bluuebunny/crossref_metadata_2025 huggingface dataset:
Desempenho de frangos de corte nas fases de crescimento e final alimentados com rações contendo soja integral extrusada em diferentes temperaturas
Authors: Oliveira Francisco Nairson de , Costa Fernando Guilherme Perazzo , Silva José Humberto Vilar da , Brandão Patrícia Araújo , Amarante Júnior Valene da Silva , Nascimento Germano Augusto Jerônimo do , Barros Leilane Rocha
Published: January 2006

O efeito da temperatura de extrusão da soja integral (SIE) sobre o desempenho e as características de carcaça de frangos de corte em diferentes fases de criação foi avaliado neste experimento. As rações, isonutritivas, foram formuladas para atender às exigências nutricionais das aves nas fases de crescimento (22 a 35 dias) e final (36 a 45 dias de idade). Os tratamentos experimentais consistiram de quatro temperaturas de extrusão da soja integral (125, 130, 135 e 140ºC), com nove repetições de 30 aves. As variáveis avaliadas foram consumo de ração (CR), ganho de peso (GP), conversão alimentar (CA) e características da carcaça (rendimentos de perna e de peito e gordura abdominal). Em ambas as fases, os tratamentos não apresentaram efeito significativo sobre as variáveis estudadas. Os resultados indicam que o processamento da soja integral com extrusão à temperatura de 125 a 140ºC pode ser utilizado visando à utilização do extrusado em dietas para frangos de corte.
The MGED Ontology: a resource for semantics-based description of microarray experiments
Authors: Whetzel Patricia L. , Parkinson Helen , Causton Helen C. , Fan Liju , Fostel Jennifer , Fragoso Gilberto , Game Laurence , Heiskanen Mervi , Morrison Norman , Rocca-Serra Philippe , Sansone Susanna-Assunta , Taylor Chris , White Joseph , Stoeckert Christian J.
Published: January 2006

Motivation: The generation of large amounts of microarray data and the need to share these data bring challenges for both data management and annotation and highlights the need for standards. MIAME specifies the minimum information needed to describe a microarray experiment and the Microarray Gene Expression Object Model (MAGE-OM) and resulting MAGE-ML provide a mechanism to standardize data representation for data exchange, however a common terminology for data annotation is needed to support these standards.

Results: Here we describe the MGED Ontology (MO) developed by the Ontology Working Group of the Microarray Gene Expression Data (MGED) Society. The MO provides terms for annotating all aspects of a microarray experiment from the design of the experiment and array layout, through to the preparation of the biological sample and the protocols used to hybridize the RNA and analyze the data. The MO was developed to provide terms for annotating experiments in line with the MIAME guidelines, i.e. to provide the semantics to describe a microarray experiment according to the concepts specified in MIAME. The MO does not attempt to incorporate terms from existing ontologies, e.g. those that deal with anatomical parts or developmental stages terms, but provides a framework to reference terms in other ontologies and therefore facilitates the use of ontologies in microarray data annotation.

Availability: The MGED Ontology version.1.2.0 is available as a file in both DAML and OWL formats at . Release notes and annotation examples are provided. The MO is also provided via the NCICB's Enterprise Vocabulary System ().

Contact:  Stoeckrt@pcbi.upenn.edu

Supplementary information: Supplementary data are available at Bioinformatics online.
To transplant or not? The importance of psychosocial and behavioural factors before lung transplantation
Authors: Dobbels F , Verleden G , Dupont L , Vanhaecke J , De Geest S
Published: January 2006

The gratifying results of lung transplantation in terms of survival and quality of life stimulate the referral of an ever-increasing number of patients with end-stage lung disease. This in turn compounds the organ shortage, which is the limiting factor in the transplantation rate. In the absence of good alternative treatment modalities, an evidence-based pretransplant screening process is a prerequisite to detennine which patients will benefit most from transplantation. Within this evidence-based screening process, medical selection criteria are well established. There is a growing awareness that psychosocial and behavioural factors may determine outcome after transplantation as well. This paper reviews the available evidence for psychosocial and behavioural factors in the screening process for lung transplantation. The relation of various factors with post-transplant outcome was explored. Psychosocial characteristics before transplantation consist of 1) anxiety and depression, 2) personality disorders, 3) neurocognitive problems, and 4) lack of social support. Pretransplant behavioural factors include 1) noncompliance with medication, 2) alcohol abuse or dependence, 3) smoking, 4) noncompliance with dietary guidelines, and 5) noncompliance with monitoring of vital parameters and infections. It appears that the lack of rigorous studies limit the feasibility of an evidence-based screening process. Prospective studies are crucial to this further investigation of the relationship between psychosocial and behavioural determinants before transplantation and outcomes after transplantation, in terms of compliance, morbidity, and mortality. Identification of modifiable risk factors for poor outcome before transplantation is a first step in developing interventions.
Addio, Adieu, Adios: A Farewell to Arms in Three Romance Languages
Authors: Azevedo Milton Mariano
Published: January 2006

The strong component of orality that pervades A Farewell to Arms creates an intensely heteroglossic text in which a variety of voices are continuously modulated. This feature challenges translators to find lexical and syntactic equivalents capable of conveying the colloquial tone of the dialogues and the contrasts among the characters' voices. Taking into account five categories of translation issues—lexical congruence, modes of address, off-color language, literary dialect, and narrative perspective—this article analyzes how that challenge is met in translations into French, Italian and Spanish.
"He Felt the Change So That It Hurt Him All Through": Sodomy and Transvestic Hallucination in Late Hemingway
Authors: Eby Carl P
Published: January 2006

