bluuebunny/crossref_metadata_2025_split huggingface dataset:
Binding of the Escherichia coli MelR protein to the melAB promoter: orientation of MelR subunits and investigation of MelR–DNA contacts
Authors: Grainger David C. , Belyaeva Tamara A. , Lee David J. , Hyde Eva I. , Busby Stephen J. W. 
Published: January 2004

SummaryThe  Escherichia  coli
MelR  protein  is  a  melibiose‐triggered transcription factor, belonging to the AraC family, that activates transcription initiation at the melAB promoter. Activation is dependent on the binding of MelR to four 18 bp sites, centred at position −42.5 (site 2′), position −62.5 (site 2), position −100.5 (site 1) and position −120.5 (site 1′) relative to the melAB transcription start point. Activation also depends on the binding of CRP to a single site located between MelR binding site 1 and site 2. All members of the AraC family contain two helix–turn–helix (HTH) motifs that contact two segments of the DNA major groove at target sites on the same DNA face. In this work, we have studied the binding of MelR to different sites at the melAB promoter, focusing on the orientation of binding of the two MelR HTH motifs, and the juxtaposition of the different bound MelR subunits with respect to each other. To do this, MelR was engineered to contain a single cysteine residue adjacent to either one or the other HTH motif. The MelR derivatives were purified, and the cysteine residues were tagged with p‐bromoacetamidobenzyl‐EDTA‐Fe, an inorganic DNA cleavage reagent. Patterns of DNA cleavage after MelR binding were then used to determine the positions of the two HTH motifs at target sites. In order to simplify our analysis, we exploited an engineered derivative of the melAB promoter in which MelR binding to site 2 and site 2′, in the absence of CRP, is sufficient for transcription activation. To assist in the interpretation of our results, we also used a shortened derivative of MelR, MelR173, that is able to bind to site 2 but not to site 2′. Our results show that MelR binds as a direct repeat to site 2 and site 2′ with the C‐terminal HTH located towards the promoter‐proximal end of each site. The orientation in which MelR binds to site 2′ appears to be determined by MelR–MelR interactions rather than by MelR–DNA interactions. In complementary experiments, we used genetic analysis to investigate the importance of different residues in the two HTH motifs of MelR. Epistasis experiments provided evidence that supports the proposed orientation of binding of MelR at its target site.
Predicting mutation frequencies in stem–loop structures of derepressed genes: implications for evolution
Authors: Wright Barbara E. , Reschke Dennis K. , Schmidt Karen H. , Reimers Jacqueline M. , Knight William 
Published: January 2004

SummaryThis work provides evidence that, during transcription, the mutability (propensity to mutate) of a base in a DNA secondary structure depends both on the stability of the structure and on the extent to which the base is unpaired. Zuker's DNA folding computer program reveals the most probable stem–loop structures (SLSs) and negative energies of folding (–ΔG) for any given nucleotide sequence. We developed an interfacing program that calculates (i) the percentage of folds in which each base is unpaired during transcription; and (ii) the mutability index (MI) for each base, expressed as an absolute value and defined as ­follows: MI = (% total folds in which the base is unpaired) × (highest –ΔG of all folds in which it is unpaired). Thus, MIs predict the relative mutation or reversion frequencies of unpaired bases in SLSs. MIs for 16 mutable bases in auxotrophs, selected during starvation in derepressed genes, are compared with 70 background mutations in lacI and ebgR that were not derepressed during mutant selection. All the results are consistent with the location of known mutable bases in SLSs. Specific conclusions are: (i) Of 16 mutable bases in transcribing genes, 87% have higher MIs than the average base of the sequence analysed, compared with 50% for the 70 background mutations. (ii) In 15 of the mutable bases of transcribing genes, the correlation between MIs and relative mutation frequencies determined experimentally is good. There is no correlation for 35 mutable bases in the lacI gene. (iii) In derepressed auxotrophs, 100% of the codons containing the mutable bases are within one codon's length of a stem, compared with 53% for the background mutable bases in lacI. (iv) The data suggest that environmental stressors may cause as well      as     select     mutations     in     derepressed     genes. The implications of these results for evolution are discussed.
The regulation of EFG1 in white–opaque switching in Candida albicans involves overlapping promoters
Authors: Lachke Salil A. , Srikantha Thyagarajan , Soll David R. 
Published: January 2004

