virus 1 (HIV-1) clade A vaccine in clinical trials: stimulation of HIV-specific T-cell responses by DNA and recombinant modified vaccinia virus Ankara (MVA) vaccines in humans
Authors: Mwau Matilu , Cebere Inese , Sutton Julian , Chikoti Priscilla , Winstone Nicola , Wee Edmund G.-T. , Beattie Tara , Chen Yun-Hsiang , Dorrell Lucy , McShane Helen , Schmidt Claudia , Brooks Mary , Patel Sandip , Roberts Joanna , Conlon Christopher , Rowland-Jones Sarah L. , Bwayo Job J. , McMichael Andrew J. , Hanke Tomáš
Published: March 2004

The immunogenicities of candidate DNA- and modified vaccinia virus Ankara (MVA)-vectored human immunodeficiency virus (HIV) vaccines were evaluated on their own and in a prime–boost regimen in phase I clinical trials in healthy uninfected individuals in the United Kingdom. Given the current lack of approaches capable of inducing broad HIV-neutralizing antibodies, the pTHr.HIVA DNA and MVA.HIVA vaccines focus solely on the induction of cell-mediated immunity. The vaccines expressed a common immunogen, HIVA, which consists of consensus HIV-1 clade A Gag p24/p17 proteins fused to a string of clade A-derived epitopes recognized by cytotoxic T lymphocytes (CTLs). Volunteers' fresh peripheral blood mononuclear cells were tested for HIV-specific responses in a validated gamma interferon enzyme-linked immunospot (ELISPOT) assay using four overlapping peptide pools across the Gag domain and three pools of known CTL epitopes present in all of the HIVA protein. Both the DNA and the MVA vaccines alone and in a DNA prime–MVA boost combination were safe and induced HIV-specific responses in 14 out of 18, seven out of eight and eight out of nine volunteers, respectively. These results are very encouraging and justify further vaccine development.
Interaction of VPg-Pro of Turnip mosaic virus with the translation initiation factor 4E and the poly(A)-binding protein in planta
Authors: Léonard Simon , Viel Catherine , Beauchemin Chantal , Daigneault Nicole , Fortin Marc G. , Laliberté Jean-François
Published: March 2004

The viral protein linked to the genome (VPg) ofTurnip mosaic virus(TuMV) interactsin vitrowith the translation eukaryotic initiation factor (eIF) 4E. In the present study, we investigated the consequence of TuMV infection on eIF4E expression. Two isomers are present in plants, namely eIF4E and eIF(iso)4E. Expression of the latter was detected in both TuMV-infected and mock-inoculatedBrassica perviridisplants, but expression of eIF4E was found only in infected plants. Membranes from TuMV-infected or mock-inoculated tissues were separated by sucrose gradient centrifugation and fractions were collected. Immunoblot analyses showed that 6K2-VPg-Pro/VPg-Pro polyproteins were associated with endoplasmic reticulum membranes and were the viral forms likely to interact with eIF(iso)4E and eIF4E.In plantainteraction between 6K2-VPg-Pro/VPg-Pro and eIF(iso)4E/eIF4E was confirmed by co-purification by metal chelation chromatography. The poly(A)-binding protein (PABP) was also found to co-purify with VPg-Pro. Direct interaction between VPg-Pro and PABP was shown by an ELISA-based binding assay. These experiments suggest that a multi-protein complex may form around VPg-Pro of TuMV.
The influenza A virus NS1 protein binds small interfering RNAs and suppresses RNA silencing in plants
Authors: Bucher Etienne , Hemmes Hans , de Haan Peter , Goldbach Rob , Prins Marcel
Published: March 2004

RNA silencing comprises a set of sequence-specific RNA degradation pathways that occur in a wide range of eukaryotes, including animals, fungi and plants. A hallmark of RNA silencing is the presence of small interfering RNA molecules (siRNAs). The siRNAs are generated by cleavage of larger double-stranded RNAs (dsRNAs) and provide the sequence specificity for degradation of cognate RNA molecules. In plants, RNA silencing plays a key role in developmental processes and in control of virus replication. It has been shown that many plant viruses encode proteins, denoted RNA silencing suppressors, that interfere with this antiviral response. Although RNA silencing has been shown to occur in vertebrates, no relationship with inhibition of virus replication has been demonstrated to date. Here we show that the NS1 protein of human influenza A virus has an RNA silencing suppression activity in plants, similar to established RNA silencing suppressor proteins of plant viruses. In addition, NS1 was shown to be capable of binding siRNAs. The data presented here fit with a potential role for NS1 in counteracting innate antiviral responses in vertebrates by sequestering siRNAs.
Human influenza virus NS1 protein enhances viral pathogenicity and acts as an RNA silencing suppressor in plants
Authors: Delgadillo M. Otilia , Sáenz Pilar , Salvador Beatriz , García Juan Antonio , Simón-Mateo Carmen
Published: March 2004

