bluuebunny/crossref_metadata_2025 huggingface dataset:
17β-Estradiol attenuates PDGF signaling in vascular smooth muscle cells at the postreceptor level
Authors: Kappert Kai , Caglayan Evren , Huntgeburth Michael , Bäumer Anselm T. , Sparwel Jan , Uebel Manuela , Rosenkranz Stephan
Published: October 2005

Estrogens are known to display significant vasoprotective effects in premenopausal women. PDGF is an important mediator of vascular smooth muscle cell (VSMC) migration and proliferation, and thus atherogenesis. We analyzed the effects of 17β-estradiol (E2) on β-PDGF receptor (β-PDGFR) expression/activation and PDGF-dependent VSMC proliferation, migration, and downstream signaling events. Pretreatment of VSMCs with E2 (0.3 μM–0.1 mM) for 24 h concentration-dependently inhibited PDGF-induced proliferation and migration up to 85.5 ± 15.8% and 79.4 ± 9.8%, respectively (both P < 0.05). These effects were prevented by coincubation with the ER antagonist ICI-182780. E2 did not alter β-PDGFR expression, nor did it impair the ligand-induced tyrosine phosphorylation of the β-PDGFR and consecutive binding of the receptor-associated signaling molecules Src homology region 2-containing phosphatase-2, PLC-γ, phosphatidylinositol 3-kinase, and RasGAP. Thus estrogens inhibited PDGF-induced cellular responses at the postreceptor level. Although stimulation of VSMCs with PDGF-BB led to a transient increase of rac-1 activity, pretreatment with E2 for 24 h concentration-dependently inhibited PDGF-induced rac-1 activation. Furthermore, inhibition of rac-1 by Clostridium sordellii lethal toxin or overexpression of dominant-negative rac-1 (rac-N17) significantly inhibited PDGF-induced VSMC migration, indicating that rac-1 activity is essential for PDGF-dependent cellular responses. E2 did not further reduce PDGF-induced migration in rac-N17-overexpressing cells, suggesting that it diminishes VSMC migration by altering rac-1 activity. We conclude that E2 attenuates PDGF-dependent cellular functions of VSMCs downstream of the β-PDGFR via inhibition of rac-1. These observations offer a molecular explanation for the vasoprotective effects of estrogens.
Characteristics and actions of NAD(P)H oxidase on the sarcoplasmic reticulum of coronary artery smooth muscle
Authors: Yi Xiu-Yu , Li Victoria X. , Zhang Fan , Yi Fan , Matson Daniel R. , Jiang Ming Tao , Li Pin-Lan
Published: October 2005

It has been reported that nonmitochondrial NAD(P)H oxidases make an important contribution to intracellular O2−· in vascular tissues and, thereby, the regulation of vascular function. Topological analyses have suggested that a well-known membrane-associated NAD(P)H oxidase may not release O2−· into the cytosol. It is imperative to clarify the source of intracellular O2−· associated with this enzyme and its physiological significance in vascular cells. The present study hypothesized that an NAD(P)H oxidase on the sarcoplasmic reticulum (SR) in coronary artery smooth muscle (CASM) regulates SR ryanodine receptor (RyR) activity by producing O2−· locally. Western blot analysis was used to detect NAD(P)H oxidase subunits in purified SR from CASM. Fluorescent spectrometric analysis demonstrated that incubation of SR with NADH time dependently produced O2−·, which could be substantially blocked by the specific NAD(P)H oxidase inhibitors diphenylene iodonium and apocynin and by SOD or its mimetic tiron. This SR NAD(P)H oxidase activity was also confirmed by HPLC analysis of conversion of NADH to NAD+. In experiments of lipid bilayer channel reconstitution, addition of NADH to the cis solution significantly increased the activity of RyR/Ca2+release channels from these SR preparations from CASM, with a maximal increase in channel open probability from 0.0044 ± 0.0005 to 0.0213 ± 0.0018; this effect of NADH was markedly blocked in the presence of SOD or tiron or the NAD(P)H oxidase inhibitors diphenylene iodonium, N-vanillylnonanamide, and apocynin. These results suggest that a local NAD(P)H oxidase system on SR from CASM regulates RyR/Ca2+channel activity and Ca2+release from SR by producing O2−·.
Hypoxia-induced reactive oxygen species downregulate ETBreceptor-mediated contraction of rat pulmonary arteries
Authors: Wang Xiaohua , Tong Mei , Chinta Shashi , Raj J. Usha , Gao Yuansheng
Published: October 2005