Summary EFG1
, which encodes a
trans
‐acting factor, is expressed as a more abundant 3.2 kb transcript in the white phase and as a less abundant 2.2 kb transcript in the opaque phase of the white–opaque transition in
Candida albicans
. To understand how alternative phase‐specific mRNAs are transcribed from the same gene locus, the 2320 bp upstream region of the gene was functionally characterized by analysing the ­activity of deletion derivatives in a luciferase‐based reporter system. The white phase‐specific promoter contained three discrete sequences involved in white phase‐specific activation, between −2022 and −1809 bp (AR1), between −1809 and −1727 bp (AR2) and between −922 and −840 bp (AR3). A higher resolution deletion and mutation analysis of AR2 revealed two regions between −1809 and −1787 bp and between −1764 and −1728 bp that are responsible for AR2 activation. Targeting of promoter constructs to the ectopic
ADE2
genomic site and the 3′ end of the
EFG1
genomic site revealed a positional requirement for white phase‐regulated activation specific for the AR2 region of the promoter. Gel mobility shift assays using AR2 revealed a white phase‐specific activation complex. No discrete activation sequences were identified in the overlapping promoter of the opaque phase‐specific
EFG1 transcript. The strength of opaque phase activation was directly proportional to the length of the promoter. Northern analysis excluded the possibility of an opaque phase‐specific repressor. These results demonstrate overlapping promoters for white and opaque phase‐specific expression of the gene for the transcription factor Efg1, with distinctly different mechanisms of phase‐specific activation.
Salmonella effectors translocated across the vacuolar membrane interact with the actin cytoskeleton
Authors: Miao Edward A. , Brittnacher Mitchell , Haraga Andrea , Jeng Robert L. , Welch Matthew D. , Miller Samuel I. 
Published: January 2004

SummaryA family of nine Salmonella typhimurium type III secretion effectors with a conserved amino‐terminus have been defined. Three family members (SifA, SifB and SseJ) have previously been demonstrated to localize to the Salmonella‐containing vacuole and to Salmonella‐induced filaments. In contrast, we demonstrate that two other family members, SspH2 and SseI, co‐localized with the polymerizing actin cytoskeleton. These proteins also interacted with the mammalian actin cross‐linking protein filamin in the yeast two‐hybrid assay through their highly conserved amino‐terminal domains. This amino‐terminus was sufficient to direct localization to the polymerizing actin cytoskeleton, suggesting that the interaction with filamin is important for this subcellular localization. In addition, SspH2 co‐localized with vacuole‐associated actin polymerizations (VAP) induced by intracellular bacteria through the Salmonella pathogenicity island (SPI)‐2 type III secretion system (TTSS). SspH2 interacted with the actin‐binding protein profilin in the yeast two‐hybrid assay and by affinity chromatography. This interaction was highly specific to SspH2 and was mediated by its carboxy‐terminus. Furthermore, SspH2 inhibited the rate of actin polymerization in vitro, suggesting that it functions to reduce or remodel VAP. Strains with mutations in sspH2 and sseI retained the ability to form VAP. However, a third intracellular virulence factor, spvB, which ADP‐ribosylates actin, strongly inhibited VAP formation in HeLa cells, suggesting a more subtle effect for SspH2 and SseI on the actin cytoskeleton.
Child and Adolescent Mental Health within Primary Care: A Study of General Practitioners’ Perceptions
Authors: Cockburn Katrina , Bernard Paul 
Published: January 2004

Background:  Primary Care teams play an important role in the provision of mental health care to children and young people.Methods:  We developed and distributed a questionnaire to all General Practitioners within one Health Authority area.Results:  Many of the respondents rated as less than satisfactory their competence and their knowledge and skills in important areas of child and adolescent mental health practice. A significant minority expressed a high level of interest in child and adolescent mental health and most respondents reported that they would value further training.Conclusions:  General Practitioners should be provided with more training and support in their role as providers of child and adolescent mental health care.
Evolving scholarly communication
Authors: Marks Jayne , Hannay Timo 
Published: January 2004

