Analysis of the mouse transcriptome based on functional annotation of 60,770 full-length cDNAs

Only a small proportion of the mouse genome is transcribed into mature messenger RNA transcripts. There is an international collaborative effort to identify all full-length mRNA transcripts from the mouse, and to ensure that each is represented in a physical collection of clones. Here we report the manual annotation of 60,770 full-length mouse complementary DNA sequences. These are clustered into 33,409 'transcriptional units', contributing 90.1% of a newly established mouse transcriptome database. Of these transcriptional units, 4,258 are new protein-coding and 11,665 are new non-coding messages, indicating that non-coding RNA is a major component of the transcriptome. 41% of all transcriptional units showed evidence of alternative splicing. In protein-coding transcripts, 79% of splice variations altered the protein product. Whole-transcriptome analyses resulted in the identification of 2,431 sense-antisense pairs. The present work, completely supported by physical clones, provides the most comprehensive survey of a mammalian transcriptome so far, and is a valuable resource for functional genomics.

Secondary structure of the third extracellular loop responsible for ligand selectivity of a mammalian gonadotropin-releasing hormone receptor

The extracellular loop 3 (ECL3) of the mammalian gonadotropin-releasing hormone receptor (GnRH-R) contains an acidic amino acid (Glu(301) in the mouse GnRH-R,) that confers agonist selectivity for Are in mammalian GnRH. It is proposed that a specific conformation of ECL3 is necessary to orientate the carboxyl side chain of the acidic residue for interaction with Arg(8) of GnRH, which is supported by decreased affinity for Arg(8) GnRH but not Gln(8) GnRH when an adjacent Pro is mutated to Ala. To probe the structural contribution of the loop domain to the proposed presentation of the carboxyl side chain, we synthesized a model peptide (CGPEMLNRVSEPGC) representing residues 293-302 of mouse ECL3, where Cys and Gly residues are added symmetrically at the N and C termini, respectively, allowing the introduction of a disulfide bridge to simulate the distances at which the ECL3 is tethered to the transmembrane domains 6 and 7 of the receptor. The ability of the ECL3 peptide to bind GnRH with low affinity was demonstrated by its inhibition of GnRH stimulation of inositol phosphate production in cells expressing the GnRH-R. The CD bands of the ECL3 peptides exhibited a superposition of predominantly unordered structure and partial contributions from beta-sheet structure. Likewise, the analysis of the amide I and amide III bands from micro-Raman and FT Raman experiments revealed mainly unordered conformations of the cyclic and of the linear peptide. NMR data demonstrated the presence of a beta-hairpin among an ensemble of largely disordered structures in the cyclic peptide. The location of the turn linking the two strands of the hairpin was assigned to the three central residues L-296, N-297, and R-298. A small population of structured species among an ensemble of predominantly random coil conformation suggests that the unliganded receptor represents a variety of structural conformers, some of which have the potential to make contacts with the ligand. We propose a mechanism of receptor activation whereby binding of the agonist to the inactive receptor state induces and stabilizes a particular structural state of the loop domain, leading to further conformational rearrangements across the transmembrane domain and signal propagating interaction with G proteins. Interaction of the Glu(301) of the receptor with Arg(8) of GnRH induces a folded configuration of the ligand. Our proposal thus suggests that conformational changes of both ligand and receptor result from this interaction.

Control of Aedes vectors of dengue in three provinces of Vietnam by use of Mesocyclops (Copepoda) and community-based methods validated by entomologic, clinical, and serological surveillance