This article calls attention to a pattern of heterosexual sodomy in four late Hemingway texts to explain the psychological mechanics of the gender transformations experienced by the male protagonists of these novels. The article suggests that these protagonists experience a sort of bodily hallucination, for which sodomy functions as a catalyst, and which amounts to nothing less than a sort of "psychotic flash" within the context of an otherwise "perverse" psychology. The paper then uses this framework to briefly link gender to trauma theory in Hemingway studies and theorize why eroticism is so much more overt and transgressive in late, as opposed to early, Hemingway.
Reaction of cyanoacetylene HCCCN(XΣ+1) with ground-state carbon atoms C(P3) in cold molecular clouds
Authors: Li H. Y. , Cheng W. C. , Liu Y. L. , Sun B. J. , Huang C. Y. , Chen K. T. , Tang M. S. , Kaiser R. I. , Chang A. H. H.
Published: January 2006

The reaction of the simplest cyanopolyyne, cyanoacetylene [HCCCN(XΣ+1)], with ground-state atomic carbon C(P3) is investigated theoretically to explore the probable routes for the depletion of the famed interstellar molecule HCCCN, and the formation of carbon-nitrogen-bearing species in extraterrestrial environments particularly of ultralow temperature. Six collision complexes (c1–c6) without entrance barrier as a result of the carbon atom addition to the π systems of HCCCN are located. The optimized geometries and harmonic frequencies of the intermediates, transition states, and products along the isomerization and dissociation pathways of each collision complex are obtained by utilizing the unrestricted B3YLP∕6-311G(d,p) level of theory, and the corresponding CCSD(T)/cc-pVTZ energies are calculated. Subsequently, with the facilitation of Rice-Ramsperger-Kassel-Marcus (RRKM) and variational RRKM rate constants at collision energy of 0–10kcal∕mol, the most probable paths for the titled reaction are determined, and the product yields are estimated. Five collision complexes (c1–c3, c5, and c6) are predicted to give the same products, a chained CCCCN (p2)+H, via the linear and most stable intermediate, HCCCCN (i2), while collision complex c4 is likely to dissociate back to C+HCCCN. The study suggests that this class of reaction is an important route to the destruction of cyanoacetylene and cyanopolyynes in general, and to the synthesis of linear carbon-chained nitriles at the temperature as low as 10K to be incorporated in future chemical models of interstellar clouds.
Isotopic replacement in ionic systems: The He2+4+He3→He3He+4+He4 reaction
Authors: Bodo Enrico , Lara Manuel , Gianturco Franco A.
Published: January 2006

Full quantum dynamics calculations have been carried out for the ionic reaction He2+4+He3 and state-to-state reactive probabilities have been obtained using both time-dependent and time-independent approaches. An accurate ab initio potential-energy surface has been employed for the present quantum dynamics and the two sets of results are shown to be in agreement with each other. The results for zero total angular momentum suggest a marked presence of atom exchange (isotopic replacement) reaction with probabilities as high as 60%. The reaction probabilities are only weakly dependent on the initial vibrational state of the reactants, while they are slightly more sensitive to the degree of rotational excitation. A brief discussion of the results for selected higher total angular momentum values is also presented, while the l-shifting approximation [S. K. Gray et al., Phys. Chem. Chem. Phys. 1, 1141 (1999)] has been used to provide estimates of the total reaction rates for the title process. Such rates are found to be large enough to possibly become experimentally accessible.
The chemical nature of the C–H⊕⋯X− (X=Cl or Br) interaction in imidazolium halide ionic liquids
Authors: Wang Yong , Li Haoran , Han Shijun
Published: January 2006

The C–H⋯X (X=Cl or Br) interaction is traditionally characterized as a relatively weak interaction. However, this interaction becomes very strong in the imidazolium-based halide ionic liquids [J. Phys. Chem. 123, 174501 (2005)]. This strong interaction had been attributed to the electrostatic interaction between the imidazolium cation and the halide anion. In this paper, the chemical nature of the C–H⊕⋯Cl− and C–H⊕⋯Br− interactions is investigated by atoms in molecules (AIM) and natural bond orbital (NBO) analyses. The AIM calculations indicate that in the EmimX complexes, the C–H⊕⋯Cl− and C–H⊕⋯Br− interactions have some covalent character, especially the C–H⊕⋯Cl− interaction. Mulliken, ChelpG charge, and natural bond orbital population analyses for these two kinds of interactions indicate that the charge transfer is important in the interaction of the cation with the anion. In addition, the NBO analysis demonstrated that the stabilization energy is due to an n→σC–H• orbital interaction. However, in the Emim2X and Emim3X complexes, the calculated results suggested a dominant electrostatic character for the C–H⊕⋯Cl− and C–H⊕⋯Br− interactions.
Plant community responses to experimental warming across the tundra biome
Authors: Walker Marilyn D. , Wahren C. Henrik , Hollister Robert D. , Henry Greg H. R. , Ahlquist Lorraine E. , Alatalo Juha M. , Bret-Harte M. Syndonia , Calef Monika P. , Callaghan Terry V. , Carroll Amy B. , Epstein Howard E. , Jónsdóttir Ingibjörg S. , Klein Julia A. , Magnússon Borgþór , Molau Ulf , Oberbauer Steven F. , Rewa Steven P. , Robinson Clare H. , Shaver Gaius R. , Suding Katharine N. , Thompson Catharine C. , Tolvanen Anne , Totland Ørjan , Turner P. Lee , Tweedie Craig E. , Webber Patrick J. , Wookey Philip A.
Published: January 2006