Production of reactive oxygen species (ROS) may be increased during hypoxia in pulmonary arteries. In this study, the role of ROS in the effect of hypoxia on endothelin (ET) type B (ETB) receptor-mediated vasocontraction in lungs was determined. In rat intrapulmonary (∼0.63 mm ID) arteries, contraction induced by IRL-1620 (a selective ETBreceptor agonist) was significantly attenuated after 4 h of hypoxia (30 mmHg Po2) compared with normoxic control (140 mmHg Po2). The effect was abolished by tiron, a scavenger of superoxide anions, but not by polyethylene glycol (PEG)-conjugated catalase, which scavenges H2O2. The hypoxic effect on ETBreceptor-mediated vasoconstriction was also abolished by endothelium denudation but not by nitro-l-arginine and indomethacin. Exposure for 4 h to exogenous superoxide anions, but not H2O2, attenuated the vasoconstriction induced by IRL-1620. Confocal study showed that hypoxia increased ROS production in pulmonary arteries that were scavenged by PEG-conjugated SOD. In endothelium-intact pulmonary arteries, the ETBreceptor protein was reduced after 4 h of exposure to hypoxia, exogenous superoxide anions, or ET-1. BQ-788, a selective ETBreceptor antagonist, prevented these effects. ET-1 production was stimulated in endothelium-intact arteries after 4 h of exposure to hypoxia or exogenous superoxide anions. This effect was blunted by PEG-conjugated SOD. These results demonstrate that exposure to hypoxia attenuates ETBreceptor-mediated contraction of rat pulmonary arteries. A hypoxia-induced production of superoxide anions may increase ET-1 release from the endothelium and result in downregulation of ETBreceptors on smooth muscle.
Transforming growth factor-β1 induces Nox4 NAD(P)H oxidase and reactive oxygen species-dependent proliferation in human pulmonary artery smooth muscle cells
Authors: Sturrock Anne , Cahill Barbara , Norman Kimberly , Huecksteadt Thomas P. , Hill Kenneth , Sanders Karl , Karwande S. V. , Stringham James C. , Bull David A. , Gleich Martin , Kennedy Thomas P. , Hoidal John R.
Published: October 2005

Transforming growth factor-β1 (TGF-β1) is abundantly expressed in pulmonary hypertension, but its effect on the pulmonary circulation remains unsettled. We studied the consequences of TGF-β1 stimulation on freshly isolated human pulmonary artery smooth muscle cells (HPASMC). TGF-β1 initially promoted differentiation, with upregulated expression of smooth muscle contractile proteins. TGF-β1 also induced expression of Nox4, the only NAD(P)H oxidase membrane homolog found in HPASMC, through a signaling pathway involving Smad 2/3 but not mitogen-activated protein (MAP) kinases. TGF-β1 likewise increased production of reactive oxygen species (ROS), an effect significantly reduced by the NAD(P)H oxidase flavoprotein inhibitor diphenylene iodonium (DPI) and by Nox4 siRNAs. In the absence of TGF-β1, Nox4 was present in freshly cultured cells but progressively lost with each passage in culture, paralleling a decrease in ROS production by HPASMC over time. At a later time point (72 h), TGF-β1 promoted HPASMC proliferation in a manner partially inhibited by Nox4 small interfering RNA and dominant negative Smad 2/3, indicating that TGF-β1 stimulates HPASMC growth in part by a redox-dependent mechanism mediated through induction of Nox4. HPASMC activation of the MAP kinases ERK1/2 was reduced by the NAD(P)H oxidase inhibitors DPI and 4-(2-aminoethyl)benzenesulfonyl fluoride, suggesting that TGF-β1 may facilitate proliferation by upregulating Nox4 and ROS production, with transient oxidative inactivation of phosphatases and augmentation of growth signaling cascades. These findings suggest that Nox4 is the relevant Nox homolog in HPASMC. This is the first observation that TGF-β1 regulates Nox4, with important implications for mechanisms of pulmonary vascular remodeling.
Expression and muscarinic receptor coupling of Lyn kinase in cultured human airway smooth muscle cells
Authors: Pertel Thomas , Zhu Defen , Panettieri Reynold A. , Yamaguchi Naoto , Emala Charles W. , Hirshman Carol A.
Published: October 2005

Src family tyrosine kinases are signaling intermediates in a diverse array of cellular events including cell differentiation, motility, proliferation, and survival. In nonairway smooth muscle cells, muscarinic receptors directly interact with Src family tyrosine kinases. As little is known about the expression and signaling of these Src family tyrosine kinases in human airway smooth muscle cells, we determined the expression of Src family members and characterized the muscarinic receptor-mediated activation of Lyn kinase in these cells. RT-PCR revealed mRNA transcripts for FYN, c- SRC, YES, FRK, and LYN. Fyn, c-Src, Yes, and Lyn were identified in cultured airway smooth muscle cells by immunoblot analysis. In both nontransformed human cultured airway smooth muscle cells and cells transduced with wild-type human Lyn kinase, carbachol increased Lyn kinase activity. Pertussis toxin pretreatment failed to block carbachol activation of Lyn kinase but did attenuate the carbachol-induced increase in ERK/MAPK phosphorylation. Moreover, carbachol inhibited adenylyl cyclase but failed to increase total inositol phosphate synthesis in these cells. The present study shows that Lyn kinase is expressed in human cultured airway smooth muscle cells at both the mRNA and protein levels and that carbachol, an M2 muscarinic receptor agonist in these cells, activates Lyn kinase by a pertussis toxin-insensitive signaling pathway.
Membrane capacitance and conductance changes parallel mucin secretion in the human airway epithelium
Authors: Danahay Henry , Atherton Hazel C. , Jackson Alan D. , Kreindler James L. , Poll Christopher T. , Bridges Robert J.
Published: October 2005