ABSTRACTNature Publishing Group and the Alliance for Cellular Signaling formed a partnership in 2001 to develop a comprehensive website for researchers in the field of cellular signalling. This website aimed to evolve the traditional form of publication (an article with supplementary information and links) into something that made better use of web and database technologies, and provided more value to scientists. This article gives an overview of the project, some of the thinking behind it, and how it has developed.
The Digital Curation Centre
Authors: Beagrie Neil 
Published: January 2004

ABSTRACTThe JISC and the EPSRCe‐science core programme will shortly establish a Digital Curation Centre to develop key research, services and generic tools needed by institutions and archiving bodies to manage digital collections over the long term. This article sets out its implications for ALPSP members and its relationship to other initiatives in e‐science and digital preservation.
Driving change in the research and HE information market
Authors: Roosendaal Hans E. 
Published: January 2004

ABSTRACTThis paper considers the concept of a federated network of repositories of information for research and higher education, and discusses the main engines for change in that market using availability and use of information as parameters. The value chain is used as an analytical instrument in considering options for, and roles and responsibilities of, the stakeholders. Strategic issues are discussed in the context of a high‐level strategy for this market and its consequences for the different stakeholders in the value chain, and in particular research and higher education institutions.
Between a rock and a hard place: the big squeeze for small publishers
Authors: Prosser David C. 
Published: January 2004

ABSTRACTThe combination of decreased income as a result of the RoweCom collapse, cuts in library budgets internationally, and the success of ‘Big Deal’ bundling arrangements has serious repercussions for smaller and society publishers. This paper argues that the scholarly communications environment is going to become increasingly hostile for these publishers and that their best chance of survival may be to adapt to new business models, in particular open access.
Publishing e‐books: OECD's pay‐per‐view and e‐library services 1998–2003
Authors: Green Toby 
Published: January 2004

ABSTRACTThe OECD's experience in transforming its stagnating book publishing business using the e‐book is described. OECD used e‐books first as a pay‐per‐view system within an online bookshop (1998). It then rejected e‐book aggregators and introduced an online library on a subscription basis (2001). OECD has moved from being a traditional print book publisher to being an online e‐book publisher, supported by print where necessary. It is now developing an e‐book account service inside its online bookshop and has the flexibility to publish a broader range of books to a wider audience than was previously possible.
The role of academic journal publications in the UK Research Assessment Exercise
Authors: Bence Valerie , Oppenheim Charles 
Published: January 2004

ABSTRACTThis article explores the evolution of the role of academic journal articles submitted to the UK's Research Assessment Exercise (RAE). By reviewing their role, it is possible to see how the changes from quantitative to qualitative assessment (and the resulting problems surrounding the definition of ‘quality’ journals) has impacted both on the journals selected by academics for publishing their research and the assessment of them. Although only one part of RAE submissions, the listing of published research outputs provides the primary evidence for research quality to most RAE panels, and is a significant driver of the final grade awarded, and thus the funding received by submitting institutions. The RAE, being a peer‐reviewed assessment exercise, mirrors in some ways the peer‐review process immured within scholarly publication. The developing role of journal publications as a vehicle for academic research output is examined via the chronology of the RAE, before assessing the current situation in which published journal output formed almost 70% of all output assessed by RAE panels in the latest exercise. The impact of this increased importance of academic journals in the assessment process is considered not only for academics but also for the wider community, i.e. publishers and libraries.
Liver Disease in Heavy Drinkers With and Without Alcohol Withdrawal Syndrome
Authors: Barrio E. , Tomé S. , Rodríguez I. , Gude F. , Sánchez‐Leira J. , Pérez‐Becerra E. , González‐Quintela A. 
Published: January 2004

