bluuebunny/crossref_metadata_2025 huggingface dataset:
Genetic Complementation Screen Identifies a Mitogen-activated Protein Kinase Phosphatase, MKP3, as a Regulator of Dopamine Transporter Trafficking
Authors: Mortensen Ole Valente , Larsen Mads Breum , Prasad Balakrishna M. , Amara Susan G.
Published: April 2008

The antidepressant and cocaine sensitive plasma membrane monoamine transporters are the primary mechanism for clearance of their respective neurotransmitters and serve a pivotal role in limiting monoamine neurotransmission. To identify molecules in pathways that regulate dopamine transporter (DAT) internalization, we used a genetic complementation screen in Xenopus oocytes to identify a mitogen-activated protein (MAP) kinase phosphatase, MKP3/Pyst1/DUSP6, as a molecule that inhibits protein kinase C–induced (PKC) internalization of transporters, resulting in enhanced DAT activity. The involvement of MKP3 in DAT internalization was verified using both overexpression and shRNA knockdown strategies in mammalian cell models including a dopaminergic cell line. Although the isolation of MKP3 implies a role for MAP kinases in DAT internalization, MAP kinase inhibitors have no effect on internalization. Moreover, PKC-dependent down-regulation of DAT does not correlate with the phosphorylation state of several well-studied MAP kinases (ERK1/2, p38, and SAPK/JNK). We also show that MKP3 does not regulate PKC-induced ubiquitylation of DAT but acts at a more downstream step to stabilize DAT at the cell surface by blocking dynamin-dependent internalization and delaying the targeting of DAT for degradation. These results indicate that MKP3 can act to enhance DAT function and identifies MKP3 as a phosphatase involved in regulating dynamin-dependent endocytosis.
Coat-Tether Interaction in Golgi Organization
Authors: Guo Yusong , Punj Vasu , Sengupta Debrup , Linstedt Adam D.
Published: April 2008

Biogenesis of the Golgi apparatus is likely mediated by the COPI vesicle coat complex, but the mechanism is poorly understood. Modeling of the COPI subunit βCOP based on the clathrin adaptor AP2 suggested that the βCOP C terminus forms an appendage domain with a conserved FW binding pocket motif. On gene replacement after knockdown, versions of βCOP with a mutated FW motif or flanking basic residues yielded a defect in Golgi organization reminiscent of that occurring in the absence of the vesicle tether p115. Indeed, βCOP bound p115, and this depended on the βCOP FW motif. Furthermore, the interaction depended on E19E21in the p115 head domain and inverse charge substitution blocked Golgi biogenesis in intact cells. Finally, Golgi assembly in permeabilized cells was significantly reduced by inhibitors containing intact, but not mutated, βCOP FW or p115 EE motifs. Thus, Golgi organization depends on mutually interacting domains in βCOP and p115, suggesting that vesicle tethering at the Golgi involves p115 binding to the COPI coat.
A Mep2-dependent Transcriptional Profile Links Permease Function to Gene Expression during Pseudohyphal Growth inSaccharomyces cerevisiae
Authors: Rutherford Julian C. , Chua Gordon , Hughes Timothy , Cardenas Maria E. , Heitman Joseph
Published: April 2008

The ammonium permease Mep2 is required for the induction of pseudohyphal growth, a process in Saccharomyces cerevisiae that occurs in response to nutrient limitation. Mep2 has both a transport and a regulatory function, supporting models in which Mep2 acts as a sensor of ammonium availability. Potentially similar ammonium permease-dependent regulatory cascades operate in other fungi, and they may also function in animals via the homologous Rh proteins; however, little is known about the molecular mechanisms that mediate ammonium sensing. We show that Mep2 is localized to the cell surface during pseudohyphal growth, and it is required for both filamentous and invasive growth. Analysis of site-directed Mep2 mutants in residues lining the ammonia-conducting channel reveal separation of function alleles (transport and signaling defective; transport-proficient/signaling defective), indicating transport is necessary but not sufficient to sense ammonia. Furthermore, Mep2 overexpression enhances differentiation under normally repressive conditions and induces a transcriptional profile that is consistent with activation of the mitogen-activated protein (MAP) kinase pathway. This finding is supported by epistasis analysis establishing that the known role of the MAP kinase pathway in pseudohyphal growth is linked to Mep2 function. Together, these data strengthen the model that Mep2-like proteins are nutrient sensing transceptors that govern cellular differentiation.
Insulin Stimulates Phosphatidylinositol 3-Phosphate Production via the Activation of Rab5
Authors: Lodhi Irfan J. , Bridges Dave , Chiang Shian-Huey , Zhang Yanling , Cheng Alan , Geletka Lynn M. , Weisman Lois S. , Saltiel Alan R.
Published: April 2008