Measurement of the magnitude and kinetics of exocytosis from intact epithelia has historically been difficult. Using well-differentiated cultures of human bronchial epithelial cells, we describe the use of transepithelial impedance analysis to enable the real-time quantification of mucin secretagogue-induced changes in membrane capacitance (surface area) and conductance. ATPγS, UTP, ionomycin, and PMA induced robust increases in total cellular capacitance that were demonstrated to be dominated by a specific increase in apical membrane surface area. The UTP-induced increase in capacitance occurred in parallel with goblet cell emptying and the secretion of mucin and was associated with decreases in apical and basolateral membrane resistances. The magnitude and kinetics of the capacitance increases were dependent on the agonist and the sidedness of the stimulation. The peak increase in capacitance induced by UTP was ∼30 mucin granule fusions per goblet cell. Secretagogue-induced decreases in apical membrane resistance were independent of exocytosis, although each of the secretagogues induced profound reductions in basolateral membrane resistance. Transepithelial impedance analysis offers the potential to study morphological and conductance changes in cultured human bronchial epithelial cells.
Euripides, Hecuba 444-6/455-7, Helen 1465-77, Bacchae 565-75
Authors:   Willink
Published: October 2005

Three more passages in the cantica of Euripides, currently unsatisfactory, are discussed and conjecturally remedied.
Lagaroi: Le temps de la re-rythmisation de l'hexamètre
Authors: Steinrück Martin
Published: October 2005

According to many metricians, the original structure of the hexameter is based on the connection of two cola, not the recurrence of six dactyls. The time of the development of the first form into the second is, however, uncertain. The main evidence in favour of a pre-homeric rerhythmisation seems to consist in the fact that most of Homer can be read as metron-shaped verses. Although this seems certain, this certainty could result from a normalizing transmission of the 'Homeric' text. The history of irregular verses, such as the so-called lagaroi, offers some evidence that there were more irregular, i.e. colon-based, verses in Homer before the long scholarly tradition corrected them into metron-based schemas.
Priscian of Lydia, Commentator on the de Anima in the Tradition of Iamblichus
Authors:   Perkams
Published: October 2005

It has been argued that Priscian of Lydia (around 530), to whom the manuscripts ascribe only two short treatises, is the author of an extended commentary on the De anima, which is transmitted under the name of Simplicius. Our analysis confirms this: Priscian's Metaphrase of Theophrastus' Physics is the text which the commentator mentions as his own work. Consequently, its author, Priscian, also wrote the De anima commentary. The parallels between both texts show that the commentator sometimes does not quote Iamblichus directly, but borrowed Iamblichean formulations from the Metaphrase. As for the dating of his works, a comparison with Damascius' writings makes it probable that his On principles is a terminus post quem for the De anima commentary and a terminus ante quem for the Metaphrase. It is likely that both works were composed before 529.
Recollection and Philosophical Reflection in Plato's Phaedo
Authors: Franklin Lee
Published: October 2005

Interpretations of recollection in the Phaedo are divided between ordinary interpretations, on which recollection explains a kind of learning accomplished by all, and sophisticated interpretations, which restrict recollection to philosophers. A sophisticated interpretation is supported by the prominence of philosophical understanding and reflection in the argument. Recollection is supposed to explain the advanced understanding displayed by Socrates and Simmias (74b2-4). Furthermore, it seems to be a necessary condition on recollection that one who recollects also perform a comparison of sensible particulars to Forms (74a5-7). I provide a new ordinary interpretation which explains these features of the argument. First, we must clearly distinguish the philosophical reflection which constitutes the argument for the Theory of Recollection from the ordinary learning which is its subject. The comparison of sensibles to Forms is the reasoning by which we see, as philosophers, that we must recollect. At the same time, we must also appreciate the continuity of ordinary and philosophical learning. Plato wants to explain the capacity for ordinary discourse, but with an eye to its role as the origin of philosophical reflection and learning. In the Phaedo, recollection has ordinary learning as its immediate explanandum, and philosophical learning as its ultimate explanandum.
LXX Syntax and the Identity of the NT Veil
Authors: Gurtner Daniel
Published: October 2005

This article argues καταπɛτασμα in the NT (Mt. 27:51a, Mk. 15:38, Lk. 23:45, Hb. 6:19, 9:3, 10:20) is best identified as the inner veil before the holy of holies based on syntactical evidence from their LXX sources.
Cosmic Lordship and Divine Gift-Giving: Psalm 68 in Ephesians 4:8
Authors: Gombis Timothy
Published: October 2005

The use of Psalm 68 in Eph. 4:8 has proved to be one of the most difficult interpretive problems in this enigmatic letter. In this article I will engage and critique a leading interpretive proposal and then offer an alternative reading that exploits the elements of divine triumph and divine warfare in Ephesians, and that satisfactorily accounts for the quotation and elaboration by the writer in vv. 9-10.
The OT Background To Armageddon (Rev. 16:16 Revisited)
Authors: Jauhiainen Marko
Published: October 2005