RNA silencing has a well-established function as an antiviral defence mechanism in plants and insects. Using anAgrobacterium-mediated transient assay, we report here that NS1 protein from human influenza A virus suppresses RNA silencing in plants in a manner similar to P1/HC-Pro protein ofTobacco etch potyvirus, a well-characterized plant virus silencing suppressor. Moreover, we have shown that NS1 protein expression strongly enhances the symptoms ofPotato virus Xin three different plant hosts, suggesting that NS1 protein could be inhibiting defence mechanisms activated in the plant on infection. These data provide further evidence that an RNA silencing pathway could also be activated as a defence response in mammals.
Autographa californica M nucleopolyhedrovirus early GP64 synthesis mitigates developmental resistance in orally infected noctuid hosts
Authors: Zhang Ji-Hong , Washburn Jan O. , Jarvis Donald L. , Volkman Loy E.
Published: March 2004

The unusual early synthesis of the Autographa californica M nucleopolyhedrovirus (AcMNPV) budded virus (BV) structural protein GP64 is an important virulence factor during oral infection of Heliothis virescens larvae. Considering the breadth of the AcMNPV host range, the importance of early GP64 synthesis in orally infected permissive hosts (Trichoplusia ni and Spodoptera exigua) from subfamilies other than that of H. virescens was assessed. An AcMNPV recombinant, having wild-type early and late GP64 synthesis, was compared with one in which only late GP64 synthesis occurred. Early GP64 synthesis was found to have more of an effect on virulence in orally inoculated T. ni than S. exigua and that virulence was dependent on two factors: the ability of the host to slough occlusion-derived virus (ODV)-infected midgut cells and the rapidity with which BV was transmitted to the tracheal cells. In both host species, insects inoculated orally with the control virus transmitted BV to tracheal cells hours before those inoculated with the gp64 temporal mutant. Moreover, with early GP64 synthesis, the lag between the onset of viral gene expression in midgut and tracheal cells was only 3–4 h, supporting the conclusion that in these insects, the first systemic infections arose from ODV-derived nucleocapsids repackaged as BV. These results provide further empirical proof that early GP64 synthesis is a component of a unique and selectively advantageous baculovirus infection strategy for exploiting larval lepidopterans by counteracting developmental resistance.
Novel herpesviruses of Suidae: indicators for a second genogroup of artiodactyl gammaherpesviruses
Authors: Ehlers Bernhard , Lowden Stewart
Published: March 2004

Five novel herpesviruses were identified in suid species from Africa (common warthog,Phacochoerus africanus) and South-East Asia (bearded pig,Sus barbatus; babirusa,Babyrousa babyrussa) by detection and analysis of their DNA polymerase genes. Three of the novel species,P. africanuscytomegalovirus 1,P. africanuslymphotropic herpesvirus 1 (PafrLHV-1) andS. barbatuslymphotropic herpesvirus 1 (SbarLHV-1), were closely related to known beta- (porcine cytomegalovirus) and gammaherpesviruses [porcine lymphotropic herpesvirus (PLHV) 1 and 3] of domestic pigs. In contrast, two novel species,S. barbatusrhadinovirus 1 (SbarRHV-1) andBabyrousa babyrussarhadinovirus 1 (BbabRHV-1), were more closely related to a ruminant gammaherpesvirus, bovine herpesvirus 4 (BoHV-4), than to the porcine gammaherpesviruses PLHV-1, -2, -3, PafrLHV-1 and SbarLHV-1. SbarRHV-1, BbabRHV-1 and BoHV-4 were therefore tentatively assigned to a novel genogroup of artiodactyl gammaherpesviruses. This latter genogroup may also contain an as yet undiscovered gammaherpesvirus of domestic pigs, thereby adding a concern to their use in xenotransplantation.
Epidemiological and sequence differences between two subtypes (Ae and Aa) of hepatitis B virus genotype A
Authors: Sugauchi Fuminaka , Kumada Hiromitsu , Acharya Subrat A. , Shrestha Santosh Man , Gamutan Maria Teresita A. , Khan Mobin , Gish Robert G. , Tanaka Yasuhito , Kato Takanobu , Orito Etsuro , Ueda Ryuzo , Miyakawa Yuzo , Mizokami Masashi
Published: March 2004