Recent observations of changes in some tundra ecosystems appear to be responses to a warming climate. Several experimental studies have shown that tundra plants and ecosystems can respond strongly to environmental change, including warming; however, most studies were limited to a single location and were of short duration and based on a variety of experimental designs. In addition, comparisons among studies are difficult because a variety of techniques have been used to achieve experimental warming and different measurements have been used to assess responses. We used metaanalysis on plant community measurements from standardized warming experiments at 11 locations across the tundra biome involved in the International Tundra Experiment. The passive warming treatment increased plant-level air temperature by 1-3°C, which is in the range of predicted and observed warming for tundra regions. Responses were rapid and detected in whole plant communities after only two growing seasons. Overall, warming increased height and cover of deciduous shrubs and graminoids, decreased cover of mosses and lichens, and decreased species diversity and evenness. These results predict that warming will cause a decline in biodiversity across a wide variety of tundra, at least in the short term. They also provide rigorous experimental evidence that recently observed increases in shrub cover in many tundra regions are in response to climate warming. These changes have important implications for processes and interactions within tundra ecosystems and between tundra and the atmosphere.
Oil market power and United States national security
Authors: Stern Roger
Published: January 2006

It is widely believed that an oil weapon could impose scarcity upon the United States. Impending resource exhaustion is thought to exacerbate this threat. However, threat seems implausible when we consider strategic deficits of prospective weapon users and the improbability of impending resource exhaustion. Here, we explore a hypothesis relating oil to national security under a different assumption, abundance. We suggest that an oil cartel exerts market power to keep abundance at bay, commanding monopoly rents [or wealth transfers (wt)] that underwrite security threats. We then compare security threats attributed to the oil weapon to those that may arise from market power. We first reexamine whether oil is abundant or scarce by reviewing current development data, then we estimate a competitive price for oil. From this, we derivewt2004collections by Persian Gulf states ≈ $132-178 × 109. We find thatwtand the behavior of states collecting it interact to actuate security threats. Threats underwritten bywtare (i) the potential for emergence of a Persian Gulf superpower and (ii) terrorism. It is therefore oil market power, not oil per se, that actuates threats. We also describe a paradox in the relation of market power to the United States' defense doctrine of force projection to preempt a Gulf superpower. Because the superpower threat derives fromwt, force alone cannot preempt it. A further paradox is that because foreign policy is premised on oil weapon fear, market power is appeased. Threats thereby grow unimpeded.
Reconciling the working strokes of a single head of skeletal muscle myosin estimated from laser-trap experiments and crystal structures
Authors: Sleep John , Lewalle Alexandre , Smith David
Published: January 2006

Myosin generates force by a rotation of its lever arm. Crystal structures of myosin II indicate an unloaded working stroke of 10–12 nm, a range confirmed by recent x-ray interference experiments. However, when an actin filament, held between two weakly, optically trapped beads is made to interact with a single head of skeletal myosin, the bead displacements have often been reported as having a mean value of 5–6 nm, a value that is commonly interpreted as the working stroke. In general, the observed displacement is not expected to be equal to the working stroke because the kinetics of the stroke is necessarily strain-dependent: this effect biases the frequency of binding events to different actin sites so that displacements smaller than the working stroke are preferentially selected. Our analysis is tailored to current trap experiments, in which the time resolution is insufficient to detect prerigor states. If the preceding transitions are in equilibrium, the mean displacement is zero, contrary to observations in the presence of ATP. However, under ATP-cycling conditions, we find that the mean displacement is deflated to 0.3–0.7 of the true working stroke, depending on the equilibrium constant of the stroke and the rate at which the first myosin product state can detach from actin. The primary working stroke of processive myosin motors as measured by optical trapping is similarly uncertain.
Ancient polymorphism reveals unidirectional breeding system shifts
Authors: Igic Boris , Bohs Lynn , Kohn Joshua R.
Published: January 2006

Loss of complex characters is thought to be irreversible (Dollo's law). However, hypotheses of irreversible evolution are remarkably difficult to test, especially when character transitions are frequent. In such cases, inference of ancestral states, in the absence of fossil evidence, is uncertain and represents the single greatest constraint for reconstructing the evolutionary history of characters. Breeding system character transitions are of particular interest because they affect the amount and distribution of genetic variation within species. Transitions from obligate outcrossing to partial or predominant self-fertilization are thought to represent one of the most common trends in flowering plants. We use the unique molecular genetic properties (manifested as deep persistent polymorphisms) of the locus that enforces outcrossing to demonstrate that its loss is irreversible in the plant family Solanaceae. We argue that current phylogenetic methods of reconstruction are potentially inadequate in cases where ancestral state information is inferred by using only the phylogeny and the distribution of character states in extant taxa. This study shows in a statistical framework that a particular character transition is irreversible, consistent with Dollo's law.
Foamy virus vector integration sites in normal human cells
Authors: Trobridge Grant D. , Miller Daniel G. , Jacobs Michael A. , Allen James M. , Kiem Hans-Peter , Kaul Rajinder , Russell David W.
Published: January 2006

