bluuebunny/crossref_metadata_2025 huggingface dataset:
A subset of bacterial inner membrane proteins integrated by the twin‐arginine translocase
Authors: Hatzixanthis Kostas , Palmer Tracy , Sargent Frank
Published: August 2003

A group of bacterial exported proteins are synthesized with N‐terminal signal peptides containing a SRRxFLK ‘twin‐arginine’ amino acid motif. Proteins bearing twin‐arginine signal peptides are targeted post‐translationally to the twin‐arginine translocation (Tat) system which transports folded substrates across the inner membrane. In Escherichia coli, most integral inner membrane proteins are assembled by a co‐translational process directed by SRP/FtsY, the SecYEG translocase, and YidC. In this work we define a novel class of integral membrane proteins assembled by a Tat‐dependent mechanism. We show that at least five E. coli Tat substrate proteins contain hydrophobic C‐terminal transmembrane helices (or ‘C‐tails’). Fusions between the identified transmembrane C‐tails and the exclusively Tat‐dependent reporter proteins TorA and SufI render the resultant chimeras membrane‐bound. Export‐linked signal peptide processing and membrane integration of the chimeras is shown to be both Tat‐dependent and YidC‐independent. It is proposed that the mechanism of membrane integration of proteins by the Tat system is fundamentally distinct from that employed for other bacterial inner membrane proteins.
CaSPA2is important for polarity establishment and maintenance inCandida albicans
Authors: Zheng Xin‐De , Wang Yan‐Ming , Wang Yue
Published: August 2003

Saccharomyces cerevisiaeSpa2p is a component of polarisome that controls cell polarity. Here, we have characterized the role of its homologue, CaSpa2p, in the polarized growth inCandida albicans. During yeast growth, GFP‐tagged CaSpa2p localized to distinct growth sites in a cell cycle‐dependent manner, while during hyphal growth it persistently localized to hyphal tips throughout the cell cycle. Persistent tip localization of the protein was also observed inCatup1Δ andCanrg1Δ, mutants constitutive for filamentous growth.Caspa2Δ exhibited defects in polarity establishment and maintenance, such as random budding and failure to confine growth to a small surface area leading to round cells with wide, elongated bud necks and markedly thicker hyphae. It was also defective in nuclear positioning, presumably a result of defective interactions between cytoplasmic microtubules with certain polarity determinants. The highly conserved SHD‐I and SHD‐V domains were found to be important and responsible for different aspects of CaSpa2p function.Caspa2Δ exhibited no virulence in the mouse systemic candidiasis model. Because of the existence of distinct growth forms and the easy control of the switch between themin vitro,C. albicansmay serve as a useful model in cell polarity research.
SpoIVB‐mediated cleavage of SpoIVFA could provide the intercellular signal to activate processing of Pro‐σK in Bacillus subtilis
Authors: Dong Tran C. , Cutting Simon M.
Published: August 2003

SpoIVB is the critical determinant for intercompartmental signalling of pro‐σK processing during sporulation in Bacillus subtilis. We show here that the SpoIVB serine peptidase can cleave the SpoIVFA protein, which is one component of the pro‐σK processing complex. SpoIVFA has been shown elsewhere (Rudner, D.Z., and Losick, R., 2002, Genes Dev 16: 1007–1018) to tether BofA and SpoIVFB in a membrane‐embedded heteroligomeric complex in which BofA directly inhibits the activity of SpoIVFB. Cleavage of SpoIVFA would provide the necessary signal to dissolve this complex and release BofA‐mediated inhibition on the zinc metalloprotease, SpoIVFB, that is responsible for cleaving pro‐σK to its mature form. We also show that the SpoIVB PDZ domain is required for self‐recognition and trans cleavage of SpoIVB and is probably also used to target an internal motif within the C‐terminal region of SpoIVFA exposed in the space between the inner and outer forespore membranes. This work reveals the mechanism of intercompartmental signalling and provides a unified model as to how σK‐directed gene expression in the mother cell is co‐ordinated with events in the forespore chamber.
Ku80 plays a role in non‐homologous recombination but is not required for T‐DNA integration inArabidopsis
Authors: Gallego M. E. , Bleuyard J.‐Y. , Daoudal‐Cotterell S. , Jallut N. , White C. I.
Published: August 2003

Chromosomal breaks are repaired by homologous recombination (HR) or non‐homologous end joining (NHEJ) mechanisms. The Ku70/Ku80 heterodimer binds DNA ends and plays roles in NHEJ and telomere maintenance in organisms ranging from yeast to humans. We have previously identified aku80mutant of the model plantArabidopsis thalianaand shown the role of Ku80 in telomere homeostasis in plant cells. We show here that this mutant is hypersensitive to the DNA‐damaging agent methyl methane sulphonate and has a reduced capacity to carry out NHEJ recombination. To understand the interplay between HR and NHEJ in plants, we measured HR in the absence of Ku80. We find that the frequency of intrachromosomal HR is not affected by the absence of Ku80. Previous work has clearly implicated the Ku heterodimer inAgrobacterium‐mediated T‐DNA transformation of yeast. Surprisingly,ku80mutant plants show no defect in the efficiency of T‐DNA transformation of plants withAgrobacterium, showing that an alternative pathway must exist in plants.
Mutation of the regulatory phosphorylation site of tobacco nitrate reductase results in constitutive activation of the enzyme in vivo and nitrite accumulation
Authors: Lillo Cathrine , Lea Unni S. , Leydecker Marie‐Thérèse , Meyer Christian
Published: August 2003