Abstract: Background: Withdrawal syndrome is a hallmark of alcohol dependence. The characteristics of alcohol consumption, closely related to dependence, could influence the development of alcoholic liver disease. The study aimed to investigate if patients with severe alcohol withdrawal syndrome have a peculiar profile of liver disease.Methods: The study included 256 heavy drinkers (aged 19–75 years, 70.3% males) admitted to an Internal Medicine Department. Patients admitted for complications of liver disease were not included. Severe alcohol withdrawal syndrome (seizures, disordered perceptions, or delirium) developed in 150 patients (58.6%). Alcohol consumption (daily quantity, duration, and pattern [regular or irregular]) was assessed by questionnaire. Liver biopsy was performed in all cases.Results: Patients with alcohol withdrawal syndrome showed a lower prevalence of liver cirrhosis and a higher prevalence of alcoholic hepatitis than patients without it. The negative association of alcohol withdrawal syndrome with liver cirrhosis persisted after we adjusted for sex, daily intake, duration, and pattern of alcohol consumption. Alcoholic hepatitis was independently associated with the irregular pattern of alcohol consumption, which was closely associated with severe alcohol withdrawal syndrome.Conclusions: The profile of liver injury is different in heavy drinkers who develop and who do not develop a severe alcohol withdrawal syndrome when admitted to the hospital.
Endogenous Opioid Receptor Genes and Alcohol Dependence Among Taiwanese Han
Authors: Loh El Wui , Fann Cathy S. J. , Chang Yuh Terng , Chang Ching Jui , Cheng Andrew T. A. 
Published: January 2004

Abstract: Background: Nonselective opioid antagonists reduce alcohol consumption under various experimental situations, and several association studies have examined possible roles of opioid receptor mu (OPRM), delta (OPRD), and kappa (OPRK) genes in the development of alcohol dependence.Methods: We examined 20 single nucleotide polymorphisms (SNPs) across the OPRM, OPRD, and OPRK genes in 158 alcohol‐dependent subjects and 149 controls. Differences in allele frequency and genotype distribution between case subjects and controls, as well as the deviation from Hardy‐Weinberg equilibrium, were examined using Fisher's exact tests.Results: No significant difference in either allele or genotype frequency was found between case subjects and controls for each of the SNPs.Conclusions: Our findings do not support a possible role of the opioid receptor genes for the proclivity to alcohol dependence in the Taiwanese Han.
Survival of AA and ANA Rats During Lifelong Ethanol Exposure
Authors: Sarviharju Maija , Riikonen Jarno , Jaatinen Pia , Sinclair David , Hervonen Antti , Kiianmaa Kalervo 
Published: January 2004

Abstract: Background: Study of the long‐term effects of chronic alcohol consumption in human populations is confounded by genetic and environmental factors.Methods: The study was intended to investigate the effects on morbidity and survival of lifetime forced ethanol consumption in male and female AA (Alko, Alcohol) and ANA (Alko, Non‐Alcohol) rats. The ethanol‐exposed rats had 12% ethanol as the only available fluid from 3 to 24 months of age. The control groups had water. Rats that died during the experiment and those that were killed at 24 months of age were all autopsied, and the pathologic findings were recorded.Results: Lifelong ethanol consumption did not change the survival rate of the rats, and had no significant effect on the rates of any of the pathologic measures in either the AA or ANA line of rats, suggesting that this may not be a good animal model for studying the detrimental effects of chronic alcohol. An unexpected, highly significant finding was observed: the AA rats, bred for high voluntary ethanol drinking, lived much longer than the ANA rats, bred for ethanol avoidance. The death rate by 24 months in the AA line was less than one‐third of that in the ANA line. This difference was found regardless of whether the animals were maintained on alcohol or water, and in both genders. The AA rats had significantly lower rates of kidney disease, benign tumors, and cardiovascular disease than the ANA animals.Conclusions: Lifelong ethanol consumption increased neither the mortality nor the morbidity of AA and ANA line of rats. Genes selected in the development of the high drinking AA line have additional effects producing rats that are healthier and living longer than the ANA rats possessing genes resulting in alcohol avoidance.
Ethanol Withdrawal Influences Survival and Morphology of Developing Rat Hippocampal Neurons In Vitro
Authors: Lindsley Tara A. , Clarke Shannon 
Published: January 2004