Foamy viruses (FVs) or spumaviruses are retroviruses that have been developed as vectors, but their integration patterns have not been described. We have performed a large-scale analysis of FV integration sites in unselected human fibroblasts (
n
= 1,008) and human CD34
+
hematopoietic cells (
n
= 1,821) by using a bacterial shuttle vector and a comparable analysis of lentiviral vector integration sites in CD34
+
cells (
n
= 1,331). FV vectors had a distinct integration profile relative to other types of retroviruses. They did not integrate preferentially within genes, despite a modest preference for integration near transcription start sites and a significant preference for CpG islands. The genomewide distribution of FV vector proviruses was nonrandom, with both clusters and gaps. Transcriptional profiling showed that gene expression had little influence on integration site selection. Our findings suggest that FV vectors may have desirable integration properties for gene therapy applications.
Antagonistic regulation of ROMK by long and kidney-specific WNK1 isoforms
Authors: Lazrak Ahmed , Liu Zhen , Huang Chou-Long
Published: January 2006

WNK kinases are serine-threonine kinases with an atypical placement of the catalytic lysine. Intronic deletions with increased expression of a ubiquitous long
WNK1
transcript cause pseudohypoaldosteronism type 2 (PHA II), characterized by hypertension and hyperkalemia. Here, we report that long WNK1 inhibited ROMK1 by stimulating its endocytosis. Inhibition of ROMK by long WNK1 was synergistic with, but not dependent on, WNK4. A smaller transcript of
WNK1
lacking the N-terminal 1-437 amino acids is expressed highly in the kidney. Whether expression of the KS-WNK1 (kidney-specific, KS) is altered in PHA II is not known. We found that KS-WNK1 did not inhibit ROMK1 but reversed the inhibition of ROMK1 caused by long WNK1. Consistent with the lack of inhibition by KS-WNK1, we found that amino acids 1-491 of the long WNK1 were sufficient for inhibiting ROMK. Dietary K
+
restriction decreases ROMK abundance in the renal cortical-collecting ducts by stimulating endocytosis, an adaptative response important for conservation of K
+
during K
+
deficiency. We found that K
+
restriction in rats increased whole-kidney transcript of long WNK1 while decreasing that of KS-WNK1. Thus, KS-WNK1 is a physiological antagonist of long WNK1. Hyperkalemia in PHA II patients with PHA II mutations may be caused, at least partially, by increased expression of long WNK1 with or without decreased expression of KS-WNK1.
Identification of a novel putative mitogen-activated kinase cascade on human chromosome 21 by computational approaches
Authors: Pellegrini-Calace Marialuisa , Tramontano Anna
Published: January 2006

Summary: Down syndrome (DS) is the most frequent form of mental retardation and is caused by chromosome 21 (HSA21) trisomy. Despite the number of known genes involved in DS and its high therapeutic interest, biological mechanisms leading to the DS phenotype are not fully clear. We present a functional hypothesis based on fold recognition and hidden Markov model techniques for four HSA21 genes located in the DS Candidate Region (DSCR). More specifically, we propose that they are members of a novel mitogen-activated protein kinase pathway with DYRK1A, SNF1LK and RIPK4 gene products being elements of the kinase cascade and the DSCR3 acting as structural scaffold for their interaction. This hypothesis finds support in various biochemical studies concerning the biological behavior and features of the involved HSA21 proteins. Our analysis calls for specifically designed experiments to validate our prediction and establish its relevance in terms of therapeutic approaches to the disease.

Contact:  anna.tramontano@uniroma1.it

Suplementary information: Supplementary data are available at Bioinformatics online.
Genetic test bed for feature selection
Authors: Choudhary Ashish , Brun Marcel , Hua Jianping , Lowey James , Suh Ed , Dougherty Edward R.
Published: January 2006

Motivation: Given a large set of potential features, such as the set of all gene-expression values from a microarray, it is necessary to find a small subset with which to classify. The task of finding an optimal feature set of a given size is inherently combinatoric because to assure optimality all feature sets of a given size must be checked. Thus, numerous suboptimal feature-selection algorithms have been proposed. There are strong impediments to evaluate feature-selection algorithms using real data when data are limited, a common situation in genetic classification. The difficulty is compound. First, there are no class-conditional distributions from which to draw data points, only a single small labeled sample. Second, there are no test data with which to estimate the feature-set errors, and one must depend on a training-data-based error estimator. Finally, there is no optimal feature set with which to compare the feature sets found by the algorithms.

Results: This paper describes a genetic test bed for the evaluation of feature-selection algorithms. It begins with a large biological feature-label dataset that is used as an empirical distribution and, using massively parallel computation, finds the top feature sets of various sizes based on a given sample size and classification rule. The user can draw random samples from the data, apply a proposed algorithm, and evaluate the proficiency of the proposed algorithm via three different measures (code provided). A key feature of the test bed is that, once a dataset is input, a single command creates the entire test bed relative to the dataset. The particular dataset used for the first version of the test bed comes from a microarray-based classification study that analyzes a large number of microarrays, prepared with RNA from breast tumor samples from each of 295 patients.

Availability: The software and supplementary material are available at

Contact:  edward@ece.tamu.edu
Automatic extension of Gene Ontology with flexible identification of candidate terms
Authors: Lee Jin-Bok , Kim Jung-jae , Park Jong C.
Published: January 2006

Motivation: Gene Ontology (GO) has been manually developed to provide a controlled vocabulary for gene product attributes. It continues to evolve with new concepts that are compiled mostly from existing concepts in a compositional way. If we consider the relatively slow growth rate of GO in the face of the fast accumulation of the biological data, it is much desirable to provide an automatic means for predicting new concepts from the existing ones.