In wild‐type Nicotiana plumbaginifolia and other higher plants, nitrate reductase (NR) is rapidly inactivated/activated in response to dark/light transitions. Inactivation of NR is believed to be caused by phosphorylation at a special conserved regulatory Ser residue, Ser 521, and interactions with divalent cations and inhibitory 14‐3‐3 proteins. A transgenic N. plumbaginifolia line (S521) was constructed where the Ser 521 had been changed by site‐directed mutagenesis into Asp. This mutation resulted in complete abolishment of inactivation in response to light/dark transitions or other treatments known to inactivate NR. During prolonged darkness, NR in wild‐type plants is in the inactivated form, whereas NR in the S521 line is always in the active form. Differences in degradation rate between NR from S521 and lines with non‐mutated NR were not found. Kinetic constants like Km values for NADH and NO3− were not changed, but a slightly different pH profile was observed for mutated NR as opposed to non‐mutated NR. Under optimal growth conditions, the phenotype of the S521 plants was not different from the wild type (WT). However, when plants were irrigated with high nitrate concentration, 150 mm, the transgenic plants accumulated nitrite in darkness, and young leaves showed chlorosis.
A new single‐locus cytogenetic mapping system for maize (Zea mays L.): overcoming FISH detection limits with marker‐selected sorghum (S. propinquum L.) BAC clones
Authors: Koumbaris George L. , Bass Hank W.
Published: August 2003

The development of a cytogenetic map for maize (Zea mays L.) is shown to be feasible by means of a combination of resources from sorghum and oat that overcome limitations of single‐copy gene detection. A maize chromosome‐addition line of oat, OMAd9.2, provided clear images of optically isolated pachytene chromosomes through a chromosome spread and painting technique. A direct labeled oligonucleotide fluorescence in situ hybridization (FISH) probe MCCY specifically stained the centromere. The arm ratio (long/short) for maize chromosome 9 in the addition line was 1.7, comparable to the range of 1.6–2.1 previously reported for maize chromosome 9. A sorghum (Sorghum propinquum L.) BAC library was screened by hybridization with each of three maize core‐bin‐marker (CBM) probes: umc109 (CBM9.01), umc192/bz1 (CBM9.02), and csu54b (CBM9.08). A single BAC clone for each marker was chosen; designated sCBM9.1, sCBM9.2, or sCBM9.8; and used as a FISH probe on pachytene spreads from OMAd9.2. In each case, discrete FISH signals were observed, and their cytogenetic positions were determined to be 9S.79 (at position 79% of the length of chromosome 9 short arm) for sCBM9.1, 9S.65 for sCBM9.2, and approximately 9L.95 for sCBM9.8. These map positions were co‐linear with linkage‐map positions for these and other loci common to the linkage and cytogenetic maps. This work represents a major breakthrough for cytogenetic mapping of the maize genome, and also provides a general strategy that can be applied to cytogenetic mapping of other plant species with relatively large and complex genomes.
Identification of a locus controlling Verticillium disease symptom response in Arabidopsis thaliana
Authors: Veronese Paola , Narasimhan Meena L. , Stevenson Rebecca A. , Zhu Jian‐K. , Weller Stephen C. , Subbarao Krishna V. , Bressan Ray A.
Published: August 2003

Verticillium dahliae Klebahn is a soil‐borne fungal pathogen causing vascular diseases. The pathogen penetrates the host through the roots, spreads through the xylem, and systemically colonizes both resistant and susceptible genotypes. To elucidate the genetic and molecular bases of plant–Verticillium interactions, we have developed a pathosystem utilizing Arabidopsis thaliana and an isolate of V. dahliae pathogenic to both cruciferous and non‐cruciferous crops. Relative tolerance (based on symptom severity) but no immunity was found in a survey of Arabidopsis ecotypes. Anthocyanin accumulation, stunting, and chlorosis were common symptoms. Specific responses of the more susceptible ecotype Columbia were induction of early flowering and dying. The more tolerant ecotype C‐24 was characterized by pathogen‐induced delay of transition to flowering and mild chlorosis symptoms. Genetic analysis indicated that a single dominant locus, Verticillium dahliae‐tolerance (VET1), likely functioning also as a negative regulator of the transition to flowering, was able to convey increased tolerance. VET1 was mapped on chromosome IV. The differential symptom responses observed between ecotypes were not correlated with different rates of fungal tissue colonization or with differential transcript accumulation of PR‐1 and PDF1.2 defense genes whose activation was not detected during the Arabidopsis–V. dahliae interaction. Impairment in salicylic acid (SA)‐ or jasmonic acid (JA)‐dependent signaling did not cause hypersensitivity to the fungal infection, whereas ethylene insensitivity led to reduced chlorosis and ABA deficiency to reduced anthocyanin accumulation. The results of this study clearly indicated that the ability of V. dahliae to induce disease symptoms is also connected to the genetic control of development and life span in Arabidopsis.
Phosphorylation of sucrose synthase at serine 170: occurrence and possible role as a signal for proteolysis
Authors: Hardin Shane C. , Tang Guo‐Qing , Scholz Anke , Holtgraewe Daniela , Winter Heike , Huber Steven C.
Published: August 2003