Phosphatidylinositol 3-phosphate (PI(3)P) plays an important role in insulin-stimulated glucose uptake. Insulin promotes the production of PI(3)P at the plasma membrane by a process dependent on TC10 activation. Here, we report that insulin-stimulated PI(3)P production requires the activation of Rab5, a small GTPase that plays a critical role in phosphoinositide synthesis and turnover. This activation occurs at the plasma membrane and is downstream of TC10. TC10 stimulates Rab5 activity via the recruitment of GAPEX-5, a VPS9 domain–containing guanyl nucleotide exchange factor that forms a complex with TC10. Although overexpression of plasma membrane-localized GAPEX-5 or constitutively active Rab5 promotes PI(3)P formation, knockdown of GAPEX-5 or overexpression of a dominant negative Rab5 mutant blocks the effects of insulin or TC10 on this process. Concomitant with its effect on PI(3)P levels, the knockdown of GAPEX-5 blocks insulin-stimulated Glut4 translocation and glucose uptake. Together, these studies suggest that the TC10/GAPEX-5/Rab5 axis mediates insulin-stimulated production of PI(3)P, which regulates trafficking of Glut4 vesicles.
Regulation of theCandida albicansCell Wall Damage Response by Transcription Factor Sko1 and PAS Kinase Psk1
Authors: Rauceo Jason M. , Blankenship Jill R. , Fanning Saranna , Hamaker Jessica J. , Deneault Jean-Sebastien , Smith Frank J. , Nantel Andre , Mitchell Aaron P.
Published: April 2008

The environmental niche of each fungus places distinct functional demands on the cell wall. Hence cell wall regulatory pathways may be highly divergent. We have pursued this hypothesis through analysis of Candida albicans transcription factor mutants that are hypersensitive to caspofungin, an inhibitor of beta-1,3-glucan synthase. We report here that mutations in SKO1 cause this phenotype. C. albicans Sko1 undergoes Hog1-dependent phosphorylation after osmotic stress, like its Saccharomyces cerevisiae orthologues, thus arguing that this Hog1-Sko1 relationship is conserved. However, Sko1 has a distinct role in the response to cell wall inhibition because 1) sko1 mutants are much more sensitive to caspofungin than hog1 mutants; 2) Sko1 does not undergo detectable phosphorylation in response to caspofungin; 3) SKO1 transcript levels are induced by caspofungin in both wild-type and hog1 mutant strains; and 4) sko1 mutants are defective in expression of caspofungin-inducible genes that are not induced by osmotic stress. Upstream Sko1 regulators were identified from a panel of caspofungin-hypersensitive protein kinase–defective mutants. Our results show that protein kinase Psk1 is required for expression of SKO1 and of Sko1-dependent genes in response to caspofungin. Thus Psk1 and Sko1 lie in a newly described signal transduction pathway.
A note on the false discovery rate and inconsistent comparisons between experiments
Authors: Higdon Roger , van Belle Gerald , Kolker Eugene
Published: April 2008

Motivation: The false discovery rate (FDR) has been widely adopted to address the multiple comparisons issue in high-throughput experiments such as microarray gene-expression studies. However, while the FDR is quite useful as an approach to limit false discoveries within a single experiment, like other multiple comparison corrections it may be an inappropriate way to compare results across experiments. This article uses several examples based on gene-expression data to demonstrate the potential misinterpretations that can arise from using FDR to compare across experiments. Researchers should be aware of these pitfalls and wary of using FDR to compare experimental results. FDR should be augmented with other measures such as p-values and expression ratios. It is worth including standard error and variance information for meta-analyses and, if possible, the raw data for re-analyses. This is especially important for high-throughput studies because data are often re-used for different objectives, including comparing common elements across many experiments. No single error rate or data summary may be appropriate for all of the different objectives.

Contact:  Eugene.Kolker@seattlechildrens.org
Prediction of disordered regions in proteins based on the meta approach
Authors: Ishida Takashi , Kinoshita Kengo
Published: April 2008

Motivation: Intrinsically disordered regions in proteins have no unique stable structures without their partner molecules, thus these regions sometimes prevent high-quality structure determination. Furthermore, proteins with disordered regions are often involved in important biological processes, and the disordered regions are considered to play important roles in molecular interactions. Therefore, identifying disordered regions is important to obtain high-resolution structural information and to understand the functional aspects of these proteins.

Results: We developed a new prediction method for disordered regions in proteins based on the meta approach and implemented a web-server for this prediction method named ‘metaPrDOS’. The method predicts the disorder tendency of each residue using support vector machines from the prediction results of the seven independent predictors. Evaluation of the meta approach was performed using the CASP7 prediction targets to avoid an overestimation due to the inclusion of proteins used in the training set of some component predictors. As a result, the meta approach achieved higher prediction accuracy than all methods participating in CASP7.

Availability:  http://prdos.hgc.jp/meta/

Contact:  t-ishida@hgc.jp
A global pathway crosstalk network
Authors: Li Yong , Agarwal Pankaj , Rajagopalan Dilip
Published: April 2008

Motivation: Given the complex nature of biological systems, pathways often need to function in a coordinated fashion in order to produce appropriate physiological responses to both internal and external stimuli. Therefore, understanding the interaction and crosstalk between pathways is important for understanding the function of both cells and more complex systems.

Results: We have developed a computational approach to detect crosstalk among pathways based on protein interactions between the pathway components. We built a global mammalian pathway crosstalk network that includes 580 pathways (covering 4753 genes) with 1815 edges between pathways. This crosstalk network follows a power-law distribution: P(k) ∼ k−γ, γ = 1.45, where P(k) is the number of pathways with k neighbors, thus pathway interactions may exhibit the same scale-free phenomenon that has been documented for protein interaction networks. We further used this network to understand colorectal cancer progression to metastasis based on transcriptomic data.