Complete nucleotide sequences of 19 hepatitis B virus (HBV) isolates of genotype A (HBV/A) were determined and analysed along with those of 20 previously reported HBV/A isolates. Of the 19 HBV/A isolates, six including three from Japan and three from the USA clustered with the 14 HBV/A isolates from Western countries. The remaining 13 isolates including four from The Philippines, two from India, three from Nepal and four from Bangladesh clustered with the six HBV/A isolates reported from The Philippines, South Africa and Malawi. Due to distinct epidemiological distributions, genotype A in the 20 HBV isolates was classified into subtype Ae (e for Europe), and that in the other 19 into subtype Aa (a for Asia and Africa) provisionally. The 19 HBV/Aa isolates had a sequence variation significantly greater than that of the 20 HBV/Ae isolates (2·5±0·3 % vs 1·1±0·6 %, P<0·0001); they differed by 5·0±0·4 % (4·1–6·4 %). The double mutation (T1762/A1764) in the core promoter was significantly more frequent in HBV/Aa isolates than in HBV/Ae isolates (11/19 or 58 % vs 5/20 or 25 %, P<0·01). In the pregenome encapsidation (ε) signal, a point mutation from G to A or T at nt 1862 was detected in 16 of the 19 (84 %) HBV/Aa isolates but not in any of the 20 HBV/Ae isolates, which may affect virus replication and translation of hepatitis B e antigen. Subtypes Aa and Ae of genotype A deserve evaluation for any clinical differences between them, with a special reference to hepatocellular carcinoma prevalent in Africa.
Fun at the debtors’ prison: undergraduates’ university experience
Authors: Curtis Susan
Published: March 2004

Discusses the results of a survey on aspects of the student experience of university at a semi‐rural faculty of a metropolitan university in England. The research found that students tend to be amassing considerable debts, have little financial parental support and some are dependent on wages from part‐time work as a source of funds. It would appear that there is some financial hardship among the students surveyed, with a significant minority paying their own tuition fees. It is a possibility that these students would not be able to afford the higher top‐up tuition fees proposed for 2006. However, it is suggested that, for most students, there is a good social life, and university remains an enjoyable experience.
Assessing the role of internships in the career‐oriented employment of graduating college students
Authors: Callanan Gerard , Benzing Cynthia
Published: March 2004

This study assessed the relationship between the completion of an internship assignment prior to graduation and subsequent employment in a career‐oriented job after graduation. Subjects for the study were 163 seniors graduating with a business degree from a large public university in the mid‐Atlantic region of the USA. Results showed that the completion of an internship assignment was linked with finding career‐oriented employment, but was not related to a higher level of confidence over personal fit with the position that was selected. The article presents implications for future research and discusses practical issues related to the use of internships and other forms of anticipatory socialization in the career management activities of college students.
An assessment of the effectiveness of e‐learning on university space planning and design
Authors: Shabha Ghasson
Published: March 2004

It is apparent that the world of education has changed over the last ten years. The emergence of newly adopted metaphors such as “virtual university”, “networked campus” and “e‐university” may have yet greater impact on a university’smodus operandiin the twenty‐first century. This, in turn, will require more flexibility to cater for varying learning, managerial and organisational requirements. In terms of space management this will shift the emphasis towards a more time‐flexible, space‐flexible and location‐flexible pattern which will becomesine qua nonfor e‐learning. One of the main questions this paper is seeking to address is “what are the wider implications of these changes on the space design and management of existing university buildings?”. An attempt will be made to examine, speculate and extrapolate the design implications on a selected building at the UCE , Perry Barr Campus in the West Midlands.
Barriers to colorectal cancer screening in a low income, urban population: a descriptive study
Authors: Brouse Corey H. , Basch Charles E. , Wolf Randi L. , Shmukler Celia
Published: March 2004