Results: We present a novel method that predicts more detailed concepts by utilizing syntactic relations among the existing concepts. We propose a validation measure for the automatically predicted concepts by matching the concepts to biomedical articles. We also suggest how to find a suitable direction for the extension of a constantly growing ontology such as GO.

Availability:  

Contact:  park@nlp.kaist.ac.kr

Supplementary information: Supplementary materials are available at Bioinformatics online.
Chloroplastic NAD(P)H Dehydrogenase in Tobacco Leaves Functions in Alleviation of Oxidative Damage Caused by Temperature Stress
Authors: Wang Peng , Duan Wei , Takabayashi Atsushi , Endo Tsuyoshi , Shikanai Toshiharu , Ye Ji-Yu , Mi Hualing
Published: January 2006

In this study, the function of the NAD(P)H dehydrogenase (NDH)-dependent pathway in suppressing the accumulation of reactive oxygen species in chloroplasts was investigated. Hydrogen peroxide accumulated in the leaves of tobacco (Nicotiana tabacum) defective in ndhC-ndhK-ndhJ (ΔndhCKJ) at 42°C and 4°C, and in that of wild-type leaves at 4°C. The maximum quantum efficiency of PSII decreased to a similar extent in both strains at 42°C, while it decreased more evidently in ΔndhCKJ at 4°C. The parameters linked to CO2 assimilation, such as the photochemical efficiency of PSII, the decrease of nonphotochemical quenching following the initial rise, and the photosynthetic O2 evolution, were inhibited more significantly in ΔndhCKJ than in wild type at 42°C and were seriously inhibited in both strains at 4°C. While cyclic electron flow around PSI mediated by NDH was remarkably enhanced at 42°C and suppressed at 4°C. The proton gradient across the thylakoid membranes and light-dependent ATP synthesis were higher in wild type than in ΔndhCKJ at either 25°C or 42°C, but were barely formed at 4°C. Based on these results, we suggest that cyclic photophosphorylation via the NDH pathway might play an important role in regulation of CO2 assimilation under heat-stressed condition but is less important under chilling-stressed condition, thus optimizing the photosynthetic electron transport and reducing the generation of reactive oxygen species.
The Arabidopsis Zinc Finger-Homeodomain Genes Encode Proteins with Unique Biochemical Properties That Are Coordinately Expressed during Floral Development
Authors: Tan Queenie K.-G. , Irish Vivian F.
Published: January 2006

Arabidopsis (Arabidopsis thaliana) contains approximately 100 homeobox genes, many of which have been shown to play critical roles in various developmental processes. Here we characterize the zinc finger-homeodomain (ZF-HD) subfamily of homeobox genes, consisting of 14 members in Arabidopsis. We demonstrate that the HDs of the ZF-HD proteins share some similarities with other known HDs in Arabidopsis, but they contain distinct features that cluster them as a unique class of plant HD-containing proteins. We have carried out mutational analyses to show that the noncanonical residues present in the HDs of this family of proteins are important for function. Yeast (Saccharomyces cerevisiae) two-hybrid matrix analyses of the ZF-HD proteins reveal that these proteins both homo- and heterodimerize, which may contribute to greater selectivity in DNA binding. These assays also show that most of these proteins do not contain an intrinsic activation domain, suggesting that interactions with other factors are required for transcriptional activation. We also show that the family members are all expressed predominantly or exclusively in floral tissue, indicating a likely regulatory role during floral development. Furthermore, we have identified loss-of-function mutations for six of these genes that individually show no obvious phenotype, supporting the idea that the encoded proteins have common roles in floral development. Based on these results, we propose the ZF-HD gene family encodes a group of transcriptional regulators with unique biochemical activities that play overlapping regulatory roles in Arabidopsis floral development.
Control of Floral Meristem Determinacy in Petunia by MADS-Box Transcription Factors
Authors: Ferrario Silvia , Shchennikova Anna V. , Franken John , Immink Richard G.H. , Angenent Gerco C.
Published: January 2006

The shoot apical meristem (SAM), a small group of undifferentiated dividing cells, is responsible for the continuous growth of plants. Several genes have been identified that control the development and maintenance of the SAM. Among these, WUSCHEL (WUS) from Arabidopsis (Arabidopsis thaliana) is thought to be required for maintenance of a stem cell pool in the SAM. The MADS-box gene AGAMOUS, in combination with an unknown factor, has been proposed as a possible negative regulator of WUS, leading to the termination of meristematic activity within the floral meristem. Transgenic petunia (Petunia hybrida) plants were produced in which the E-type and D-type MADS-box genes FLORAL BINDING PROTEIN2 (FBP2) and FBP11, respectively, are simultaneously overexpressed. These plants show an early arrest in development at the cotyledon stage. Molecular analysis of these transgenic plants revealed a possible combined action of FBP2 and FBP11 in repressing the petunia WUS homolog, TERMINATOR. Furthermore, the ectopic up-regulation of the C-type and D-type homeotic genes FBP6 and FBP7, respectively, suggests that they may also participate in a complex, which causes the determinacy in transgenic plants. These data support the model that a transcription factor complex consisting of C-, D-, and E-type MADS-box proteins controls the stem cell population in the floral meristem.
Extracellular ATP Induces the Accumulation of Superoxide via NADPH Oxidases in Arabidopsis
Authors: Song Charlotte J. , Steinebrunner Iris , Wang Xuanzhi , Stout Stephen C. , Roux Stanley J.
Published: January 2006