Contact:  yong.2.li@gsk.com

Supplementary information:  Supplementary data are available at Bioinformatics online.
“Business Responsibilities in a Divided World”: The Cold War Roots of the Corporate Social Responsibility Movement
Authors: Spector Bert
Published: April 2008

Both business executives and management scholars have, in recent years, focused a great deal of attention on the theme of corporate social responsibility (CSR). Calls for business leaders to expend resources on behalf of “social good” tend to downplay, if not ignore, what is fundamentally an ideological question: just what is a “good” society and who defines “goodness”? The ideological underpinnings of social responsibility and its relationship to the “good” society can be explored through an historical perspective. The roots of the CSR movement trace back to the early years of the Cold War. Led by Donald K David, Dean of the Harvard Business School and supported by other academics and executives given voice on the pages of the Harvard Business Review, advocates urged expanded business social responsibility as a means of aligning business interests with the defense of free-market capitalism against what was depicted as the clear-and-present danger of Soviet Communism. Today's enthusiastic calls for business to “do well by doing good” could benefit from a similar critical analysis not just of the goals of CSR but also the ideological assumptions, often unacknowledged, that underlie those goals.
Modelling combined harvest and effort regulations: the case of the Dutch beam trawl fishery for plaice and sole in the North Sea
Authors: Hoff A. , Frost H.
Published: April 2008

Hoff, A. and Frost, H. 2008. Modelling combined harvest and effort regulations: the case of the Dutch beam trawl fishery for plaice and sole in the North Sea. – ICES Journal of Marine Science, 65: 822–831. Currently, several European fishing fleets are regulated through a combination of harvest and effort control. The two regulation schemes are interrelated, i.e. a given quota limit will necessarily determine the effort used, and vice versa. It is important to acknowledge this causality when assessing combined effort and harvest regulation systems. A bioeconomic feedback model is presented that takes into account the causality between effort and harvest control by switching back and forth between the two, depending on which is the binding rule. The model consists of a biological and an economic operation module, the former simulating stock assessment and quota establishment, and the latter simulating the economic fleet dynamics. When harvest control is binding, catch is evaluated using the biological projection formula, whereas the economics-based Cobb–Douglas production function is used when effort is binding. The method is applied to the Dutch beam trawl fishery for plaice and sole in the North Sea.
Side-aspect target-strength measurements of bay anchovy (Anchoa mitchilli) and Gulf menhaden (Brevoortia patronus) derived from ex situ experiments
Authors: Boswell Kevin M. , Wilson Charles A.
Published: April 2008

Boswell, K. M., and Wilson, C. A. 2008. Side-aspect target strength measurements of bay anchovy (Anchoa mitchilli) and Gulf menhaden (Brevoortia patronus) derived from ex situ experiments. – ICES Journal of Marine Science, 65: 1012–1020. Acoustic target strengths (TSs) of bay anchovy [Anchoa mitchilli, 4.7–6.1 cm TL (total length)] and Gulf menhaden (Brevoortia patronus, 5.1–7.7 cm TL) were measured at 420 kHz during side-aspect, ex situ experiments. TS–TL relationships were derived at aspect for tethered individuals, and TS distributions were compared with those of free-swimming fish. Linear models relating TS and TL were fitted to data at two levels of pulse duration (0.1 and 0.4 ms) for both side aspect and all horizontal aspects of each species, and for pooled data from both species. At a pulse duration of 0.4 ms, the side-aspect TS–TL relationships were anchovy: TSlateral = 19.5 log10 TLcm − 62.4; menhaden: TSlateral = 26.1 log10 Lcm − 65.6; pooled: TSlateral = 32 log10 TLcm − 70.9. For all horizontal-aspect orientations (0–180° in the horizontal) the best-fit TS–TL relationships were anchovy: TSall = 20 log10 TLcm− 65.4; menhaden: TSall = 20 log10 TLcm − 64.8; pooled: TSall = 20 log10 TLcm− 65. However, the “fit” for all angles was consistently low. Predicted TS comparisons with side-aspect models presented by earlier authors suggested biases in the estimated TL. Additionally, relationships between TS and wet weight (W) were derived for each species. Variation in TS by pulse-duration level suggests the potential for the use of this factor as a time-dependent variable by which to separate echoes from bay anchovy and Gulf menhaden.
Random walk models in biology
Authors: Codling Edward A , Plank Michael J , Benhamou Simon
Published: April 2008