This study challenges the old view, recently championed by John Day, according to which 'Aρμαγɛδων (Rev. 16:16) means the 'Mount of Megiddo' and is a conflation of 'Megiddo' in Zech. 12:11 and the 'mountains of Israel' in Ezek. 38-39. Instead of betraying Zecharian and Ezekielian influence, a closer inspection of the context of Rev. 16:16 points to the Isaian and Jeremian prophecies concerning the destruction of Babylon as a more plausible background of 'Aρμαγɛδων. It is concluded that the solution to the riddle of 'Armageddon' is most likely to be found in the etymological approach and that within this approach, an interpretation of the word as a reference to the "cutting down" of the "mountain" Babylon is perhaps more attractive than other alternatives.
Methodology in Identifying Patristic Citations in NT Textual Criticism
Authors: Osburn Carroll
Published: October 2005

After advancing a preliminary list of criteria for identifying patristic biblical citations, Fee called for adequate guidelines for assessing their text critical usability. Drawing illustrations primarily from fresh assessments of references in Hippolytus, Methodius, Origen, and Epiphanius for Novum Testamentum Graecum, Osburn advances the discussion regarding citations, adaptations, allusions, reminiscences, and locutions. Simple verbal precision is inadequate to establish a reference as reflective of an exemplar. A quotation must be read in its patristic context in order to determine how the text is actually used and in what way it probably reflects a text known to the Father.
Horkheimer and Adorno on Social Change: Problems and Potential in Light of “History from Below”1
Authors: Sawchuk Dana
Published: October 2005

In this article, I critique Max Horkheimer and Theodor Adorno's post-1940 pessimism regarding the potential for social change in modern society. The basis for this critique is the “history from below” perspective, a perspective which has particular relevance to the discussion since Horkheimer and Adorno's critical theory and history from below share certain core theoretical presuppositions. History from below's re-appropriation of past struggles and relational analysis of domination provide a useful antidote to critical theory's excessive focus on past suffering and its fatalistic interpretation of domination. The history from below perspective helps to demonstrate that Horkheimer and Adorno's pessimism was neither theoretically consistent nor inevitable.
The “Invisible Hand”: Neoclassical Economics and the Ordering of Society1
Authors: Finlayson Alan Christopher , Lyson Thomas A. , Pleasant Andrew , Schafft Kai A. , Torres Robert J.
Published: October 2005

The dominant economic discourse of the industrialized world — in political, academic, and popular terms — is neoclassical economics. A founding proposition is that an “invisible hand” aggregates individual decisions driven by rational self-interest into socially optimal outcomes. We draw upon economics as well as the sociology and philosophy of science to question this fundamental proposition and investigate neoclassical economic theory's dominant position in shaping social and political organization. We document the historical and contingent processes by which the neoclassical narrative has come to dominate the discursive space of capitalist societies. We argue that, contrary to claims of value neutrality, neoclassical economics functions as a master social narrative, or a technology of power, that concentrates power by transferring socio-economic decision-making from multiple sites to the centralized nodes of global economic and political institutions. This transfer occurs through the domination of the discursive space. The “invisible hand” is power.
The Politics of Economic Restructuring: Class Antagonism and the Neo-liberal Agenda
Authors: Green Brian , Huey Laura
Published: October 2005

This paper examines the underlying class dynamics of current political-economic policymaking at the global level, with a view to laying bare the class politics of mainstream market-oriented analysis. To this end, we read the texts of leading economists and policy-makers whose work has been highly influential in the formation of government policy in North America and Western Europe as a guide to the politics of economic restructuring, and to determine how class analysis and even class antagonism are embedded in — and indeed constitutive of — the theoretical and political discourses of high-level policy-makers and advisors.
The School System as a Post-Fordist Organization: Fragmented Centralization and the Emergence of IT Specialists
Authors: Monahan Torin
Published: October 2005

This article explores organizational restructuring in one large public institution as a situated response to the instabilities brought on by globalization. Drawing upon ethnographic research with the Los Angeles public school system, I argue that a process of fragmented centralization is taking place, whereby decision-making authority is becoming more centralized while accountability for centrally made decisions is becoming more distributed. This process is propelled, in part, by the rise of a new occupational group of information technology (IT) specialists that is integrating itself into places of power and altering all aspects of organizational operations, transforming the school system into a post-Fordist organization.
Andean Silver and the Rise of the Western World
Authors: Von Der Heydt-Coca Magda
Published: October 2005

This essay analyzes the contribution of Andean silver to the mercantile accumulation in the European core countries by tracing the flow of silver from the Andes to Asia. American Silver became the empowering currency that enabled the dynamic West European countries to dominate overseas trade. I argue that the influx of American silver into Europe over a period of three hundred years not only constituted a remarkable source of wealth, but also had systemic effects on the rise of the Western World influencing the global balance of power. I present a schematic model of the mercantile accumulation in the European hegemonic centers that explains how the external stimulus was amplified through positive feedback mechanisms.
Who Was Sitting in the Theatre at Miletos? An Epigraphical Application of a Novel Theory
Authors: Baker Murray
Published: October 2005

Stephen Mitchell has proposed a broad view of the cult of the Most High God (Theos Hypsistos) in which the followers describe themselves as theosebes. This provides a robust identity for these God-fearers and allows for a wider local context for theosebes (and other God-fearer) inscriptions. Mitchell's view is applied to the famous seating inscription for Jews and God-fearers in the theatre in Miletos. Here a quiet Jewish community is associated with a community of followers of the Most High God which included prominent members. The inscription should be read as, "The place of the Jews [real Jews], who are called [are part of] the group of theosebioi [followers of the Most High God]."
Jewish Life Before the Revolt: The Archaeological Evidence
Authors: Berlin Andrea
Published: October 2005