Sequence analysis identified serine 170 (S170) of the maize (Zea mays L.) SUS1 sucrose synthase (SUS) protein as a possible, second phosphorylation site. Maize leaves contained two calcium‐dependent protein kinase activities and a calcium‐independent kinase activity with characteristics of an sucrose non‐fermenting 1 (SNF1)‐related protein kinase. Phosphorylation of the novel S170 and the known serine 15 (S15) site by these protein kinases was determined in peptide substrates and detected in SUS1 protein substrates utilizing sequence‐ and phosphorylation‐specific antibodies. We demonstrate phosphorylation of S170 in vitro and in vivo. The calcium‐dependent protein kinases phosphorylated both S170 and S15, whereas SNF1‐related protein kinase activity was restricted to S15. Calcium‐dependent protein‐kinase‐mediated S170 and S15 phosphorylation kinetics were determined in wild‐type and mutant SUS1 substrates. These analyses revealed that kinase specificity for S170 was threefold lower than that for S15, and that phosphorylation of S170 was stimulated by prior phosphorylation at the S15 site. The SUS‐binding peptides encoded by early nodulin 40 (ENOD40) specifically antagonized S170 phosphorylation in vitro. A model wherein S170 phosphorylation functions as part of a mechanism targeting SUS for proteasome‐mediated degradation is supported by the observations that SUS proteolytic fragments: (i) were detected and possessed relatively high phosphorylated‐S170 (pS170) stoichiometry; (ii) were spatially coincident with proteasome activity within developing leaves; and (iii) co‐sedimented with proteasome activity. In addition, full‐length pS170‐SUS protein was less stable than S170‐SUS in cultured leaf segments and was stabilized by proteasome inhibition. Post‐translational control of SUS protein level through pS170‐promoted proteolysis may explain the specific and significant decrease in SUS abundance that accompanies the sink‐to‐source transition in developing maize leaves.
Different pathways of homologous recombination are used for the repair of double‐strand breaks within tandemly arranged sequences in the plant genome
Authors: Orel Nadiya , Kyryk Anzhela , Puchta Holger
Published: August 2003

Different DNA repair pathways that use homologous sequences in close proximity to genomic double‐strand breaks (DSBs) result in either an internal deletion or a gene conversion. We determined the efficiency of these pathways in somatic plant cells of transgenic Arabidopsis lines by monitoring the restoration of the β‐glucuronidase (GUS) marker gene. The transgenes contain a recognition site for the restriction endonuclease I‐SceI either between direct GUS repeats to detect deletion formation (DGU.US), or within the GUS gene to detect gene conversion using a nearby donor sequence in direct or inverted orientation (DU.GUS and IU.GUS). Without expression of I‐SceI, the frequency of homologous recombination (HR) was low and similar for all three constructs. By crossing the different lines with an I‐SceI expressing line, DSB repair was induced, and resulted in one to two orders of magnitude higher recombination frequency. The frequencies obtained with the DGU.US construct were about five times higher than those obtained with DU.GUS and IU.GUS, irrespective of the orientation of the donor sequence. Our results indicate that recombination associated with deletions is the most efficient pathway of homologous DSB repair in plants. However, DSB‐induced gene conversion seems to be frequent enough to play a significant role in the evolution of tandemly arranged gene families like resistance genes.
The SOC1 MADS‐box gene integrates vernalization and gibberellin signals for flowering in Arabidopsis
Authors: Moon Jihyun , Suh Sung‐Suk , Lee Horim , Choi Kyu‐Ri , Hong Choo Bong , Paek Nam‐Chon , Kim Sang‐Gu , Lee Ilha
Published: August 2003

The floral transition in Arabidopsis is regulated by at least four flowering pathways: the long‐day, autonomous, vernalization, and gibberellin (GA)‐dependent pathways. Previously, we reported that the MADS‐box transcription factor SUPPRESSOR OF OVEREXPRESSION OF CO 1 (SOC1) integrates the long‐day and vernalization/autonomous pathways. Here, we present evidences that SOC1 also integrates signaling from the GA‐dependent pathway, a major flowering pathway under non‐inductive short days. Under short days, the flowering time of GA‐biosynthetic and ‐signaling mutants was well correlated with the level of SOC1 expression; overexpression of SOC1 rescued the non‐flowering phenotype of ga1‐3, and the soc1 null mutant showed reduced sensitivity to GA for flowering. In addition, we show that vernalization‐induced repression of FLOWERING LOCUS C (FLC), an upstream negative regulator of SOC1, is not sufficient to activate SOC1; positive factors are also required. Under short days, the GA pathway provides a positive factor for SOC1 activation. In contrast to SOC1, the GA pathway does not regulate expression of other flowering integrators FLC and FT. Our results explain why the GA pathway has a strong effect on flowering under short days and how vernalization and GA interact at the molecular level.
The Arabidopsis TDS4 gene encodes leucoanthocyanidin dioxygenase (LDOX) and is essential for proanthocyanidin synthesis and vacuole development
Authors: Abrahams Sharon , Lee Elizabeth , Walker Amanda R. , Tanner Gregory J. , Larkin Philip J. , Ashton Anthony R.
Published: August 2003