Mathematical modelling of the movement of animals, micro-organisms and cells is of great relevance in the fields of biology, ecology and medicine. Movement models can take many different forms, but the most widely used are based on the extensions of simple random walk processes. In this review paper, our aim is twofold: to introduce the mathematics behind random walks in a straightforward manner and to explain how such models can be used to aid our understanding of biological processes. We introduce the mathematical theory behind the simple random walk and explain how this relates to Brownian motion and diffusive processes in general. We demonstrate how these simple models can be extended to include drift and waiting times or be used to calculate first passage times. We discuss biased random walks and show how hyperbolic models can be used to generate correlated random walks. We cover two main applications of the random walk model. Firstly, we review models and results relating to the movement, dispersal and population redistribution of animals and micro-organisms. This includes direct calculation of mean squared displacement, mean dispersal distance, tortuosity measures, as well as possible limitations of these model approaches. Secondly, oriented movement and chemotaxis models are reviewed. General hyperbolic models based on the linear transport equation are introduced and we show how a reinforced random walk can be used to model movement where the individual changes its environment. We discuss the applications of these models in the context of cell migration leading to blood vessel growth (angiogenesis). Finally, we discuss how the various random walk models and approaches are related and the connections that underpin many of the key processes involved.
Combining experiments and modelling to understand size regulation inDictyostelium discoideum
Authors: Jang Wonhee , Gomer Richard H
Published: April 2008

Little is known about how the sizes of specific organs and tissues are regulated. To try to understand these mechanisms, we have been using a combination of modelling and experiments to study the simple systemDictyostelium discoideum, which forms approximately 20 000 cell groups. We found that cells secrete a factor, and as the number of cells increases, the concentration of the factor increases. Diffusion calculations indicated that this lets cells sense the local cell density. Computer simulations predicted, and experiments then showed, that this factor decreases cell–cell adhesion and increases random cell motility. In a group, adhesion forces keep cells together, while random motility forces cause cells to pull apart and separate from each other. As the group size increases above a threshold, the factor concentration goes above a threshold and the cells switch from an adhered state to a separated state. This causes excessively large groups to break apart and/or dissipate, creating an upper limit to group size. In this review, we focus on how computer simulations made testable predictions that led the way to understanding the size regulation mechanism mediated by this factor.
Tomographic techniques for the study of exceptionally preserved fossils
Authors: Sutton Mark D
Published: April 2008

Three-dimensional fossils, especially those preserving soft-part anatomy, are a rich source of palaeontological information; they can, however, be difficult to work with. Imaging of serial planes through an object (tomography) allows study of both the inside and outside of three-dimensional fossils. Tomography may be performed using physical grinding or sawing coupled with photography, through optical techniques of serial focusing, or using a variety of scanning technologies such as neutron tomography, magnetic resonance imaging and most usefully X-ray computed tomography. This latter technique is applicable at a variety of scales, and when combined with a synchrotron X-ray source can produce very high-quality data that may be augmented by phase-contrast information to enhance contrast. Tomographic data can be visualized in several ways, the most effective of which is the production of isosurface-based ‘virtual fossils’ that can be manipulated and dissected interactively.
The immunology of Epstein–Barr virus infection
Authors: Moss Denis J. , Burrows Scott R. , Silins Sharon L. , Misko Ihor , Khanna Rajiv
Published: April 2008

Epstein–Barr virus is a classic example of a persistent human virus that has caught the imagination of immunologists, virologists and oncologists because of the juxtaposition of a number of important properties. First, the ability of the virus to immortalize B lymphocytesin vitrohas provided an antigen presenting cell in which all the latent antigens of the virus are displayed and are available for systematic study. Second, the virus presents an ideal system for studying the immune parameters that maintain latency and the consequences of disturbing this cell–virus relationship. Third, this wealth of immunological background has provided a platform for elucidating the role of the immune system in protection from viral–associated malignancies of B cell and epithelial cell origin. Finally, attention is now being directed towards the development of vaccine formulations which might have broad application in the control of human malignancies.
Violent behaviour among people with schizophrenia: a framework for investigations of causes, and effective treatment, and prevention
Authors: Hodgins Sheilagh
Published: April 2008

Robust evidence has accumulated showing that individuals who develop schizophrenia are at elevated risk when compared to the general population to engage in violence towards others. This violence impacts negatively on victims as well as perpetrators and poses a significant financial burden to society. It is posited that among violent offenders with schizophrenia there are three distinct types defined by the age of onset of antisocial and violent behaviour. The early starters display a pattern of antisocial behaviour that emerges in childhood or early adolescence, well before illness onset, and that remains stable across the lifespan. The largest group of violent offenders with schizophrenia show no antisocial behaviour prior to the onset of the illness and then repeatedly engage in aggressive behaviour towards others. A small group of individuals who display a chronic course of schizophrenia show no aggressive behaviour for one or two decades after illness onset and then engage in serious violence, often killing, those who care for them. We hypothesize that both the developmental processes and the proximal factors, such as symptoms of psychosis and drug misuse, associated with violent behaviour differ for the three types of offenders with schizophrenia, as do their needs for treatment.
Roles of the plasticity regions of Helicobacter pylori in gastroduodenal pathogenesis
Authors: Yamaoka Yoshio
Published: April 2008