We describe remarkable success in controlling dengue vectors, Aedes aegypti (L.) and Aedes albopictus (Skuse), in 6 communes with 11,675 households and 49,647 people in the northern provinces of Haiphong, Hung Yen, and Nam Dinh in Vietnam. The communes were selected for high-frequency use of large outdoor concrete tanks and wells. These were found to be the source of 49.6-98.4% of Ae. aegypti larvae, which were amenable to treatment with local Mesocyclops, mainly M. woutersi Van der Velde, M. aspericornis (Daday) and M. thermocyclopoides Harada. Knowledge, attitude, and practice surveys were performed to determine whether the communities viewed dengue and dengue hemorrhagic fever as a serious health threat; to determine their knowledge of the etiology, attitudes, and practices regarding control methods including Mesocyclops; and to determine their receptivity to various information methods. On the basis of the knowledge, attitude, and practice data, the community-based dengue control program comprised a system of local leaders, health volunteer teachers, and schoolchildren, supported by health professionals. Recycling of discards for economic gain was enhanced, where appropriate, and this, plus 37 clean-up campaigns, removed small containers unsuitable for Mesocyclops treatment. A previously successful eradication at Phan Boi village (Hung Yen province) was extended to 7 other villages forming Di Su commune (1,750 households) in the current study. Complete control was also achieved in Nghia Hiep (Hung Yen province) and in Xuan Phong (Nam Dinh province); control efficacy was greater than or equal to 99.7% in the other 3 communes (Lac Vien in Haiphong, Nghia Dong, and Xuan Kien in Nam Dinh). Although tanks and wells were the key container types of Ae. aegypti productivity, discarded materials were the source of 51% of the standing crop of Ae. albopictus. Aedes albopictus larvae were eliminated from the 3 Nam Dinh communes, and 86-98% control was achieved in the other 3 communes. Variable dengue attack rates made the clinical and serological comparison of control and untreated communes problematic, but these data indicate that clinical surveillance by itself is inadequate to monitor dengue transmission.

Conceptual modelling and ontology: Possibilities and pitfalls

Conceptual modelling is an activity undertaken during information systems development work to build a representation of selected semantics about some real-world domain. Ontological theories have been developed to account for the structure and behavior of the real world in general. In this paper, I discuss why ontological theories can be used to inform conceptual modelling research, practice, and pedagogy. I provide examples from my research to illustrate how a particular ontological theory has enabled me to improve my understanding of certain conceptual modelling practices and grammars. I describe, also, how some colleagues and I have used this theory to generate several counter-intuitive, sometimes surprising predictions about widely advocated conceptual modelling practices - predictions that subsequently were supported in empirical research we undertook. Finally, I discuss several possibilities and pitfalls I perceived to be associated with our using ontological theories to underpin research on conceptual modelling.

An investigation into commitment in non-Western industrial marketing relationships

This paper reports an investigation into the antecedents of commitment in non-Western industrial marketing relationships. The authors draw the antecedents from extant literature and posit that commitment is related to trust (integrity and reliability), communication quality, conflict, and similarity (social, ethnic, and economic). It is further argued that trust mediates the effects of communication, conflict, and similarity on commitment. As an extension, the authors examine the moderating effects of normative contracts (an implicit understanding of roles and responsibilities) on the construct interrelationships. The hypotheses are tested using data collected from approximately 150 industrial marketing relationships sampled from overseas Chinese firms. The results generally support the authors' framework; however, the mediating hypotheses are not supported. There is evidence of systematic differences in the effects of the studied antecedents on commitment and trust. Furthermore, a multigroup analysis provides evidence of significant moderating effects due to contracting mode. The study provides new insights into the theory and practice of industrial marketing. (C) 2003 Elsevier Science Inc. All rights reserved.

Distribution and abundance of dugongs in Moreton Bay, Queensland, Australia

Dugong abundances in Moreton Bay (south-east Queensland) were estimated during six bi- monthly aerial surveys throughout 1995. Sampling intensity ranged between 20 and 80% for different sampling zones within the Bay, with a mean intensity of 40.5%. Population estimates for dugongs were corrected for perception bias ( the proportion of animals visible in the transect that were missed by observers), and standardised for availability bias ( the proportion of animals that were invisible due to water turbidity) with survey and species-specific correction factors. Population estimates for dugongs in Moreton Bay ranged from 503 +/- 64 (s.e.) in July to 1019 +/- 166 in January. The highest uncorrected count was 857 dugongs in December. This is greater than previous population estimates, suggesting that either previous surveys have underestimated abundance and/or that this population may have increased through recruitment, immigration, or a combination of both. The high degree of variation in population estimates between surveys may be due to temporal differences in distribution and herding behaviour. In winter, dugongs were found in smaller herds and were dispersed over a wider area than in summer. The Eastern Banks region of the bay supported 80 - 98% of the dugong population at any one time. Within this region, there were several dugong 'hot spots' that were visited repeatedly by large herds. These 'hot spots' contained seagrass communities that were dominated by species that dugongs prefer to eat. The waters of Rous Channel, South Passage and nearby oceanic waters are also frequently inhabited by dugongs in the winter months. Dugongs in other parts of Moreton Bay were at much lower densities than on the Eastern Banks.