In this paper, I present material remains relevant to understanding Jewish life in Palestine in the century or so before the Revolt. These remains comprise tangible data by which Jewish praxis, actual behavior, as well as attitudes, can be identified and dated. From the early-mid first century B.C.E., Jews adopted what I call 'household Judaism'—using mikva'ot as well as locally manufactured oil, wine, and cooking vessels—in order to incorporate a religious sensibility into their daily lives. At the end of the first century B.C.E. and in the early first century C.E., they began using plain oil lamps and stone dishes as markers of ethnic solidarity and, perhaps, religious attitudes. Throughout these years, most Jews in Jerusalem and Judea followed identical burial practices, with understated funerals at undecorated family tombs. 'Household Judaism' developed outside halakhic or priestly concerns. The remains reflect shared beliefs as well as a broad desire for material possessions that would encode and reflect religious unity and identity.Other remains, however, tell a different story. In country villas and in the Upper City of Jerusalem, wealthy Jews embraced the use of decorated table vessels, Italian-style cooking pans, and foreign modes of dining. They built elaborate display tombs whose large courtyards and impressive façades provided a classicizing backdrop for staged funeral ceremonies. These sorts of remains are rare or absent in rural Judea, Jewish Galilee, and Gaulanitis. The archaeological evidence thus provides an eyewitness view of a population strongly unified in religious practices but sharply divided by cultural ethic. Over the course of the first century that division may have helped weaken the link between position and moral authority, which in turn created a space and a platform for new voices and different agendas.
Some Observations of Philo's De Gigantibus and Evil Spirits in Second Temple Judaism
Authors: Wright Archie
Published: October 2005

The following discussion delineates Philo's interpretation of Genesis 6:1-4 in his various treatises. The presentation provides a brief description of his understanding of the journey of the soul, which includes the origin of the soul, its place in the heavens, its time on earth, and its eventual return to the divine realm. Throughout the discussion, I will introduce various points of the interpretation of the Genesis passage found in the Watcher tradition of 1 Enoch and its adaptation in various documents in Second Temple Jewish literature. In doing so, I will highlight the similarities and differences between the interpretations which suggest Philo had knowledge of some form of the Watcher tradition and was perhaps attempting to write a corrective of its understanding of the problem of evil and the cause of human suffering in the first century C.E.
Differentiation of regions with atypical oligonucleotide composition in bacterial genomes
Authors: Reva Oleg N , Tümmler Burkhard
Published: October 2005

Background

Complete sequencing of bacterial genomes has become a common technique of present day microbiology. Thereafter, data mining in the complete sequence is an essential step. New in silico methods are needed that rapidly identify the major features of genome organization and facilitate the prediction of the functional class of ORFs. We tested the usefulness of local oligonucleotide usage (OU) patterns to recognize and differentiate types of atypical oligonucleotide composition in DNA sequences of bacterial genomes.

Results

A total of 163 bacterial genomes of eubacteria and archaea published in the NCBI database were analyzed. Local OU patterns exhibit substantial intrachromosomal variation in bacteria. Loci with alternative OU patterns were parts of horizontally acquired gene islands or ancient regions such as genes for ribosomal proteins and RNAs. OU statistical parameters, such as local pattern deviation (D), pattern skew (PS) and OU variance (OUV) enabled the detection and visualization of gene islands of different functional classes.

Conclusion

A set of approaches has been designed for the statistical analysis of nucleotide sequences of bacterial genomes. These methods are useful for the visualization and differentiation of regions with atypical oligonucleotide composition prior to or accompanying gene annotation.
CoaSim: A flexible environment for simulating genetic data under coalescent models
Authors: Mailund Thomas , Schierup Mikkel H , Pedersen Christian NS , Mechlenborg Peter JM , Madsen Jesper N , Schauser Leif
Published: October 2005

Background

Coalescent simulations are playing a large role in interpreting large scale intra-specific sequence or polymorphism surveys and for planning and evaluating association studies. Coalescent simulations of data sets under different models can be compared to the actual data to test the importance of different evolutionary factors and thus get insight into these.

Results

We have created the CoaSim application as a flexible environment for Monte Carlo simulation of various types of genetic data under equilibrium and non-equilibrium coalescent processes for a variety of applications. Interaction with the tool is through the Guile version of the Scheme scripting language. Scheme scripts for many standard and advanced applications are provided and these can easily be modified by the user for a much wider range of applications. A graphical user interface with less functionality and flexibility is also included. It is primarily intended as an exploratory and educational tool

Conclusion

CoaSim is a powerful tool because of its flexibility and ease of use. This is illustrated through very varied uses of the application, e.g. evaluation of association mapping methods, parametric bootstrapping, and design and choice of markers for specific questions
ProfNet, a method to derive profile-profile alignment scoring functions that improves the alignments of distantly related proteins
Authors: Ohlson Tomas , Elofsson Arne
Published: October 2005

Background

Profile-profile methods have been used for some years now to detect and align homologous proteins. The best such methods use information from the background distribution of amino acids and substitution tables either when constructing the profiles or in the scoring. This makes the methods dependent on the quality and choice of substitution table as well as the construction of the profiles.