Extracellular ATP can serve as a signaling agent in animal cells, and, as suggested by recent reports, may also do so in plant cells. In animal cells it induces the production of reactive oxygen species through the mediation of NADPH oxidase. Similarly, here we report that in leaves of Arabidopsis (Arabidopsis thaliana), applied ATP, but not AMP or phosphate, induces the accumulation of superoxide (O2  −) in a biphasic, dose-dependent manner, with a threshold at 500 nm ATP. This effect did not require ATP hydrolysis for it was mimicked by ATPγS. ATP also induced increased levels of Arabidopsis respiratory burst oxidase homolog D (AtrbohD) mRNA, but ATP-treated plants that had disrupted AtrbohD and AtrbohF genes did not accumulate O2  −, indicating that NADPH oxidases are responsible for the induced O2  − accumulation. Inhibitors of mammalian P2-type ATP receptors abolished ATP-induced O2  − production, suggesting that the ATP effects may be mediated through P2-like receptors in plants. Cytosolic Ca2+ and calmodulin are likely to help transduce the ATP responses, as they do in animal cells, because a Ca2+ channel blocker, a Ca2+ chelator, and calmodulin antagonist all reduced ATP-induced O2  − accumulation. Furthermore, ATP treatment enhanced the expression of genes that are induced by wounds and other stresses. The ATP measured at wound sites averaged 40 μ  m, well above the level needed to induce O2  − accumulation and gene expression changes. Transgenic plants overexpressing an apyrase gene had reduced O2  − production in response to applied ATP and wounding. Together, these data suggest a possible role for extracellular ATP as a signal potentially in wound and stress responses.
Forward Genetic Analysis of the Circadian Clock Separates the Multiple Functions of ZEITLUPE
Authors: Kevei Éva , Gyula Péter , Hall Anthony , Kozma-Bognár László , Kim Woe-Yeon , Eriksson Maria E. , Tóth Réka , Hanano Shigeru , Fehér Balázs , Southern Megan M. , Bastow Ruth M. , Viczián András , Hibberd Victoria , Davis Seth J. , Somers David E. , Nagy Ferenc , Millar Andrew J.
Published: January 2006

The circadian system of Arabidopsis (Arabidopsis thaliana) includes feedback loops of gene regulation that generate 24-h oscillations. Components of these loops remain to be identified; none of the known components is completely understood, including ZEITLUPE (ZTL), a gene implicated in regulated protein degradation. ztl mutations affect both circadian and developmental responses to red light, possibly through ZTL interaction with PHYTOCHROME B (PHYB). We conducted a large-scale genetic screen that identified additional clock-affecting loci. Other mutants recovered include 11 new ztl alleles encompassing mutations in each of the ZTL protein domains. Each mutation lengthened the circadian period, even in dark-grown seedlings entrained to temperature cycles. A mutation of the LIGHT, OXYGEN, VOLTAGE (LOV)/Period-ARNT-Sim (PAS) domain was unique in retaining wild-type responses to red light both for the circadian period and for control of hypocotyl elongation. This uncoupling of ztl phenotypes indicates that interactions of ZTL protein with multiple factors must be disrupted to generate the full ztl mutant phenotype. Protein interaction assays showed that the ztl mutant phenotypes were not fully explained by impaired interactions with previously described partner proteins Arabidopsis S-phase kinase-related protein 1, TIMING OF CAB EXPRESSION 1, and PHYB. Interaction with PHYB was unaffected by mutation of any ZTL domain. Mutation of the kelch repeat domain affected protein binding at both the LOV/PAS and the F-box domains, indicating that interaction among ZTL domains leads to the strong phenotypes of kelch mutations. Forward genetics continues to provide insight regarding both known and newly discovered components of the circadian system, although current approaches have saturated mutations at some loci.
Identification and Characterization of the Arabidopsis Orthologs of Nuclear Transport Factor 2, the Nuclear Import Factor of Ran
Authors: Zhao Qiao , Leung Sara , Corbett Anita H. , Meier Iris
Published: January 2006

Ran is a multifunctional small GTPase that is involved in nucleocytoplasmic transport, mitotic spindle assembly, and nuclear envelope formation. Nuclear import of Ran relies on a small RanGDP-binding protein, Nuclear Transport Factor 2 (NTF2). Three proteins are expressed in Arabidopsis (Arabidopsis thaliana) that show significant sequence similarity to human and yeast (Saccharomyces cerevisiae) NTF2. Here, we demonstrate that two of them, AtNTF2a and AtNTF2b, can functionally replace the essential NTF2 gene in yeast. Consistent with this finding, both AtNTF2a and AtNTF2b interact with yeast and Arabidopsis Ran. The third NTF2-related protein, AtNTL, does not functionally replace NTF2 in yeast. Similar to yeast NTF2-green fluorescent protein (GFP), AtNTF2a-GFP and AtNTF2b-GFP accumulate at the nuclear rim. The AtNTF2a E38K and E91K mutants, which fail to bind Ran, are not functional in yeast, indicating conservation of the requirement for these key amino acids in plants and yeast. AtNTF2a overexpression, but not AtNTF2aE38K overexpression, blocks nuclear import of a plant transcription factor in Nicotiana benthamiana leaves, indicating that excess AtNTF2a disrupts nuclear import in a Ran-binding-dependent manner. On the basis of these results, we propose that AtNTF2a and AtNTF2b function in Ran import in Arabidopsis and that nuclear import of Ran is functionally conserved in plants.
Stimulation of CD8+T Cells following Diphtheria Toxin-Mediated Antigen Delivery into Dendritic Cells
Authors: Shaw Christine A. , Starnbach Michael N.
Published: January 2006