Direct visualization of Ras proteins in spatially distinct cell surface microdomains

Localization of signaling complexes to specific micro-domains coordinates signal transduction at the plasma membrane. Using immunogold electron microscopy of plasma membrane sheets coupled with spatial point pattern analysis, we have visualized morphologically featureless microdomains including lipid rafts, in situ and at high resolution. We find that an inner-plasma membrane lipid raft marker displays cholesterol-dependent clustering in microdomains with a mean diameter of 44 nm that occupy 35% of the cell surface. Cross-linking an outer-leaflet raft protein results in the redistribution of inner leaflet rafts, but they retain their modular structure. Analysis of Ras microlocalization shows that inactive H-ras is distributed between lipid rafts and a cholesterol-independent micro-domain. Conversely, activated H-ras and K-ras reside predominantly in nonoverlapping, cholesterol-independent microdomains. Galectin-1 stabilizes the association of activated H-ras with these nonraft microdomains, whereas K-ras clustering is supported by farnesylation, but not geranylgeranylation. These results illustrate that the inner plasma membrane comprises a complex mosaic of discrete microdomains. Differential spatial localization within this framework can likely account for the distinct signal outputs from the highly homologous Ras proteins.

Phylogeny and classification of the Digenea (Platyhelminthes : Trematoda)

Complete small subunit ribosomal RNA gene (ssrDNA) and partial (D1-D3) large subunit ribosomal RNA gene (lsrDNA) sequences were used to estimate the phylogeny of the Digenea via maximum parsimony and Bayesian inference. Here we contribute 80 new ssrDNA and 124 new lsrDNA sequences. Fully complementary data sets of the two genes were assembled from newly generated and previously published sequences and comprised 163 digenean taxa representing 77 nominal families and seven aspidogastrean outgroup taxa representing three families. Analyses were conducted on the genes independently as well as combined and separate analyses including only the higher plagiorchiidan taxa were performed using a reduced-taxon alignment including additional characters that could not be otherwise unambiguously aligned. The combined data analyses yielded the most strongly supported results and differences between the two methods of analysis were primarily in their degree of resolution. The Bayesian analysis including all taxa and characters, and incorporating a model of nucleotide substitution (general-time-reversible with among-site rate heterogeneity), was considered the best estimate of the phylogeny and was used to evaluate their classification and evolution. In broad terms, the Digenea forms a dichotomy that is split between a lineage leading to the Brachylaimoidea, Diplostomoidea and Schistosomatoidea (collectively the Diplostomida nomen novum (nom. nov.)) and the remainder of the Digenea (the Plagiorchiida), in which the Bivesiculata nom. nov. and Transversotremata nom. nov. form the two most basal lineages, followed by the Hemiurata. The remainder of the Plagiorchiida forms a large number of independent lineages leading to the crown clade Xiphidiata nom. nov. that comprises the Allocreadioidea, Gorgoderoidea, Microphalloidea and Plagiorchioidea, which are united by the presence of a penetrating stylet in their cercariae. Although a majority of families and to a lesser degree, superfamilies are supported as currently defined, the traditional divisions of the Echinostomida, Plagiorchiida and Strigeida were found to comprise non-natural assemblages. Therefore, the membership of established higher taxa are emended, new taxa erected and a revised, phylogenetically based classification proposed and discussed in light of ontogeny, morphology and taxonomic history. (C) 2003 Australian Society for Parasitology Inc. Published by Elsevier Science Ltd. All rights reserved.

Acoustic analysis of speech through a hearing aid: perceptual effects of changes with two-channel compression

Compression amplification significantly alters the acoustic speech signal in comparison to linear amplification. The central hypothesis of the present study was that the compression settings of a two-channel aid that best preserved the acoustic properties of speech compared to linear amplification would yield the best perceptual results, and that the compression settings that most altered the acoustic properties of speech compared to linear would yield significantly poorer speech perception. On the basis of initial acoustic analysis of the test stimuli recorded through a hearing aid, two different compression amplification settings were chosen for the perceptual study. Participants were 74 adults with mild to moderate sensorineural hearing impairment. Overall, the speech perception results supported the hypothesis. A further aim of the study was to determine if variation in participants' speech perception with compression amplification (compared to linear amplification) could be explained by the individual characteristics of age, degree of loss, dynamic range, temporal resolution, and frequency selectivity; however, no significant relationships were found.