Putative virulence genes of Helicobacter pylori are generally classified into three categories: strain-specific genes, phase-variable genes and genes with variable structures/genotypes. Among these, there has recently been considerable interest in strain-specific genes found outside of the cag pathogenicity island, especially genes in the plasticity regions. Nearly half of the strain-specific genes of H. pylori are located in the plasticity regions in strains 26695 and J99. Strain HPAG1, however, seems to lack a typical plasticity region; instead it has 43 HPAG1-specific genes which are either undetectable or incompletely represented in the genomes of strains 26695 and J99. Recent studies showed that certain genes or combination of genes in this region may play important roles in the pathogenesis of H. pylori-associated gastroduodenal diseases. Most previous studies have focused on the plasticity region in strain J99 (jhp0914–jhp0961) and the jhp0947 gene and the duodenal ulcer promoting (dupA) gene are good candidate markers for gastroduodenal diseases although there are some paradoxical findings. The jhp0947 gene is reported to be associated with an increased risk of both duodenal ulcers and gastric cancers, whereas the dupA gene, which encompasses jhp0917 and jhp0918, is reported to be associated with an increased risk of duodenal ulcers and protection against gastric cancers. In addition, recent studies showed that approximately 10–30 % of clinical isolates possess a 16.3 kb type IV secretion apparatus (tfs3) in the plasticity region. Studies on the plasticity region have only just begun, and further investigation is necessary to elucidate the roles of genes in this region in gastroduodenal pathogenesis.
Signal-mediated interactions between Pseudomonas aeruginosa and Candida albicans
Authors: McAlester Gordon , O'Gara Fergal , Morrissey John P.
Published: April 2008

Pseudomonas aeruginosa causes infections in a wide variety of hosts and is the leading cause of mortality in cystic fibrosis (CF) patients. Although most clinical isolates of P. aeruginosa share common virulence determinants, it is known that strains evolve and change phenotypically during CF lung infections. These changes can include alterations in the levels of N-acyl homoserine lactones (HSLs), which are secreted signal molecules. In the CF lung, fungi, especially Candida albicans and Aspergillus fumigatus, may coexist with P. aeruginosa but the implications for disease are not known. Recent studies have established that signalling can occur between P. aeruginosa and C. albicans, with the bacterial molecule 3-oxo-C12HSL affecting Candida morphology, and the fungal metabolite farnesol reducing levels of the Pseudomonas quinolone signal and pyocyanin in Pseudomonas. Whether these interactions are common and typical in clinical strains of P. aeruginosa was addressed using CF isolates that produced varied levels of HSLs. It was found that, whereas some clinical P. aeruginosa strains affected C. albicans morphology, others did not. This correlated closely with the amounts of 3-oxo-C12HSL produced by the isolates. Furthermore, it was established that signalling is bidirectional and that the C. albicans molecule farnesol inhibits swarming motility in P. aeruginosa CF strains. This work demonstrates that clinical isolates of these opportunistic pathogens can interact in strain-specific ways via secreted signals and illustrates the importance of studying these interactions to fully understand the microbial contribution to disease in polymicrobial infections.
Extracellular proteolytic activities expressed by Bacillus pumilus isolated from endodontic and periodontal lesions
Authors: Johnson Blair T. , Shaw Lindsey N. , Nelson Daniel C. , Mayo John A.
Published: April 2008

The purpose of the present study was to identify 12Bacillusisolates that had been obtained from root canals of teeth requiring endodontic therapy and from periodontal pockets in severe marginal periodontitis, and to determine whether these isolates exhibited extracellular proteolytic activity and, usingin vitroassays, whether any such activity could degrade substrates that would be pathophysiologically relevant with regard to the production of endodontic and periodontal lesions. Biochemical and carbohydrate fermentation patterns were used in the identification of all strains, which was confirmed by determination of the16S rRNA gene sequence for strain BJ0055. Screening for production of extracellular proteolytic activity by all strains was done with a general proteinase substrate. All isolates were identified as representingBacillus pumilusand all exhibited extracellular proteolytic activity. The putative pathophysiological relevance of extracellular proteinase production in strain BJ0055 was assessed using fluorophore-labelled elastin and collagen and several chromogenic peptides. Probable classes of proteinases acting on each substrate were investigated using class-specific inhibitors. Activity–pH profiles were determined in buffers at different pH values. Extracellular activities that were caseinolytic, elastinolytic, collagenolytic, glutamyl endopeptidase-like, and alanyl tripeptidyl peptidase-like were observed. No trypsin-like activities were detected. Serine- and chymotrypsin-like serine proteinase activities were detected, with activity observed at neutral and alkaline, but not acidic, pH.B. pumilusstrains isolated from endodontic and periodontal lesions exhibited extracellular activities that degrade elastin, collagen and other substrates. These activities may be virulence factors that contribute to tissue damage in apical periodontitis and severe marginal periodontitis.
Analysis of Helicobacter pylori isolates from Chile: occurrence of selective type 1 Lewis b antigen expression in lipopolysaccharide
Authors: Altman Eleonora , Fernández Heriberto , Chandan Vandana , Harrison Blair A. , Schuster Myra Wilson , Rademacher Laura Otth , Toledo Claudio
Published: April 2008