Recognition and clearance of many intracellular pathogens requires the activation and subsequent effector functions of CD8+T lymphocytes. To stimulate CD8+T cells by immunization, the target antigens must be delivered into the cytosol of host cells. There they can be processed into peptides and presented in the context of major histocompatibility complex class I molecules to antigen-specific CD8+T cells. One method of delivering antigens into the cytosol is to fuse them to modified bacterial toxins that are able to enter mammalian cells. The expression pattern of the toxin receptors in the host will determine the cell population that the toxin fusion protein targets and will thus restrict antigen-specific T-cell recognition to the same population. In this study we describe the development and characterization of a diphtheria toxin (DT)-based antigen delivery system. Using CD11c-DTR transgenic mice that express the DT receptor in dendritic cells (DC), this system allows for targeted delivery of CD8+T-cell antigen to DC. We show that antigen-specific CD8+T cells proliferate in CD11c-DTR mice following immunization with catalytically inactive DT-antigen fusion proteins. We also show that a toxin-based system that restricts antigen delivery to DC results in more robust antigen-specific CD8+T-cell proliferation than a toxin-based system that does not restrict delivery to a particular cell type. These results have implications for vaccine design, and they suggest that use of a toxin-based vector to target antigen to DC may be an effective way to induce a CD8+T-cell response.
Human Monoclonal Anti-Protective Antigen Antibody Completely Protects Rabbits and Is Synergistic with Ciprofloxacin in Protecting Mice and Guinea Pigs against Inhalation Anthrax
Authors: Peterson Johnny W. , Comer Jason E. , Noffsinger David M. , Wenglikowski Autumn , Walberg Kristin G. , Chatuev Bagram M. , Chopra Ashok K. , Stanberry Lawrence R. , Kang Angray S. , Scholz Wolfgang W. , Sircar Jagadish
Published: January 2006

Prevention of inhalation anthrax requires early and extended antibiotic therapy, and therefore, alternative treatment strategies are needed. We investigated whether a human monoclonal antibody (AVP-21D9) to protective antigen (PA) would protect mice, guinea pigs, and rabbits against anthrax. Control animals challenged with
Bacillus anthracis
Ames spores by the intranasal route died within 3 to 7 days. AVP-21D9 alone provided minimal protection against anthrax in the murine model, but its efficacy was notably better in guinea pigs. When Swiss-Webster mice, challenged with five 50% lethal doses (LD
50
s) of anthrax spores, were given a single 16.7-mg/kg of body weight AVP-21D9 antibody dose combined with ciprofloxacin (30 mg/kg/day for 6 days) 24 h after challenge, 100% of the mice were protected for more than 30 days, while ciprofloxacin or AVP-21D9 alone showed minimal protection. Similarly, when AVP-21D9 antibody (10 to 50 mg/kg) was combined with a low, nonprotective dose of ciprofloxacin (3.7 mg/kg/day) and administered to guinea pigs for 6 days, synergistic protection against anthrax was observed. In contrast, a single dose of AVP-21D9 antibody (1, 5, 10, or 20 mg/kg) but not 0.2 mg/kg alone completely protected rabbits against challenge with 100 LD
50
s of
B. anthracis
Ames spores, and 100% of the rabbits survived rechallenge. Further, administration of AVP-21D9 (10 mg/kg) to rabbits at 0, 6, and 12 h after challenge with anthrax spores resulted in 100% survival; however, delay of antibody treatment by 24 and 48 h reduced survival to 80% and 60%, respectively. Serological analysis of sera from various surviving animals 30 days postprimary infection showed development of a species-specific PA enzyme-linked immunosorbent assay antibody titer that correlated with protection against reinfection. Taken together, the effectiveness of human anti-PA antibody alone or in combination with low ciprofloxacin levels may provide the basis for an improved strategy for prophylaxis or treatment following inhalation anthrax infection.
Identification of Proteins Secreted via
Vibrio parahaemolyticus
Type III Secretion System 1
Authors: Ono Takahiro , Park Kwon-Sam , Ueta Mayumi , Iida Tetsuya , Honda Takeshi
Published: January 2006

Vibrio parahaemolyticus
, a gram-negative marine bacterium, is an important pathogen causing food-borne gastroenteritis or septicemia. Recent genome sequencing of the RIMD2210633 strain (a Kanagawa phenomenon-positive clinical isolate of serotype O3:K6) revealed that the strain has two sets of gene clusters that encode the type III secretion system (TTSS) apparatus. The first cluster, TTSS1, is located on the large chromosome, and the second, TTSS2, is on the small chromosome. Previously, we reported that TTSS1 is involved in the cytotoxicity of the RIMD2210633 strain against HeLa cells. Here, we analyzed proteins secreted via the TTSS apparatus encoded by TTSS1 by using two-dimensional gel electrophoresis and identified the proteins encoded by genes VP1680, VP1686, and VPA450. To investigate the roles of those secreted proteins, we constructed and analyzed a series of deletion mutants. Flow cytometry analysis using fluorescence-activated cell sorting with fluorescein isothiocyanate-labeled annexin V demonstrated that the TTSS1-dependent cell death was by apoptosis. The cytotoxicity to HeLa cells was related to one of the newly identified secreted proteins encoded by VP1680. Adenylate cyclase fusion protein studies proved that the newly identified secreted proteins were translocated into HeLa cells. Thus, these appear to be the TTSS effector proteins in
V. parahaemolyticus
.
TheBordetella bronchisepticaType III Secretion System Inhibits Gamma Interferon Production That Is Required for Efficient Antibody-Mediated Bacterial Clearance
Authors: Pilione Mylisa R. , Harvill Eric T.
Published: January 2006