A unit cell waveguide model of a reflectarray formed by microstrip patches and slots

An equivalent unit cell waveguide approach (WGA) is described to obtain reflection coefficient phase curves for designing a microstrip patch reflectarray supported by a ground plane with periodic apertures or slots. Based on the presented theory, a computer algorithm for determining the reflection coefficient of a plane wave normally incident on a multi-layer structure of patches and apertures is developed. The validity of the developed algorithm is verified by comparing the obtained results with those published in the literature and the ones generated by Agilent High Frequency Structure Simulator (HFSS). A good agreement in all the presented examples is obtained, proving that the developed theory and computer algorithm can be an effective tool for designing multi-layer microstrip reflectarrays with a periodically perforated ground plane. (C) 2003 Wiley Periodicals, Inc.

ADH genotype does not modify the effects of alcohol on high-density lipoprotein

Background: Alcohol consumption has beneficial effects on mortality which are mainly due to reduction in cardiovascular disease. These are believed to be due, at least in part, to the increase in plasma high-density lipoprotein (HDL) which is associated with alcohol consumption. It has been proposed that ADH3 genotype modifies the relationships between alcohol intake and cardiovascular disease by altering the HDL response to alcohol. The aim of this paper was to test for effects of ADH2 and ADH3 genotypes on the response of HDL components to habitual alcohol consumption. Methods: Adult male and female subjects were genotyped for ADH2 and ADH3; and plasma HDL cholesterol, apolipoprotein A-I, and apolipoprotein A-II were measured. Nine hundred one subjects had both ADH2 and ADH3 genotypes and HDL cholesterol results, while 753 had both genotypes and all three lipid results. The effect of alcohol intake on the three measured HDL components, and a factor score derived from them, was estimated for each of the ADH2 and ADH3 genotype groups. Results: All the measured components of HDL increased with increasing alcohol consumption over the range of intakes studied, 0-4 drinks per day. There were no significant interactions between alcohol consumption and ADH2 or ADH3 genotypes. Conclusions: The concept that alcohol dehydrogenase genotype and alcohol metabolic rate modify the effects of alcohol on plasma HDL concentration is not supported by our results.

Towards the on-farm conservation of the assassin bug Pristhesancus plagipennis (Walker) (Hemiptera : Reduviidae) during winter using crop plants as refuges

Habitat instability associated with seasonal crop succession in broad-acre farming systems presents a problem for the conservation and utilisation of beneficial insects in annual field crops. The present paper describes two experiments used to measure the potential of seven plant species to be utilised as winter refuges to support and conserve the predatory bug Pristhesancus plagipennis (Walker). In the first experiment, replicated plots of canola (Brassica napus ), red salvia (Salvia coccinea ), niger (Guizotia abyssinica ), linseed (Linum usitatissimum ), lupins (Lupinus angustifolius ), and lucerne (Medicago falcata ) were planted in a randomized experiment during Autumn 1998. Upon crop establishment, adults and nymphs of P. plagipennis were released into treatment plots and their numbers were assessed, along with those of their potential prey, throughout the ensuing winter months. Post-release sampling suggested that canola and niger retained a proportion of adult P. plagipennis , while niger, lucerne and canola retained some nymphs. The other plant species failed to support P. plagipennis nymphs and adults postrelease. In the second experiment, niger was compared with two lines of sunflower (Helianthus annus ). Both sunflower lines harboured significantly higher (P < 0.05) densities of P. plagipennis nymphs than did niger. The more successful refuge treatments (sunflower, niger and canola) had an abundance of yellow flowers that were attractive to pollinating insects, which served as supplementary prey on which P. plagipennis were observed to feed. Sunflower and niger also supported high densities of the prey insect Creontiades dilutus (Stal) and provided protective leafy canopies which supplied shelter during the winter months. The potential and limitations for using each plant species as a winter refuge to retain P. plagipennis during winter are discussed.