Previous studies have shown that the LPS of Helicobacter pylori isolated from North American and European hosts predominantly expresses type 2 Lewis x (Lex) and Ley epitopes, whilst the LPS from Asian strains has the capacity to express type 1 Lea and Leb structures. The aim of this study was to evaluate the expression of Le antigens and the cytotoxin-associated antigen (CagA) by H. pylori isolates from Chile. A total of 38 isolates were screened. The expression of Le antigens and CagA was determined by whole-cell indirect ELISA, using commercially available monoclonal anti-Le and polyclonal anti-CagA antibodies. LPS profiles of H. pylori isolates were assessed by gel electrophoresis and Western blotting. Expression of Lex and/or Ley epitopes was confirmed in 32/38 isolates (84 %), whilst 9/38 isolates (24 %) expressed type 1 Leb blood group determinants, in addition to type 2 Lex and Ley structures. Six strains (16 %) were non-typeable. The majority of H. pylori strains examined were CagA-positive (83.3 %).
Identification of ancient remains through genomic sequencing
Authors: Blow Matthew J. , Zhang Tao , Woyke Tanja , Speller Camilla F. , Krivoshapkin Andrei , Yang Dongya Y. , Derevianko Anatoly , Rubin Edward M.
Published: April 2008

Studies of ancient DNA have been hindered by the preciousness of remains, the small quantities of undamaged DNA accessible, and the limitations associated with conventional PCR amplification. In these studies, we developed and applied a genomewide adapter-mediated emulsion PCR amplification protocol for ancient mammalian samples estimated to be between 45,000 and 69,000 yr old. Using 454 Life Sciences (Roche) and Illumina sequencing (formerly Solexa sequencing) technologies, we examined over 100 megabases of DNA from amplified extracts, revealing unbiased sequence coverage with substantial amounts of nonredundant nuclear sequences from the sample sources and negligible levels of human contamination. We consistently recorded over 500-fold increases, such that nanogram quantities of starting material could be amplified to microgram quantities. Application of our protocol to a 50,000-yr-old uncharacterized bone sample that was unsuccessful in mitochondrial PCR provided sufficient nuclear sequences for comparison with extant mammals and subsequent phylogenetic classification of the remains. The combined use of emulsion PCR amplification and high-throughput sequencing allows for the generation of large quantities of DNA sequence data from ancient remains. Using such techniques, even small amounts of ancient remains with low levels of endogenous DNA preservation may yield substantial quantities of nuclear DNA, enabling novel applications of ancient DNA genomics to the investigation of extinct phyla.
Reduction of Plastid-Localized Carbonic Anhydrase Activity Results in Reduced Arabidopsis Seedling Survivorship
Authors: Ferreira Fernando J. , Guo Cathy , Coleman John R.
Published: April 2008

Carbonic anhydrase (CA; EC 4.2.1.1) catalyzes the interconversion of CO2 and HCO3  − and is a major protein constituent of the C3 higher plant chloroplast where it is presumed to play a role in photosynthetic carbon assimilation. In this study, we have used both RNA antisense and gene knockout lines to specifically reduce the activity of the chloroplast βCA1 polypeptide (At3g01500) in the model plant Arabidopsis (Arabidopsis thaliana). Although able to germinate, seedling establishment of transgenic plants is significantly reduced relative to wild-type plants when grown at ambient levels of CO2. Growth at elevated (1,500 μL L−1) CO2 or on plates supplemented with sucrose restores seedling establishment rates to wild-type levels. Seed from wild-type and transgenic plants exhibited no significant differences in seed protein, lipid content, or reserve mobilization during seedling growth. βCA1-deficient seedlings do, however, exhibit reduced capacity for light-dependent 14CO2 assimilation prior to the development of true leaves. The small number of surviving seedlings able to grow and develop are phenotypically similar to wild-type plants, even when subsequently grown at subambient levels of CO2. Microarray analysis of mature leaves of βCA1-deficient plants shows some differences in transcript abundance, particularly with genes involved in ethylene signaling and response. The data suggest that reduced levels of seedling establishment by βCA1-deficient plants could be the result of poor cotyledon photosynthetic performance at the onset of phototrophic growth and prior to the development of true leaves.
Stable interactions between the transmembrane domains of the adenosine A2Areceptor
Authors: Thévenin Damien , Lazarova Tzvetana
Published: April 2008

G‐protein‐coupled receptors (GPCRs) must properly insert and fold in the membrane to adopt a stable native structure and become biologically active. The interactions between transmembrane (TM) helices are believed to play a major role in these processes. Previous studies in our group showed that specific interactions between TM helices occur, leading to an increase in helical content, especially in weakly helical TM domains, suggesting that helix–helix interactions in addition to helix–lipid interactions facilitate helix formation. They also demonstrated that TM peptides interact in a similar fashion in micelles and lipid vesicles, as they exhibit relatively similar thermal stability and α‐helicity inserted in SDS micelles to that observed in liposomes. In this study, we perform an analysis of pairwise interactions between peptides corresponding to the seven TM domains of the human A2Areceptor (A2AR). We used a combination of Förster resonance energy transfer (FRET) measurement and circular dichroism (CD) spectroscopy to detect and analyze these interactions in detergent micelles. We found that strong and specific interactions occur in only seven of the 28 possible peptide pairs. Furthermore, not all interactions, identified by FRET, lead to a change in helicity. Our results identify stabilizing contacts that are likely related to the stability of the receptor and that are consistent with what is known about the three‐dimensional structure and stability of rhodopsin and the β2adrenergic receptor.
Structural effects of Parkinson's disease linked DJ‐1 mutations
Authors: Malgieri Gaetano , Eliezer David
Published: April 2008