The anthocyanin and proanthocyanidin (PA) biosynthetic pathways share common intermediates until leucocyanidin, which may be used by leucoanthocyanidin dioxygenase (LDOX) to produce anthocyanin, or the enzyme leucoanthocyanidin reductase (LAR) to produce catechin, a precursor of PA. The Arabidopsis mutant tannin deficient seed 4 (tds4‐1) has a reduced PA level and altered pattern PA accumulation. We identified the TDS4 gene as LDOX by complementation of the tds4‐1 mutation either with a cosmid encoding LDOX or a 35S:LDOX construct. Independent Arabidopsis lines with a T‐DNA insertion in the LDOX gene had a similar phenotype, and one was allelic to tds4‐1. The seed phenotype of ban tds4 double mutants showed that LDOX precedes BANYULS (BAN) in the PA pathway, confirming recent biochemical characterisation of BAN as an anthocyanidin reductase. Double mutant analysis was also used to order the other TDS genes. Analysis of the PA intermediates in tds4‐1 revealed three dimethylaminocinnamaldehyde (DMACA) reacting compounds that accumulated in extracts from developing seeds. Analysis of Arabidopsis PA and its precursors indicates that Arabidopsis, unlike many other plants, exclusively uses the epicatechin and not the catechin pathway to PA. Transmission electron microscopy (TEM) showed that the pattern observed when seeds of tds4 were stained with DMACA was a result of the accumulation of PA intermediates in the cytoplasm of endothelial cells. Fluorescent marker dyes were used to show that tds4 endothelial cells had multiple small vacuoles, instead of a large central vacuole as observed in the wild types (WT). These results show that in addition to its established role in the formation of anthocyanin, LDOX is also part of the PA biosynthesis pathway.
ars1, an Arabidopsis mutant exhibiting increased tolerance to arsenate and increased phosphate uptake
Authors: Lee David A. , Chen Alice , Schroeder Julian I.
Published: August 2003

Arsenic is one of the most toxic pollutants at contaminated sites, yet little is known about the mechanisms by which certain plants survive exposure to high arsenic levels. To gain insight into the mechanisms of arsenic tolerance in plants, we developed a genetic screen to isolate Arabidopsis thaliana mutants with altered tolerance to arsenic. We report here on the isolation of a mutant arsenic resisant 1 (ars1) with increased tolerance to arsenate. ars1 germinates and develops under conditions that completely inhibit growth of wild‐type plants and shows a semi‐dominant arsenic resistance phenotype. ars1 accumulates levels of arsenic similar to that accumulated by wild‐type plants, suggesting that ars1 plants have an increased ability to detoxify arsenate. However, ars1 plants produce phytochelatin levels similar to levels produced by the wild type, and the enhanced resistance of ars1 is not abolished by the γ‐glutamylcysteine synthetase inhibitor l‐buthionine sulfoxime (BSO). Furthermore, ars1 plants do not show resistance to arsenite or other toxic metals such as cadmium and chromium. However, ars1 plants do show a higher rate of phosphate uptake than that shown by wild‐type plants, and wild‐type plants grown with an excess of phosphate show increased tolerance to arsenate. Traditional models of arsenate tolerance in plants are based on the suppression of phosphate uptake pathways and consequently on the reduced uptake of arsenate. Our data suggest that arsenate tolerance in ars1 could be due to a new mechanism mediated by increased phosphate uptake in ars1. Models discussing how increased phosphate uptake could contribute to arsenate tolerance are discussed.
Nuclear translocation of the photoreceptor phytochrome B is necessary for its biological function in seedling photomorphogenesis
Authors: Huq Enamul , Al‐Sady Bassem , Quail Peter H.
Published: August 2003

The phytochrome (phy) family of sensory photoreceptors (phyA to phyE in Arabidopsis) enables plants to optimize their growth and development under natural light environments. Subcellular localization studies have shown that the photoreceptor molecule is induced to translocate from cytosol to nucleus by light, but direct evidence of the functional relevance of this translocation has been lacking. Here, using a glucocorticoid receptor‐based fusion protein system, we demonstrate that both photoactivation and nuclear translocation combined are necessary and sufficient for the biological function of phyB. Conversely, neither artificial nuclear translocation of non‐photoactivated phyB nor artificial retention of photoactivated phyB in the cytosol provides detectable biological activity. Together these data indicate that signal transfer from photoactivated phyB to its primary signaling partner(s) is localized in the nucleus, and conversely suggest the absence of a cytosolic pathway from photoactivated phyB to light‐responsive genes.
A novel procedure for the localization of viral RNAs in protoplasts and whole plants
Authors: Zhang Fengli , Simon Anne E.
Published: August 2003

Analysis of virus spread using co‐expressed reporter proteins has provided important details on cell‐to‐cell and long‐distance movement of viruses in plants. However, most viruses cannot tolerate insertion of large non‐viral segments or loss of any open‐reading frames, procedures required to detect viruses non‐evasively. A technique used to localize mRNAs intracellularly in yeast has been modified for detection of viral RNAs in whole plants. The technique makes use of the binding of the coat protein of MS2 bacteriophage (CPMS2) to a 19 base hairpin (hp). A fusion protein, consisting of the CPMS2, green fluorescent protein (GFP), and a nuclear localization signal (NLS), was nuclear‐localized upon transient expression in protoplasts. However, addition of the hp to the 3′ untranslated region of Turnip crinkle virus (TCV‐hp) and co‐transfection of the virus and fusion protein construct into protoplasts resulted in the re‐location of GFP to the cytoplasm. Neither the insertion of the hp nor the interaction with the fusion protein impaired any viral functions. Transgenic plants expressing the GFP‐NLS‐CPMS2 fusion protein were generated, and GFP was detected in nuclei of young plant cells. Foci of GFP cytoplasmic fluorescence were detected in TCV‐hp‐inoculated leaves at 2 days post‐inoculation. Later, GFP was detected in young leaves near the midvein and in the base (support) cells of trichomes in the vicinity of secondary and tertiary veins. In older leaves, cytoplasmic GFP could be visualized throughout many of the leaves. This technique should be amenable for detection of any virus with a transformable plant (or animal) host and may also prove useful for localizing properly engineered host RNAs.
Factors associated with the difference in score between women's and doctors' decisional conflict about hormone therapy: a multilevel regression analysis
Authors: Légaré France , Tremblay Stéphane , O'Connor Annette M. , Graham Ian D. , Wells Georges A. , Jacobsen Mary Jane
Published: August 2003

Objective: To explore factors associated with the difference in score between women's and doctors’ decisional conflict about hormone therapy (HT).