Here, we introduce a novel method called ProfNet that is used to derive a profile-profile scoring function.

The method optimizes the discrimination between scores of related and unrelated residues and it is fast and straightforward to use. This new method derives a scoring function that is mainly dependent on the actual alignment of residues from a training set, and it does not use any additional information about the background distribution.

Results

It is shown that ProfNet improves the discrimination of related and unrelated residues. Further it can be used to improve the alignment of distantly related proteins.

Conclusion

The best performance is obtained using superfamily related proteins in the training of ProfNet, and a classifier that is related to the distance between the structurally aligned residues. The main difference between the new scoring function and a traditional profile-profile scoring function is that conserved residues on average score higher with the new function.
Prediction of β-barrel membrane proteins by searching for restricted domains
Authors: Mirus Oliver , Schleiff Enrico
Published: October 2005

Background

The identification of β-barrel membrane proteins out of a genomic/proteomic background is one of the rapidly developing fields in bioinformatics. Our main goal is the prediction of such proteins in genome/proteome wide analyses.

Results

For the prediction of β-barrel membrane proteins within prokaryotic proteomes a set of parameters was developed. We have focused on a procedure with a low false positive rate beside a procedure with lowest false prediction rate to obtain a high certainty for the predicted sequences. We demonstrate that the discrimination between β-barrel membrane proteins and other proteins is improved by analyzing a length limited region. The developed set of parameters is applied to the proteome of E. coli and the results are compared to four other described procedures.

Conclusion

Analyzing the β-barrel membrane proteins revealed the presence of a defined membrane inserted β-barrel region. This information can now be used to refine other prediction programs as well. So far, all tested programs fail to predict outer membrane proteins in the proteome of the prokaryote E. coli with high reliability. However, the reliability of the prediction is improved significantly by a combinatory approach of several programs. The consequences and usability of the developed scores are discussed.
The modeled structure of the RNA dependent RNA polymerase of GBV-C Virus suggests a role for motif E in Flaviviridae RNA polymerases
Authors: Ferron François , Bussetta Cécile , Dutartre Hélène , Canard Bruno
Published: October 2005

Background

The Flaviviridae virus family includes major human and animal pathogens. The RNA dependent RNA polymerase (RdRp) plays a central role in the replication process, and thus is a validated target for antiviral drugs. Despite the increasing structural and enzymatic characterization of viral RdRps, detailed molecular replication mechanisms remain unclear. The hepatitis C virus (HCV) is a major human pathogen difficult to study in cultured cells. The bovine viral diarrhea virus (BVDV) is often used as a surrogate model to screen antiviral drugs against HCV. The structure of BVDV RdRp has been recently published. It presents several differences relative to HCV RdRp. These differences raise questions about the relevance of BVDV as a surrogate model, and cast novel interest on the "GB" virus C (GBV-C). Indeed, GBV-C is genetically closer to HCV than BVDV, and can lead to productive infection of cultured cells. There is no structural data for the GBV-C RdRp yet.

Results

We show in this study that the GBV-C RdRp is closest to the HCV RdRp. We report a 3D model of the GBV-C RdRp, developed using sequence-to-structure threading and comparative modeling based on the atomic coordinates of the HCV RdRp structure. Analysis of the predicted structural features in the phylogenetic context of the RNA polymerase family allows rationalizing most of the experimental data available. Both available structures and our model are explored to examine the catalytic cleft, allosteric and substrate binding sites.

Conclusion

Computational methods were used to infer evolutionary relationships and to predict the structure of a viral RNA polymerase. Docking a GTP molecule into the structure allows defining a GTP binding pocket in the GBV-C RdRp, such as that of BVDV. The resulting model suggests a new proposition for the mechanism of RNA synthesis, and may prove useful to design new experiments to implement our knowledge on the initiation mechanism of RNA polymerases.
Evaluating eukaryotic secreted protein prediction
Authors: Klee Eric W , Ellis Lynda BM
Published: October 2005

Background

Improvements in protein sequence annotation and an increase in the number of annotated protein databases has fueled development of an increasing number of software tools to predict secreted proteins. Six software programs capable of high throughput and employing a wide range of prediction methods, SignalP 3.0, SignalP 2.0, TargetP 1.01, PrediSi, Phobius, and ProtComp 6.0, are evaluated.

Results

Prediction accuracies were evaluated using 372 unbiased, eukaryotic, SwissProt protein sequences. TargetP, SignalP 3.0 maximum S-score and SignalP 3.0 D-score were the most accurate single scores (90–91% accurate). The combination of a positive TargetP prediction, SignalP 2.0 maximum Y-score, and SignalP 3.0 maximum S-score increased accuracy by six percent.

Conclusion

Single predictive scores could be highly accurate, but almost all accuracies were slightly less than those reported by program authors. Predictive accuracy could be substantially improved by combining scores from multiple methods into a single composite prediction.
Regulation of actin cytoskeleton architecture by Eps8 and Abi1
Authors: Roffers-Agarwal Julaine , Xanthos Jennifer B , Miller Jeffrey R
Published: October 2005

Background

The actin cytoskeleton participates in many fundamental processes including the regulation of cell shape, motility, and adhesion. The remodeling of the actin cytoskeleton is dependent on actin binding proteins, which organize actin filaments into specific structures that allow them to perform various specialized functions. The Eps8 family of proteins is implicated in the regulation of actin cytoskeleton remodeling during cell migration, yet the precise mechanism by which Eps8 regulates actin organization and remodeling remains elusive.