Variation of bulk properties of anaerobic granules with wastewater type

Development of a granular sludge with high strength, high biological activity and a narrow settling distribution is necessary for optimal operation of high-rate upflow anaerobic treatment systems. Several studies have compared granules produced from different wastewaters but these have largely been from laboratory-fed reactors or compared granules from full-scale reactors fed similar wastewater types. Though two authors have commented on the inferiority of granules produced by a protein-based feed, the properties of these granules have not been characterised. In this paper, granules from full-scale reactors treating fruit and vegetable cannery effluent, two brewery effluents and a pig abattoir (slaughterhouse) were compared in terms of basic composition, size distribution, density, settling velocity, shear strength, and EPS content. The results supported previous qualitative observations by other researchers that indicate granule properties depend more on wastewater type rather than reactor design or operating conditions such as pre-acidification level. The cannery-fed granules bad excellent shear strength, settling distribution and density. Granules from the two brewery-fed reactors had statistically the same bulk properties, which were still acceptable for upflow applications. The protein-grown granule had poor strength and settling velocity. (C) 2001 Elsevier Science Ltd. All rights reserved.

Development of a quality use of medicines coding system to rate clinical pharmacists' medication review recommendations

Objective: To develop a 'quality use of medicines' coding system for the assessment of pharmacists' medication reviews and to apply it to an appropriate cohort. Method: A 'quality use of medicines' coding system was developed based on findings in the literature. These codes were then applied to 216 (111 intervention, 105 control) veterans' medication profiles by an independent clinical pharmacist who was supported by a clinical pharmacologist with the aim to assess the appropriateness of pharmacy interventions. The profiles were provided for veterans participating in a randomised, controlled trial in private hospitals evaluating the effect of medication review and discharge counselling. The reliability of the coding was tested by two independent clinical pharmacists in a random sample of 23 veterans from the study population. Main outcome measure: Interrater reliability was assessed by applying Cohen's kappa score on aggregated codes. Results: The coding system based on the literature consisted of 19 codes. The results from the three clinical pharmacists suggested that the original coding system had two major problems: (a) a lack of discrimination for certain recommendations e. g. adverse drug reactions, toxicity and mortality may be seen as variations in degree of a single effect and (b) certain codes e. g. essential therapy were in low prevalence. The interrater reliability for an aggregation of all codes into positive, negative and clinically non-significant codes ranged from 0.49-0.58 (good to fair). The interrater reliability increased to 0.72-0.79 (excellent) when all negative codes were excluded. Analysis of the sample of 216 profiles showed that the most prevalent recommendations from the clinical pharmacists were a positive impact in reducing adverse responses (31.9%), an improvement in good clinical pharmacy practice (25.5%) and a positive impact in reducing drug toxicity (11.1%). Most medications were assigned the clinically non-significant code (96.6%). In fact, the interventions led to a statistically significant difference in pharmacist recommendations in the categories; adverse response, toxicity and good clinical pharmacy practice measured by the quality use of medicine coding system. Conclusion: It was possible to use the quality use of medicine coding system to rate the quality and potential health impact of pharmacists' medication reviews, and the system did pick up differences between intervention and control patients. The interrater reliability for the summarised coding system was fair, but a larger sample of medication regimens is needed to assess the non-summarised quality use of medicines coding system.

Identification of a novel protein kinase mediating Akt survival signaling to the ATM protein

We identified a novel human AMP-activated protein kinase (AMPK) family member, designated ARK5, encoding 661 amino acids with an estimated molecular mass of 74 kDa. The putative amino acid sequence reveals 47, 45.8, 42.4, and 55% homology to AMPK-alpha1, AMPK-alpha2, MELK and SNARE respectively, suggesting that it is a new member of the AMPK family. It has a putative Akt phosphorylation motif at amino acids 595600, and Ser(600) was found to be phosphorylated by active Akt resulting in the activation of kinase activity toward the SAMS peptide, a consensus AMPK substrate. During nutrient starvation, ARK5 supported the survival of cells in an Akt-dependent manner. In addition, we also demonstrated that ARK5, when activated by Akt, phosphorylated the ATM protein that is mutated in the human genetic disorder ataxia-telangiectasia and also induced the phosphorylation of p53. On the basis of our current findings, we propose that a novel AMPK family member, ARK5, is the tumor cell survival factor activated by Akt and acts as an ATM kinase under the conditions of nutrient starvation.