Design: Secondary analysis. Setting and participants: family doctors were randomized to prepare women for counselling about HT using either a decision aid or a pamphlet.

Main variables studied: After each counselling session, decisional conflict was assessed in women and doctors using the Decisional Conflict Scale (DCS) and the Provider Decision Process Assessment Instrument (PDPAI), respectively. The difference in score between the DCS and PDPAI was computed and entered as the dependent variable in a multilevel regression analysis.

Main outcome results: A total of 40 doctors and 167 women were included in the analysis. The intra‐doctor correlation coefficient was 0.25. Factors associated with women experiencing higher decisional conflict than their doctor were: age of doctor >45 years, women who were undecided about the best choice after the counselling session, women with a university degree and women who said that their doctor usually does not give them control over treatment decision. Factors associated with doctors experiencing more decisional conflict than women were: doctors who were undecided about the quality of the decision, length of visit <30 min and women who thought that the decision was shared with their doctor.

Conclusion: In order to reduce the disparities between women's and doctors’ decisional conflict about HT, interventions aimed at raising awareness of doctors about shared decision‐making should be encouraged.
Isolation and Antimicrobial Susceptibility of Aeromonas salmonicida in rainbow trout (Oncorhynchus mykiss) in Turkey Hatchery Farms
Authors: Kırkan Ş. , Göksoy E. Ö. , Kaya O.
Published: August 2003

Three Aeromonas salmonicida strains were isolated from the livers of 265 rainbow trouts sampled. The antibiotic susceptibility test results showed that A. salmonicida strains were susceptible to streptomycine and ciprofloxacin. However, they were resistant to amoxycilline + clavulanic acid, penicillin, erythromycine, oxytetracycline and cefuroxime sodium.
The Effect of Commercial Steam Pasteurization on the Levels of Enterobacteriaceae and Escherichia coli on Naturally Contaminated Beef Carcasses
Authors: Minihan D. , Whyte P. , O'Mahony M. , Collins J. D.
Published: August 2003

The aim of the study was to assess the reduction achieved by steam pasteurization of beef carcasses of Escherichia coli, Enterobacteriaceae and total aerobic mesophilic plate counts (APCs). In total, 30 carcass halves were exposed to steam pasteurization (90°C, 10 s exposure time) and the 30 corresponding carcass halves remained as untreated controls. The neck, midline and rump were sampled on each carcass half. Significant reductions in E. coli incidence (P < 0.05) and counts, 0.5 log10 CFU 1000 cm−2 (P < 0.05), were observed on rump sites only. Significant reductions (>0.8 log10 CFU 1000 cm−2) of Enterobacteriaceae were observed at all carcass sites sampled (P < 0.05). Enterobacteriaceae reductions (>2 log10 CFU 1000 cm−2) were highly significant at the more contaminated sites (P < 0.001). Reductions in total APCs were inconsistent. Steam pasteurization significantly reduced the level of E. coli and Enterobacteriaceae at more contaminated sites, but did not result in complete decontamination. Therefore, steam pasteurization should be classed as an aid to hygienic beef processing, but not as a critical control point.
Positive Identification of Newcastle Disease Virus Vaccine Strains and Detection of Contamination in Vaccine Batches by Restriction Site Analysis of the Matrix Protein Gene
Authors: Farsang A. , Wehmann E. , Soós T. , Lomniczi B.
Published: August 2003

Twelve vaccine batches prepared from avirulent vaccine strains of Newcastle disease virus produced by seven manufacturers were identified by analysis of the matrix (M) protein gene with restriction enzymes MboI and HinfI. The analyses have revealed the presence of the strain indicated by the manufacturers (namely B‐1, LaSota or Ulster 2C), except in one case when the vaccine contained strain V4 Queensland instead of VGGA as indicated. In addition, several batches of both monovalent and combined vaccines containing strain LaSota of the same company consistently disclosed contamination with strain B‐1. The mixed nature of the preparations was verified not only by the dual patterns of restriction fragments but also by separating the two components and identifying them individually. Restriction analysis of the M gene, by allowing positive identification of each of the lentogenic vaccine strains, should provide an improvement in controlling vaccine batches by revealing homologous contaminants or exchange of the vaccine strain.
Nitric oxide, human diseases and the herbal products that affect the nitric oxide signalling pathway
Authors: Achike Francis I , Kwan Chiu‐Yin
Published: August 2003

1. Nitric oxide (NO) is formed enzymatically from l‐arginine in the presence of nitric oxide synthase (NOS). Nitric oxide is generated constitutively in endothelial cells via sheer stress and blood‐borne substances. Nitric oxide is also generated constitutively in neuronal cells and serves as a neurotransmitter and neuromodulator in non‐adrenergic, non‐cholinergic nerve endings. Furthermore, NO can also be formed via enzyme induction in many tissues in the presence of cytokines.

2. The ubiquitous presence of NO in the living body suggests that NO plays an important role in the maintenance of health. Being a free radical with vasodilatory properties, NO exerts dual effects on tissues and cells in various biological systems. At low concentrations, NO can dilate the blood vessels and improve the circulation, but at high concentrations it can cause circulatory shock and induce cell death. Thus, diseases can arise in the presence of the extreme ends of the physiological concentrations of NO.