Results

Here, we show that Eps8 promotes the assembly of actin rich filopodia-like structures and actin cables in cultured mammalian cells and Xenopus embryos, respectively. The morphology of actin structures induced by Eps8 was modulated by interactions with Abi1, which stimulated formation of actin cables in cultured cells and star-like structures in Xenopus. The actin stars observed in Xenopus animal cap cells assembled at the apical surface of epithelial cells in a Rac-independent manner and their formation was accompanied by recruitment of N-WASP, suggesting that the Eps8/Abi1 complex is capable of regulating the localization and/or activity of actin nucleators. We also found that Eps8 recruits Dishevelled to the plasma membrane and actin filaments suggesting that Eps8 might participate in non-canonical Wnt/Polarity signaling. Consistent with this idea, mis-expression of Eps8 in dorsal regions of Xenopus embryos resulted in gastrulation defects.

Conclusion

Together, these results suggest that Eps8 plays multiple roles in modulating actin filament organization, possibly through its interaction with distinct sets of actin regulatory complexes. Furthermore, the finding that Eps8 interacts with Dsh and induced gastrulation defects provides evidence that Eps8 might participate in non-canonical Wnt signaling to control cell movements during vertebrate development.
Macrodissection versus microdissection of rectal carcinoma: minor influence of stroma cells to tumor cell gene expression profiles
Authors: de Bruin Elza C , van de Pas Simone , Lips Esther H , van Eijk Ronald , van der Zee Minke MC , Lombaerts Marcel , van Wezel Tom , Marijnen Corrie AM , van Krieken J Han JM , Medema Jan Paul , van de Velde Cornelis JH , Eilers Paul HC , Peltenburg Lucy TC
Published: October 2005

Background

The molecular determinants of carcinogenesis, tumor progression and patient prognosis can be deduced from simultaneous comparison of thousands of genes by microarray analysis. However, the presence of stroma cells in surgically excised carcinoma tissues might obscure the tumor cell-specific gene expression profiles of these samples. To circumvent this complication, laser microdissection can be performed to separate tumor epithelium from the surrounding stroma and healthy tissue. In this report, we compared RNAs isolated from macrodissected, of which only surrounding healthy tissue had been removed, and microdissected rectal carcinoma samples by microarray analysis in order to determine the most reliable approach to detect the expression of tumor cell-derived genes by microarray analysis.

Results

As microdissection yielded low tissue and RNA quantities, extra rounds of mRNA amplification were necessary to obtain sufficient RNA for microarray experiments. These second rounds of amplification influenced the gene expression profiles. Moreover, the presence of stroma cells in macrodissected samples had a minor contribution to the tumor cell gene expression profiles, which can be explained by the observation that more RNA is extracted from tumor epithelial cells than from stroma.

Conclusion

These data demonstrate that the more convenient procedure of macrodissection can be adequately used and yields reliable data regarding the identification of tumor cell-specific gene expression profiles.
EST analysis in Ginkgo biloba: an assessment of conserved developmental regulators and gymnosperm specific genes
Authors: Brenner Eric D , Katari Manpreet S , Stevenson Dennis W , Rudd Stephen A , Douglas Andrew W , Moss Walter N , Twigg Richard W , Runko Suzan J , Stellari Giulia M , McCombie WR , Coruzzi Gloria M
Published: October 2005

Background

Ginkgo biloba L. is the only surviving member of one of the oldest living seed plant groups with medicinal, spiritual and horticultural importance worldwide. As an evolutionary relic, it displays many characters found in the early, extinct seed plants and extant cycads. To establish a molecular base to understand the evolution of seeds and pollen, we created a cDNA library and EST dataset from the reproductive structures of male (microsporangiate), female (megasporangiate), and vegetative organs (leaves) of Ginkgo biloba.

Results

RNA from newly emerged male and female reproductive organs and immature leaves was used to create three distinct cDNA libraries from which 6,434 ESTs were generated. These 6,434 ESTs from Ginkgo biloba were clustered into 3,830 unigenes. A comparison of our Ginkgo unigene set against the fully annotated genomes of rice and Arabidopsis, and all available ESTs in Genbank revealed that 256 Ginkgo unigenes match only genes among the gymnosperms and non-seed plants – many with multiple matches to genes in non-angiosperm plants. Conversely, another group of unigenes in Gingko had highly significant homology to transcription factors in angiosperms involved in development, including MADS box genes as well as post-transcriptional regulators. Several of the conserved developmental genes found in Ginkgo had top BLAST homology to cycad genes. We also note here the presence of ESTs in G. biloba similar to genes that to date have only been found in gymnosperms and an additional 22 Ginkgo genes common only to genes from cycads.