Mutations in the protein DJ‐1 are associated with familial forms of Parkinson's disease, indicating that DJ‐1 may be involved in pathways related to the etiology of this disorder. Here we have used solution state NMR and circular dichroism spectroscopies to evaluate the extent of structural perturbations associated with five different Parkinson's disease linked DJ‐1mutations: L166P, E64D, M26I, A104T, and D149A. Comparison of the data with those obtained for the wild‐type protein shows that the L166P mutation leads to severe and global destabilization and unfolding of the protein structure, while the structure of the E64D mutation, as expected, is nearly unperturbed. Interestingly, the remaining three mutants all show different degrees of structural perturbation, which are accompanied by a reduction in the thermodynamic stability of the protein. The observed structural and thermodynamic differences are likely to underlie any functional variations between these mutants and the wild type, which in turn are likely responsible for the pathogenicity of these mutations.
Human homolog of NOTUM, overexpressed in hepatocellular carcinoma, is regulated transcriptionally by β‐catenin/TCF
Authors: Torisu Yuichi , Watanabe Akira , Nonaka Aya , Midorikawa Yutaka , Makuuchi Masatoshi , Shimamura Takahiro , Sugimura Haruhiko , Niida Atsushi , Akiyama Tetsu , Iwanari Hiroko , Kodama Tatsuhiko , Zeniya Mikio , Aburatani Hiroyuki
Published: April 2008

The Drosophila Notum gene, which is regulated by the Wingless pathway, encodes a secreted hydrolase that modifies heparan sulfate proteoglycans. In comparative analysis of the gene expression profiles in primary human hepatocellular carcinomas (HCC) and normal organs, we observed that the human ortholog of Drosophila Notum was overexpressed markedly in a subset of HCC, but expressed rarely in adult normal tissues. Immunoblotting confirmed the overexpression of NOTUM protein in 12 of 40 primary HCC cases (30%). High levels of NOTUM protein were significantly associated with intracellular (nuclear or cytoplasmic) accumulation of β‐catenin protein: all 10 HCC with high intracellular β‐catenin also had high NOTUM expression, whereas only 2 of 30 cases (6.7%) without intracellular β‐catenin had high NOTUM expression (P < 0.00001). NOTUM expression in HepG2 cells was downregulated significantly by induction of a dominant‐negative mutant of TCF4, a β‐catenin partner. In vivo binding of the β‐catenin/TCF complex to the NOTUM promoter was demonstrated by chromatin immunoprecipitation in HepG2 and SW480 cells, where canonical Wnt signaling is activated constitutively. These findings provide evidence that NOTUM is a novel target of β‐catenin/TCF4 and is upregulated in Wnt/β‐catenin signaling‐activated HCC. (Cancer Sci 2008; 99: 1139–1146)
Endosomal sorting complex required for transport proteins in cancer pathogenesis, vesicular transport, and non‐endosomal functions
Authors: Tanaka Nobuyuki , Kyuuma Masanao , Sugamura Kazuo
Published: April 2008

Endosomal sorting complex required for transport (ESCRT) proteins form a multicomplex sorting machinery that controls multivesicular body (MVB) formation and the sorting of ubiquitinated membrane proteins to the endosomes. Being sorted to the MVB generally results in the lysosome‐dependent degradation of cell‐surface receptors, and defects in this machinery induce dysregulated receptor traffic and turnover. Recent lessons from gene targeting and silencing methodologies have implicated the ESCRT in normal development, cell differentiation, and growth, as well as in the budding of certain enveloped viruses. Furthermore, it is becoming apparent that the dysregulation of ESCRT proteins is involved in the development of various human diseases, including many types of cancers and neurodegenerative disorders. Here, we summarize the roles of ESCRT proteins in MVB sorting processes and the regulation of tumor cells, and we discuss some of their other functions that are unrelated to vesicular transport. (Cancer Sci 2008; 99: 1293–1303)
HrpN of Erwinia amylovora functions in the translocation of DspA/E into plant cells
Authors: BOCSANCZY ANA M. , NISSINEN RIITTA M. , OH CHANG‐SIK , BEER STEVEN V.
Published: April 2008

The type III secretion system (T3SS) is required by plant pathogenic bacteria for the translocation of certain bacterial proteins to the cytoplasm of plant cells or secretion of some proteins to the apoplast. The T3SS of Erwinia amylovora, which causes fire blight of pear, apple and other rosaceous plants, secretes DspA/E, which is an indispensable pathogenicity factor. Several other proteins, including HrpN, a critical virulence factor, are also secreted by the T3SS. Using a CyaA reporter system, we demonstrated that DspA/E is translocated into the cells of Nicotiana tabacum‘Xanthi’. To determine if other T3‐secreted proteins are needed for translocation of DspA/E, we examined its translocation in several mutants of E. amylovora strain Ea321. DspA/E was translocated by both hrpW and hrpK mutants, although with some delay, indicating that these two proteins are dispensable in the translocation of DspA/E. Remarkably, translocation of DspA/E was essentially abolished in both hrpN and hrpJ mutants; however, secretion of DspA/E into medium was not affected in any of the mentioned mutants. In contrast to the more virulent strain Ea273, secretion of HrpN was abolished in a hrpJ mutant of strain Ea321. In addition, HrpN was weakly translocated into plant cytoplasm. These results suggest that HrpN plays a significant role in the translocation of DspA/E, and HrpJ affects the translocation of DspA/E by affecting secretion or stability of HrpN. Taken together, these results explain the critical importance of HrpN and HrpJ to the development of fire blight.
Indirect carrier detection of canine haemophilia A using factor VIII microsatellite markers
Authors: Brooks M. B. , MacNguyen R. , Hall R. , Gupta R. , Booth J. G.
Published: April 2008