3. The NO signalling pathway has, in recent years, become a target for new drug development. The high level of flavonoids, catechins, tannins and other polyphenolic compounds present in vegetables, fruits, soy, tea and even red wine (from grapes) is believed to contribute to their beneficial health effects. Some of these compounds induce NO formation from the endothelial cells to improve circulation and some suppress the induction of inducible NOS in inflammation and infection.

4. Many botanical medicinal herbs and drugs derived from these herbs have been shown to have effects on the NO signalling pathway. For example, the saponins from ginseng, ginsenosides, have been shown to relax blood vessels (probably contributing to the antifatigue and blood pressure‐lowering effects of ginseng) and corpus cavernosum (thus, for the treatment of men suffering from erectile dysfunction; however, the legendary aphrodisiac effect of ginseng may be an overstatement). Many plant extracts or purified drugs derived from Chinese medicinal herbs with proposed actions on NO pathways are also reviewed.
Choline potentiates the pressor response evoked by glycyl‐glutamine or naloxone in haemorrhaged rats
Authors: Gürün M Sibel , Millington William R , Ulus Ismail H
Published: August 2003

1. Severe blood loss initially lowers arterial pressure through a central mechanism that is thought to involve opioid and cholinergic neurons. The present study tested the hypothesis that simultaneous administration of a cholinergic agonist and an opioid receptor antagonist would produce a synergistic effect in the treatment of haemorrhage. Specifically, we tested whether choline, a precursor of acetylcholine, potentiates the pressor effect of the β‐endorphin derived peptide glycyl‐glutamine (Gly‐Gln) or the opioid receptor antagonist naloxone following acute haemorrhage.

2. Conscious rats were treated intracerebroventricularly (i.c.v.) with choline chloride (180 nmol) alone or combined with Gly‐Gln (10 nmol) or naloxone (10 nmol) 2 min after blood withdrawal (2.5 mL/100 g bodyweight over 20 min) was completed; mean arterial pressure and heart rate were monitored for 30 min.

3. Combined treatment with choline and Gly‐Gln elevated mean arterial pressure but did not affect heart rate significantly. Choline and Gly‐Gln had no effect on cardiovascular function when administered alone to haemorrhaged rats or when given together to normotensive animals. Choline also potentiated the pressor and tachycardic effect of naloxone in haemorrhaged rats.

4. These data show that choline potentiates the pressor effect of Gly‐Gln and naloxone in haemorrhaged rats.
Combined pharmacological preconditioning with a G‐protein‐coupled receptor agonist, a mitochondrial KATP channel opener and a nitric oxide donor mimics ischaemic preconditioning
Authors: Otani Hajime , Okada Takayuki , Fujiwara Hiroyoshi , Uchiyama Takamichi , Sumida Tomohiko , Kido Masakuni , Imamura Hiroji
Published: August 2003

1. Although pharmacological preconditioning (PPC) has emerged as an alternative to ischaemic preconditioning (IPC) in cardioprotection, the efficacy of PPC compared with IPC has not been investigated. Because IPC is mediated by complex signalling cascades arising from multiple triggers, we have hypothesized that combined PPC is necessary to mimic IPC.

2. Isolated and perfused rat hearts underwent IPC by three cycles of 5 min ischaemia and 5 min reperfusion before 30 min global ischaemia followed by 120 min reperfusion. Adenosine (30 µmol/L), diazoxide (50 µmol/L) and s‐nitroso‐N‐acetylpenicillamine (SNAP; 50 µmol/L) were added for 25 min just before (pretreatment modality) or 45 min before (PPC modality) the index ischaemia.

3. Ischaemic preconditioning significantly improved isovolumic left ventricular (LV) function and reduced infarct size. Although pretreatment with adenosine, diazoxide or SNAP alone was capable of reducing infarct size, PPC with each drug alone or in a combination of two drugs except for diazoxide plus SNAP failed to reduce infarct size. In contrast, PPC in combination with adenosine, diazoxide and SNAP (triple combination PPC) conferred significant improvement of LV function and reduction of infarct size that was as effective as IPC.

4. Cardioprotection afforded by triple combination PPC was abolished by the Gi/o‐protein inhibitor pertussis toxin, the mitochondiral KATP channel inhibitor 5‐hydroxydecanoate or the nitric oxide (NO) scavenger 2‐(4‐carboxyphenyl)‐4,4,5,5‐tetramethyl imidazoline‐1‐oxyl 3‐oxide (carboxy‐PTIO).

5. Protein kinase C (PKC)‐ε in the particulate fraction was activated throughout preconditioning ischaemia and reperfusion. Although PKC‐ε was activated during treatment with adenosine, diazoxide or SNAP alone, it was inactivated after washout. In contrast, PKC‐ε remained activated after triple combination PPC. The PKC inhibitor chelerythrine abolished activation of PKC‐ε and cardioprotection afforded by IPC and triple combination PPC.