Conclusion

Our analysis of an EST dataset from G. biloba revealed genes potentially unique to gymnosperms. Many of these genes showed homology to fully sequenced clones from our cycad EST dataset found in common only with gymnosperms. Other Ginkgo ESTs are similar to developmental regulators in higher plants. This work sets the stage for future studies on Ginkgo to better understand seed and pollen evolution, and to resolve the ambiguous phylogenetic relationship of G. biloba among the gymnosperms.
Comparison between Gram stain and culture for the characterization of vaginal microflora: Definition of a distinct grade that resembles grade I microflora and revised categorization of grade I microflora
Authors: Verhelst Rita , Verstraelen Hans , Claeys Geert , Verschraegen Gerda , Van Simaey Leen , De Ganck Catharine , De Backer Ellen , Temmerman Marleen , Vaneechoutte Mario
Published: October 2005

Background

The microbiological diagnosis of bacterial vaginosis is usually made using Nugent's criteria, a useful but rather laborious scoring system based on counting bacterial cell types on Gram stained slides of vaginal smears. Ison and Hay have simplified the score system to three categories and added a fourth category for microflora with a predominance of the Streptococcus cell type. Because in the Nugent system several cell types are not taken into account for a final score, we carried out a detailed assessment of the composition of the vaginal microflora in relation to standard Gram stain in order the improve the diagnostic value of the Gram stain. To this purpose we compared Gram stain based categorization of vaginal smears with i) species specific PCR for the detection of Gardnerella vaginalis and Atopobium vaginae and with ii) tDNA-PCR for the identification of most cultivable species.

Results

A total of 515 samples were obtained from 197 pregnant women, of which 403 (78.3%) were categorized as grade I microflora, 46 (8.9%) as grade II, 22 (4.3%) as grade III and 8 (1.6%) as grade IV, according to the criteria of Ison and Hay. Another 36 samples (7.0%) were assigned to the new category 'grade I-like', because of the presence of diphtheroid bacilli cell types. We found that 52.7% of the grade I-like samples contained Bifidobacterium spp. while L. crispatus was present in only 2.8% of the samples and G. vaginalis and A. vaginae were virtually absent; in addition, the species diversity of this category was similar to that of grade II specimens.

Based on the presence of different Lactobacillus cell types, grade I specimens were further characterized as grade Ia (40.2%), grade Iab (14.9%) and grade Ib (44.9%). We found that this classification was supported by the finding that L. crispatus was cultured from respectively 87.0% and 76.7% of grade Ia and Iab specimens while this species was present in only 13.3% of grade Ib specimens, a category in which L. gasseri and L. iners were predominant.

Conclusion

Further refinement of Gram stain based grading of vaginal smears is possible by distinguishing additional classes within grade I smears (Ia, Iab and Ib) and by adding a separate category, designated grade I-like. A strong correlation was found between grade Ia and the presence of L. crispatus and between grade I-like and the presence of bifidobacteria. This refinement of Gram stain based scoring of vaginal smears may be helpful to improve the interpretation of the clinical data in future studies, such as the understanding of response to treatment and recurrence of bacterial vaginosis in some women, and the relationship between bacterial vaginosis and preterm birth.
ABC transporter FtsABCD of Streptococcus pyogenes mediates uptake of ferric ferrichrome
Authors: Hanks Tracey S , Liu Mengyao , McClure Michael J , Lei Benfang
Published: October 2005

Background

The Streptococcus pyogenes or Group A Streptococcus (GAS) genome encodes three ABC transporters, namely, FtsABCD, MtsABC, and HtsABC, which share homology with iron transporters. MtsABC and HtsABC are believed to take up ferric (Fe3+) and manganese ions and heme, respectively, while the specificity of FtsABCD is unknown.

Results

Recombinant FtsB, the lipoprotein component of FtsABCD, was found to bind Fe3+ ferrichrome in a 1:1 stoichiometry. To investigate whether FtsABCD transports Fe3+ ferrichrome, GAS isogenic strains defective in lipoprotein gene ftsB and permease gene ftsC were generated, and the effects of the mutations on uptake of Fe3+ ferrichrome were examined using radioactive 55Fe3+ ferrichrome. FtsB was produced in the wild-type strain but not in the ftsB mutant, confirming the ftsB inactivation. While wild-type GAS took up 3.6 × 104 Fe3+ ferrichrome molecules per bacterium per min at room temperature, the ftsB and ftsC mutants did not have a detectable rate of Fe3+ ferrichrome uptake. The inactivation of ftsB or ftsC also decreased 55Fe3+ ferrichrome uptake by >90% under growth conditions in the case of limited uptake time. Complementation of the ftsB mutant with a plasmid carrying the ftsB gene restored FtsB production and 55Fe3+ ferrichrome association at higher levels compared with the parent strain. The inactivation of mtsA and htsA and Fe-restricted conditions enhanced the production of FtsB and Fe3+ ferrichrome uptake.

Conclusion

The FtsB protein bound Fe3+ ferrichrome, and inactivation of ftsB or ftsC, but not htsA or mtsA, diminished Fe3+ ferrichrome uptake, indicating that FtsABCD, but not HtsABC and MtsABC, is the transporter that takes up Fe3+ ferrichrome in GAS. Fe acquisition systems are virulence factors in many bacterial pathogens and are attractive vaccine candidates. The elucidation of the FtsABCD specificity advances the understanding of Fe acquisition processes in GAS and may help evaluating the GAS Fe acquisition systems as vaccine candidates.