A panel of factor VIII microsatellite markers was developed for indirect carrier detection of canine haemophilia A (factor VIII deficiency). A total of 78 dogs, representing 14 different breed variants of haemophilia A, were genotyped at six intragenic factor VIII marker loci. The markers spanned approximately 110 kb and were located in the 5′ UTR of the factor VIII (F8) gene and within introns 6, 10, 12, 14 and 21. The observed heterozygosity (n = 39 females) for these markers was 0.675, 0.82, 0.868, 0.692, 0.473 and 0.775 respectively. The affected males of each breed variant had unique marker haplotypes. In addition, the marker haplotypes varied for two unrelated haemophilic Jack Russell terriers, compatible with independent mutation events causing haemophilia in different breeds and different families. A three‐marker panel (markers within introns 6, 10 and 21) was informative for 37 of the 39 females. The haemophilia‐associated haplotype was defined for six breed variants based on the genotypes of an affected male and a clear male sibling, with successful carrier detection of female siblings in each pedigree. Our results demonstrate an apparent allelic heterogeneity in canine haemophilia A; however, an indirect method based on a three‐marker panel is feasible to facilitate carrier detection and genetic counselling.
Role of pore pressure generation in sediment transport within half‐grabens
Authors: You Yao , Person Mark
Published: April 2008

Sediment transport and overpressure generation are coupled primary through the impact of effective stress on subsidence and compaction. Here, we use mathematical modeling to explore the interactions between groundwater flow and diffusion‐controlled sediment transport within alluvial basins. Because of lateral variation in permeability, proximal basin facies will have pore pressure close to hydrostatic levels while distal fine‐grained facies can reach near lithostatic levels. Lateral variation in pore pressure leads to differential compaction, which deforms basins in several ways. Differential compaction reduces basin size, bends isochron surfaces across the sand–clay interface, restricts basinward progradation of sand facies, and reduces the amplitude of oscillation in the lateral position of the sand–clay interface especially in the deepest part of the section even when temporal sediment supply are held constant. Overpressure generation was found to be sensitive to change in sediment supply in permeable basins (at least 10−17 m2 in our model). We found that during basin evolution, temporal variations in overpressure and sediment supply fluctuations are not necessarily in phase with each other, especially in tight (low permeability) basins (<10−17 m2 in our model).
Pharmacogenetics of aspirin resistance: a comprehensive systematic review
Authors: Goodman Timothy , Ferro Albert , Sharma Pankaj
Published: April 2008

WHAT IS ALREADY KNOWN ABOUT THIS SUBJECT

• A genetic basis for aspirin resistance has been postulated to exist.

• The individual studies published have been too small to allow reliable conclusions to be drawn.

• In order to clarify what, if any, genetic basis exists for aspirin resistance, we have undertaken, to our knowledge, the largest and most comprehensive systematic review conducted to‐date in this field.

WHAT THIS STUDY ADDS

• There is a significant association between the PlA1/A2 molecular variant and aspirin resistance in healthy subjects, with the effect diminishing in the presence of cardiovascular disease.

• However further studies are needed to confirm this.

• We argue that investigators now need to agree on a standard technique to measure and define aspirin resistance.

AIMSThe aim was to perform a systematic review of all candidate gene association studies in aspirin resistance.

METHODSElectronic databases were searched up until 1 December 2007 for all studies investigating any candidate gene for aspirin resistance in humans. Aspirin resistance was required to have been measured by a standardized laboratory technique to be included in the analysis.

RESULTSWithin 31 studies, 50 polymorphisms in 11 genes were investigated in 2834 subjects. The PlA1/A2 polymorphism in the GPIIIa platelet receptor was the most frequently investigated, with 19 studies in 1389 subjects. The PlA1/A2 variant was significantly associated with aspirin resistance when measured in healthy subjects [odds ratio (OR) 2.36, 95% confidence interval (CI) 1.24, 4.49;P = 0.009]. Combining genetic data from all studies (comprising both healthy subjects and those with cardiovascular disease) reduced the observed effect size (OR 1.14, 95% CI 0.84, 1.54;P = 0.40). Moreover, the observed effect of PlA1/A2 genotype varied depending on the methodology used for determining aspirin sensitivity/resistance. No significant association was found with aspirin resistance in four other investigated polymorphisms in the COX‐1, GPla, P2Y1 or P2Y12 genes.

CONCLUSIONSOur data support a genetic association between the PlA1/A2 molecular variant and aspirin resistance in healthy subjects, with the effect diminishing in the presence of cardiovascular disease. The laboratory methodology used influences the detection of aspirin resistance. However, as heterogeneity was significant and our results are based on a limited number of studies, further studies are required to confirm our findings.