Several species of pathogenic microorganisms have developed strategies to survive and persist in vital organs which are normally maintained as sterile by the generation of strong immune responses. Here, we report an immunomodulation involving theBordetella bronchisepticatype III secretion system (TTSS) which contributes to bacterial survival in the lower respiratory tract of the host. The prolonged persistence ofB. bronchisepticathat was observed in gamma interferon (IFN-γ)−/−mice indicates that the efficient clearance of bacteria from the lower respiratory tract requires not only B cells and antibodies but also IFN-γ production. Our data also suggest that interleukin-10 (IL-10)-producing splenocytes are generated early during infection and that IL-10 inhibits IFN-γ-producing cells and delays the clearance ofB. bronchisepticafrom the lungs. The TTSS ofB. bronchisepticainhibits the generation of IFN-γ-producing splenocytes and is required for long-term bacterial persistence in the lower respiratory tract in wild-type mice. This suggests that a mechanism involving the modulation of IFN-γ production by the TTSS facilitatesB. bronchisepticasurvival in the lower respiratory tract.
Role for Inducible Costimulator in Control ofSalmonella entericaSerovar Typhimurium Infection in Mice
Authors: Vidric Mariana , Bladt Anna Tafuri , Dianzani Umberto , Watts Tania H.
Published: January 2006

Inducible costimulator (ICOS) is expressed on activated T cells and plays a key role in sustaining and enhancing the effector function of CD4 T cells. Given the function of this molecule in sustaining T-cell responses, we reasoned that ICOS might play an important role in a prolonged infection model, such asSalmonellainfection of mice. To test this hypothesis, wild-type (WT) and ICOS-deficient (ICOS−/−) mice were infected systemically with aSalmonella entericaserovar Typhimurium strain expressing the chicken ovalbumin gene (Salmonella-OVA). ICOS−/−mice exhibited greater splenomegaly than WT mice and showed delayed bacterial clearance. The acquired immune response in this model was slow to develop. Maximal T-cell responses toSalmonella-OVA were detected at 3 weeks postinfection in both WT and ICOS−/−mice. CD4 T-cell-dependent gamma interferon production and a class switch to immunoglobulin G2a were severely reduced in ICOS−/−mice. ICOS−/−mice also exhibited a substantial defect in antigen-specific CD8 T-cell responses. In vitro, the effect of anti-ICOS on CD8 T-cell division was greater when CD8 T cells rather than CD4 T cells expressed ICOS, suggesting that the in vivo effects of ICOS on CD8 T cells could be direct. Taken together, these studies show that ICOS plays a critical role in control ofSalmonellainfection in mice, with effects on antibody, Th1, and CD8 T-cell responses.
Construction and Phase I Clinical Evaluation of the Safety and Immunogenicity of a Candidate Enterotoxigenic
Escherichia coli
Vaccine Strain Expressing Colonization Factor Antigen CFA/I
Authors: Turner Arthur K. , Beavis Juliet C. , Stephens Jonathan C. , Greenwood Judith , Gewert Cornelia , Thomas Nicola , Deary Alison , Casula Gabriella , Daley Alexandra , Kelly Paul , Randall Roger , Darsley Michael J.
Published: January 2006

Oral delivery of toxin-negative derivatives of enterotoxigenic
Escherichia coli
(ETEC) that express colonization factor antigens (CFA) with deletions of the
aroC
,
ompC
,
ompF
, and toxin genes may be an effective approach to vaccination against ETEC-associated diarrhea. We describe the creation and characterization of an attenuated CFA/I-expressing ETEC vaccine candidate, ACAM2010, from a virulent isolate in which the heat-stable enterotoxin (ST) and CFA/I genes were closely linked and on the same virulence plasmid as the enteroaggregative
E. coli
heat-stable toxin (EAST1) gene. A new suicide vector (pJCB12) was constructed and used to delete the ST and EAST1 genes and to introduce defined deletion mutations into the
aroC
,
ompC
, and
ompF
chromosomal genes. A phase I trial, consisting of an open-label dose escalation phase in 18 adult outpatient volunteers followed by a placebo-controlled double-blind phase in an additional 31 volunteers, was conducted. The vaccine was administered in two formulations, fresh culture and frozen suspension. These were both well tolerated, with no evidence of significant adverse events related to vaccination. Immunoglobulin A (IgA) and IgG antibody-secreting cells specific for CFA/I were assayed by ELISPOT. Positive responses (greater than twofold increase) were seen in 27 of 37 (73%) subjects who received the highest dose level of vaccine (nominally 5 × 10
9
CFU). Twenty-nine of these volunteers were secreting culturable vaccine organisms at day 3 following vaccination; five were still positive on day 7, with a single isolation on day 13. This live attenuated bacterial vaccine is safe and immunogenic in healthy adult volunteers.