6. These results demonstrate that combined PPC with a G‐protein‐coupled receptor agonist, a mitochondrial KATP channel opener and an NO donor is necessary to mimic IPC and such synergistic cardioprotection is associated with enhanced and sustained activation of PKC‐ε.
Clarification of regional and local in situ stresses using the compact conical‐ended borehole overcoring technique and numerical analysis
Authors: Kang Seong‐Seung , Kim Jun‐Mo , Kaneko Katsuhiko , Sugawara Katsuhiko , Obara Yuzo
Published: August 2003

Abstract Stress measurement is performed to estimate the states of in situ rock stress at the Torigata open‐pit limestone mine in Japan using the compact conical‐ended borehole overcoring (CCBO) technique. A set of back and forward analyses are then carried out to evaluate the states of regional and local in situ rock stresses and the mine‐induced rock slope stability using a 3‐D finite element model. The maximum horizontal local in situ rock stress measured by the CCBO technique acts in the northeast–southwest direction. The horizontal regional tectonic stresses obtained by the back analysis are in good agreement with those of the horizontal local in situ rock stress measured by the CCBO technique. However, the horizontal regional tectonic stress is more compressive than the horizontal local in situ rock stress. This is because the horizontal regional stress due to gravity is not considered in the back‐analyzed horizontal regional tectonic stress, but it is included in the local in situ rock stress measured by the CCBO technique. The local stress obtained by the forward analysis, especially its horizontal components, is in good agreement with the horizontal local in situ rock stress measured by the CCBO technique, and the magnitude of the vertical normal stress increases more rapidly than those of the horizontal normal stresses with depth. As a result, the ratio of the horizontal normal stress to the vertical normal stress is largest at the nearest excavation level and decreases with depth. This means that the stress field within the mine‐induced rock slope is affected by the horizontal components of the local in situ rock stress.
Cenozoic deformation history of the Tancheng–Lujiang Fault Zone, north China, and dynamic implications
Authors: Zhang Yueqiao , Shi Wei , Dong Shuwen
Published: August 2003

Abstract Based on a field analysis of slip vectors from Cretaceous and Tertiary rocks and coupled with rift basin analysis in north China, the Cenozoic deformation history of the Tancheng–Lujiang (Tan–Lu) Fault Zone can be divided into three main phases: early Tertiary normal faulting and northwest–southeast extension; Miocene normal faulting and northeast–southwest to north‐northeast–south‐southwest extension; and Quaternary dextral strike–slip faulting and east‐northeast transpression. The early Tertiary extension, which was responsible for rift basin formation in north China, originated from back‐roll mantle convection induced by westward subduction of the Pacific Plate beneath the Asia continent. The Miocene extension occurred possibly in association with the process of the Japan Sea opening. The Quaternary dextral slip was mainly localized along the middle part of the Tan–Lu Fault Zone and resulted from the far‐field effect of the late‐stage India–Eurasia convergence.
Lithofacies and eruption ages of Late Cretaceous caldera volcanoes in the Himeji–Yamasaki district, southwest Japan: Implications for ancient large‐scale felsic arc volcanism
Authors: Yamamoto Takahiro
Published: August 2003

Abstract The Himeji–Yamasaki region in the Inner Zone of southwest Japan is underlain mainly by Late Cretaceous volcanic rocks called the Ikuno Group or the Hiromine and Aioi Groups. A new stratigraphic and geochronological study shows that the volcanic rocks in this area consist of 15 eroded caldera volcanoes between 82 and 65 Ma; they are, in order of decreasing age, the Hiromine, Hoden, Ibo, Okawachi, Seppikosan, Hayashida, Shinokubi, Fukusaki, Kurooyama, Ise, Fukadanigawa, Nagusayama, Matobayama, Yumesaki and Mineyama Formations. These calderas vary in diameter from 1 to 20 km and are bounded by steep unconformities; they coalesce and overlap each other. The individual caldera fills are composed mainly of single voluminous pyroclastic flow deposits, which are often interleaved with debris avalanche deposits and occasionally underlie lacustrine deposits. The intracaldera pyroclastic flow deposits are made up of massive, welded or non‐welded tuff breccia to lapilli tuff, and are characterized by their great thickness. The debris avalanche deposits are ill‐sorted breccia, generated by the collapse of the caldera wall toward the caldera floor during the pyroclastic‐flow eruption. The large calderas that are more than 10 km in diameter contain original values of approximately 100 km3 of intracaldera pyroclastic flow deposits. These large calderas are similar to the well‐known Valles‐type calderas in their dimensions, although it is uncertain whether their caldera floors are coherent plates or incoherent pieces. Conversely, the small calderas have diatreme‐like subsurface structures. The variety of the caldera volcanoes in this area is caused by the difference in the volume of caldera‐forming pyroclastic eruptions, as the large and small calderas coexisted. The caldera‐forming eruption rates in Late Cretaceous southwest Japan, including the studied area, were similar to those in late Cenozoic central Andes and northeast Honshu arc, Japan, but obviously smaller than those of late Cenozoic intracratonic caldera clusters in western North America and the Quaternary extensional volcanic arcs in Taupo, New Zealand. The widespread Late Cretaceous felsic igneous rocks in southwest Japan were generated by a long‐term accumulation of low‐rate granitic magmatism at the eastern margin of the Eurasian Plate.
Evolution of the Komiji Syncline in the North Fossa Magna, central Japan: Paleomagnetic and K–Ar age insights
Authors: Niitsuma Sachiko , Niitsuma Nobuaki , Saito Kazuo
Published: August 2003