The purpose of this study was to describe barriers to colorectal cancer (CRC) screening among a low income, urban sample. Participants were active members, or dependents, of a health care workers' union the New York City metropolitan area. The sample comprised 60 men and women of various ethnicity. A panel design was used with telephone contacts for up to 6 months. Data were collected during each telephone interaction regarding barriers associated with receipt of CRC screening. The majority of the barriers were based on influences of significant others and social environment, time constraints, other health problems, stress, and crisis.
Comparative analysis of isolated cellular organelles by means of soft X‐ray contact microscopy with laser‐plasma source and transmission electron microscopy
Authors: Limongi T. , Palladino L. , Tomassetti G. , Reale L. , Cesare P. , Flora F. , Aimola P. , Ragnelli A. M.
Published: March 2004

Soft X‐ray contact microscopy (SXCM) is, at present, a useful tool for the examination at submicrometre resolution of biological systems maintained in their natural hydrated conditions. Among current X‐ray‐generating devices, laser‐plasma sources are now easily available and, owing to their pulse nature, offer the opportunity to observe living biological samples before radiation damage occurs, even if the resolution achievable is not as high as with synchrotron‐produced X‐rays. To assess the potential of laser‐plasma source SXCM in the study of cellular organelles, we applied it for the analysis of chloroplasts extracted from spinach leaves and mitochondria isolated from bovine heart and liver. X‐ray radiation was generated by a nanosecond laser‐plasma source, produced by a single shot excimer XeCl laser focused onto an yttrium target. The images obtained with SXCM were then compared with those produced by transmission electron microscopy observation of the same samples prepared with negative staining, a technique requiring no chemical fixation, in order to facilitate their interpretation and test the applicability of SXCM imaging.
Use of truncated pyramid representation methodology in three‐dimensional reconstruction: an example
Authors: Papadimitriou C. , Yapijakis C. , Davaki P.
Published: March 2004

This paper describes a new methodology for three‐dimensional (3D) representation of biological structures contained in a series of sections, using an illustrative example. Spatial reconstruction of a specific area of an astrocytoma biopsy was carried out with alignment of the serial sections at an accuracy of 0.01% (or 1 µm cm−1), using the truncated pyramid representation (TPR) methodology. TPR includes: (a) serial tissue sectioning in a ribbon form; (b) alignment of the serial sections based on the properties of a ‘truncated pyramid’; (c) identification and localization of structures in every section using a field frame, and representation of the contours of the structures in every section as topographic contours (charting); (d) artificial reconstruction of the missing space between serial sections, by drawing intermediate contours based on the prototype contours of successive sections in order to provide smoother and more elegant representation of the volumes (complementation); and (e) 3D reconstruction. Application of TPR in a selected area of the astrocytoma enabled us to observe the morphology and spatial distribution of neoplastic astrocytic nuclei, which encircled an adjacent blood vessel.
Magnetic characterization of microscopic particles by MO‐SNOM
Authors: Schoenmaker J. , Lancarotte M. S. , Seabra A. C. , Souche Y. , Santos A. D.
Published: March 2004

This paper reports on the development of a magneto‐optical scanning near‐field optical microscope and the experimental near‐field study of the domain structure for a model magnetic particle of 16 × 16 µm2 of a Co70.4Fe4.6Si15B10 amorphous thin film, deposited on a silicon substrate. We present the topographic, optical and magneto‐optical differential susceptibility (MODS) images of the particle. Imaging by using the local MODS reveals the domain structure. These images are also used for positioning the tip in order to acquire local hysteresis loops, with submicrometre spatial resolution.
Interaction of mannan‐binding lectin with Trichinella spiralis glycoproteins, a possible innate immune mechanism
Authors: Gruden‐Movsesijan Alisa , Petrovic Miomir , Sofronic‐Milosavljevic Ljiljana
Published: March 2004