Abstract: Background: Previous studies in this laboratory have shown that, like their counterparts in vivo, fetal rat hippocampal pyramidal neurons in culture develop abnormally small dendritic arbors when exposed to ethanol. This study asked whether ethanol's inhibitory effects on dendritic development differ when the duration of ethanol exposure and timing of withdrawal are varied to correspond with early versus later stages of development and whether ethanol withdrawal influences survival of these neurons.Methods: We compared neurons exposed continuously for 6 or 14 days to ethanol (70 mM) with neurons transferred from ethanol‐containing medium to control medium either 1 day after adding ethanol (before dendrites elongated) or 6 days after adding ethanol (after dendrites began elongating). We then performed morphometric and cell density analyses at 6 and 14 days using digital images of neurons immunostained with microtubule‐associated protein 2 (MAP2) to visualize dendrites.Results: Continuous exposure to ethanol decreased the length and number of dendrites formed but had no effect on neuron survival compared with controls without ethanol. Dendritic length was less inhibited when ethanol was withdrawn after 1 day, but the number of dendrites per cell was unchanged compared with neurons continuously exposed to ethanol. Withdrawal from ethanol at 1 day slightly enhanced the survival of neurons assessed at 14 days compared with neurons in control medium and with neurons exposed continuously to ethanol. In contrast, withdrawal from ethanol at 6 days severely decreased the number of neurons at 14 days.Conclusions: These results suggest that dendrites can achieve normal length when ethanol exposure is limited to only 1 day and withdrawal occurs before dendrites begin elongating. However, a persistent reduction in dendrite number results in smaller overall dendritic arbor size. Although continuous exposure to ethanol has little effect on neuron survival in these cultures, and exposure limited to 1 day followed by withdrawal can be neuroprotective against cell death associated with increased time in culture, longer exposure before withdrawal can trigger cell death.
Fetal Alcohol Exposure Alters GAP‐43 Phosphorylation and Protein Kinase C Responses to Contextual Fear Conditioning in the Hippocampus of Adult Rat Offspring
Authors: Tanner Daniel C. , Githinji Ann W. , Young Elizabeth A. , Meiri Karina , Savage Daniel D. , Perrone‐Bizzozero Nora I. 
Published: January 2004

Abstract:Background:The growth‐ and plasticity‐associated protein GAP‐43 plays a significant role in the establishment and remodeling of neuronal connections. We have previously shown that GAP‐43 levels, protein kinase C (PKC) activity, and GAP‐43 phosphorylation increase during contextual fear conditioning and that fetal alcohol exposure (FAE) decreases PKC activity and GAP‐43 phosphorylation in the hippocampus of adult offspring. Drawing on these observations, we hypothesized that FAE manifests its cognitive impairment by disrupting PKC activation and membrane translocation, thereby decreasing GAP‐43 phosphorylation and function.Methods:Three groups of pregnant rat dams (FAE and two control diet groups) were placed on different diet regimens. Offspring from each of these groups were placed into each of four test groups, a contextual fear conditioned (CFC) group, a naïve unhandled group, and two nonlearning stress control groups. Hippocampi were dissected, homogenized, and used to prepare a cytosolic and a membrane fraction. These fractions were probed for total GAP‐43, PKC‐phosphorylated GAP‐43, and several PKC subtypes. PKC activity also was measured in total homogenates.Results:Compared with both control diet groups, FAE animals showed a deficit in the activation of PKC in the hippocampus at 24 hr but not at 1.5 hr after CFC. Likewise, we found that the amount of GAP‐43 and its phosphorylation were decreased 24 hr after CFC in FAE rats but not at early times after training. Analysis of the translocation of various PKC isoforms revealed that FAE animals had decreased levels of membrane‐bound PKC β2and PKC ε 24 hr after CFC.Conclusions:Considering the role of PKC activation and GAP‐43 phosphorylation in synaptic plasticity, our results suggest that deficient translocation of PKC β2and PKC ε in the hippocampus may mediate the electrophysiological and behavioral deficits observed in fetal alcohol exposed animals.