Abstract We carried out paleomagnetic measurements and K–Ar dating on Neogene andesitic lavas and sills of the Shigarami Formation in North Fossa Magna, central Japan. The Shigarami Formation is distributed in the axial part of the Komiji Syncline in the folding zone of the southwestern North Fossa Magna. Results of the present study indicate that the Komiji Syncline was formed shortly after 4.42 ± 0.12 Ma during the Pliocene. The sedimentary rocks of the Shigarami Formation consist of shallow marine and fluvial deposits. Intrusions of andesitic sills are found in the shallow marine deposits and two andesitic lava flows are present in the fluvial deposits. Oriented samples were taken from the sills at four sites and from the lavas at three sites. The samples produced stable remanent magnetization through stepwise alternating field and thermal demagnetizations. Results of a positive fold test indicate that the stable remanent magnetizations concentrate around a mean reversed polarity of declination = 169.0°, inclination = −58.5° and 95% confidence limit = 9.0° after corrections have been made according to the direction of the bedding of the sedimentary rocks. Four fresh samples were selected for K–Ar dating from the samples used for paleomagnetic measurements. The groundmass of three samples taken from the sills yield ages of 4.42 ± 0.12, 4.49 ± 0.22 and 4.69 ± 0.13 Ma, whereas the one taken from the lower lava has an age of 5.91 ± 0.26 Ma. We believe that the Komiji Syncline was formed after the emplacement of lavas and sills in the area, because the descending Miocene strata were folded concordantly with the Shigarami Formation. The Pliocene and Pleistocene strata rest unconformably on the folded strata. The deformation might have progressed during the Pliocene, then slowed down in the Early Pleistocene. Our results suggest that the northwestward motion of the Philippine Sea Plate and the collision of the Tanzawa Block affected not only the South Fossa Magna, but also the North Fossa Magna.
Ultrahigh‐pressure metamorphic records hidden in zircons from amphibolites in Sulu Terrane, eastern China
Authors: Liu Fulai , Zhang Zeming , Katayama Ikuo , Xu Zhiqin , Maruyama Shigenori
Published: August 2003

Abstract The amphibolites occur sporadically as thin layers and blocks throughout the Sulu Terrane, eastern China. All analyzed amphibolite from outcrop and drill cores from prepilot drill hole CCSD‐PP1 and CCSD‐PP2, Chinese Continental Scientific Drilling Project in the Sulu Terrane, are retrograded eclogites overprinted by amphibolite‐facies retrograde metamorphism, with characteristic mineral assemblages of amphibole +  plagioclase + epidote ± quartz ± biotite ± ilmenite ± titanite. However, coesite and coesite‐bearing ultrahigh‐pressure (UHP) mineral assemblages are identified by Raman spectroscopy and electron microprobe analysis as inclusions in zircons separated from these amphibolites. In general, coesite and other UHP mineral inclusions are preserved in the cores and mantles of zircons, whereas quartz inclusions occur in the rims of the same zircons. The UHP mineral assemblages consist mainly of coesite + garnet + omphacite +  rutile, coesite + garnet + omphacite, coesite + garnet + omphacite + phengite + rutile +  apatite, coesite + omphacite + rutile and coesite + magnesite. Compositions of analyzed mineral inclusions are very similar to those of matrix minerals from Sulu eclogites. These UHP mineral inclusion assemblages yield temperatures of 631–780°C and pressures of ≥2.8 × 103 MPa, representing the P–T conditions of peak metamorphism of these rocks, which are consistent with those (T = 642–726°C; P ≥ 2.8 × 103 MPa) deduced from adjacent eclogites. These data indicate that the amphibolites are the retrogressive products of UHP eclogites.
Lithology and palynology of Neogene sediments on the narrow edge of the Kitakami Massif (basement rocks), northeast Japan: Significant change for depositional environments as a result of plate tectonics
Authors: Yagishita Koji , Obuse Akiko , Kurita Hiroshi
Published: August 2003

Abstract A controversial stratigraphic section, the Taneichi Formation, is exposed along the Pacific Coast of northeastern Honshu, the main island of the Japanese Archipelago. Although most sediments of the formation have long been dated as late Cretaceous, the northern section of it has been assigned to (i) the Upper Cretaceous; (ii) the Paleogene; or (iii) the Neogene. In the present report, we present the data of palynological and sedimentological studies, showing that the northern section should be assigned to the Neogene. A more important point in the present study is that we invoke some basic principles of fluvial sedimentology to resolve this stratigraphic subject. The lignite layers full of Paleogene–Miocene dinoflagellate cysts and pollen assemblages drape over the boulder‐sized (>40 cm in diameter) clasts in the northern section. However, the layers totally consist of aggregates of small lignite chips, indicating that the lignites are allochthonous materials. The mega‐clasts with derived microfossils in the lignites are thought to have been deposited as Neogene fluvial (flood) sediments in the newly formed Japanese Archipelago. Prior to the Miocene, the northern Honshu was part of the Eurasian Plate, thus the boulder‐sized clasts cannot be envisaged as long river flood deposits along the continental Paleogene Pacific Coast. Instead, the mega‐clasts with the draping lignites were probably derived from nearby Miocene highlands in the newly born island arc.
Why evidence‐based practice now?: a polemic1
Authors: Walker Kim
Published: August 2003

Evidence‐based practice (EBP) first appeared on the healthcare horizon just over a decade ago. In 2003 its presence has intensified and extended beyond its initial relation to medicine embracing as it does now, nursing and the allied health disciplines. In this paper, I contend that its appearance and subsequent growth and development are the effects of potent ‘regimes of truth’, four of which bear the names: positivism, empiricism, pragmatism and economic rationalism. My aim is to show how EBP generates the controversy it does because its nature and methods are inextricably interwoven with the way it has become politicised and professionalised. This exegesis is an attempt to outline how the combined effects of the four forms of rationality mentioned above allow for both the methods and objectives of EBP to be constructed as they are, while at the same moment producing the particular effects of knowledge and power in terms of who sells and who buys the idea of EBP in the culture of contemporary healthcare.