Complex and variable glycoconjugates presented by parasitic nematodes during infection are very important in the host–parasite interplay. Predominantly carbohydrate‐rich antigens are involved in the stimulation and modulation of the stage‐specific immune response of the host. The non‐specific innate immune system, however, acts as the first line of host defence against pathogens, before the appearance of antigen‐specific responses. The functional entities of the innate system are lectins that recognize the surface ligands of pathogens: mannan‐binding lectin (MBL) is a key recognition element involved in binding oligosaccharide structures exposed on microorganisms. In the present study we investigated whether MBL binds to the parasitic nematode Trichinella spiralis (T. spiralis). Since the parasite is coated with mannose‐containing glycans, these structures could represent potential ligands for MBL and contribute to activation of the innate immune response of the host. Histochemical staining revealed MBL on the surface and internal organs of T. spiralis muscle larvae. MBL bound in a mannose‐inhibitable manner to both crude extracts of T. spiralis muscle larvae and larvae excretory/secretory products. Western blot analyses showed that MBL recognized glycoproteins from all stages of T. spiralis. In vitro complement activation assays suggested that MBL is capable of fixing complement components on T. spiralis crude extract coated plates and activating the complement cascade through the ‘lectin pathway’.
Aspartyl proteases from the intestinal brush border of Haemonchus contortus as protective antigens for sheep
Authors: Smith W. D. , Skuce P. J. , Newlands G. F. J. , Smith S. K. , Pettit D.
Published: March 2004

A novel pepsin‐like aspartyl protease was identified as a component of Haemonchus galactose‐containing glycoprotein (H‐gal‐GP), which is an integral membrane glycoprotein complex located on the intestinal cells of Haemonchus contortus, and a highly protective antigen for sheep. This molecule, designated HcPEP2, showed 50% sequence identity with a previously described aspartyl protease from H‐gal‐GP known as HcPEP1. Fractions of H‐gal‐GP, either containing both HcPEP1 and 2 or other lower molecular weight components of the complex, were evaluated as protective antigens in immunization – challenge trials in sheep. When separated from the rest of the complex by gel filtration in 8 m urea, the HcPEP1 and 2 fraction significantly reduced H. contortus egg counts by 48% and worm numbers by 36%, but the lower molecular weight components were not significantly protective. However, the HcPEP1 and 2 fraction did not protect if electro‐eluted from SDS‐dissociated H‐gal‐GP, nor did bacterially expressed recombinant HcPEP1, suggesting that conformational epitopes are important for inducing immunity.
North American Natural Gas Demand — Outlook 2020
Authors: Ghouri Salman Saif
Published: March 2004

This paper analyses natural gas demand in the United States of America, Canada and Mexico for the period 1980–2001. Visual inspections of the historical trends for natural gas demand, GDP and natural gas prices provide an initial assessment of the relationship between these variables.

The paper then estimates various regressions, using the Almon polynomial distributed lag model. Several regressions, utilizing different combinations of explanatory variables, were run with or without an autoregressive scheme of various orders. The best‐fitting model in each country was selected, and this was later used to generate natural gas demand forecasts for the respective countries to 2020.

The results show that the income elasticity for Canada is more than unity (1.4), compared with less then unity for the USA (0.8) and Mexico (0.65). Changes in natural gas consumption generally lag by a few years, before the full impact of the changes in gas prices is realised. The length of lags varies between countries, but generally the effect of a price change on natural gas consumption dissipates after three‐to‐four years. Population is one of the important explanatory variables for Mexico, while it is not significant for the USA and Canada.

The model predicts that natural gas demand in the USA (base case) will increase at an annualised growth rate of 1.9 per cent and, by 2020, should reach 919 billion cubic metres. This is not very different to the forecast of the US Department of Energy's Energy Information Administration (EIA) (900 bcm). Similarly, the base case forecast for Canada to 2020 is 128 bcm, compared with that of the EIA at 125 bcm. The forecast for Mexico (92 bcm) is significantly different to that of the EIA (120), perhaps due to different economic growth assumptions.

The paper concludes that there is an urgent need to enhance exploration efforts and investment in liquefied natural gas (LNG) terminals, to be able to receive LNG from multiple sources, particularly in the USA and Mexico.
Beautiful and not So Beautiful Minds: An Introductory Essay on Economic Theory and the Supply of Oil
Authors: Banks Ferdinand E.
Published: March 2004

Both the teaching and the studying of energy economics have been severely disadvantaged by the emphasis placed on the Hotelling theory of resource exhaustion. For example, in the study of oil and gas economics, this unrealistic construction is fundamentally irrelevant, because it does not consider fixed investment, nor the (real) options associated with such things as starting or interrupting production. In addition, both economics students and teachers have failed to recognize the crucial importance of the reserves‐to production ratio, as well as the research of M. King Hubbert. Among other things, this oversight has led to an irrational optimism, where expectations of the future availability of oil are concerned. To be more specific, the probability that the oil pessimists are correct is large enough to justify the attachment of a sizable discount factor to any claim of future oil abundance.