The Aedes aegypti larval transcriptome: a comparative perspective with emphasis on trypsins and the domain structure of peritrophins

The genome sequence of Aedes aegypti was recently reported. A significant amount of Expressed Sequence Tags (ESTs) were sequenced to aid in the gene prediction process. In the present work we describe an integrated analysis of the genomic and EST data, focusing on genes with preferential expression in larvae (LG), adults (AG) and in both stages (SG). A total of 913 genes (5.4% of the transcript complement) are LG, including ion transporters and cuticle proteins that are important for ion homeostasis and defense. From a starting set of 245 genes encoding the trypsin domain, we identified 66 putative LG, AG, and SG trypsins by manual curation. Phylogenetic analyses showed that AG trypsins are divergent from their larval counterparts (LG), grouping with blood-induced trypsins from Anopheles gambiae and Simulium vittatum. These results support the hypothesis that blood-feeding arose only once, in the ancestral Culicomorpha. Peritrophins are proteins that interlock chitin fibrils to form the peritrophic membrane (PM) that compartmentalizes the food in the midgut. These proteins are recognized by having chitin-binding domains with 6 conserved Cys and may also present mucin-like domains (regions expected to be highly O-glycosylated). PM may be formed by a ring of cells (type 2, seen in Ae. aegypti larvae and Drosophila melanogaster) or by most midgut cells (type 1, found in Ae. aegypti adult and Tribolium castaneum). LG and D. melanogaster peritrophins have more complex domain structures than AG and T. castaneum peritrophins. Furthermore, mucin-like domains of peritrophins from T. castaneum (feeding on rough food) are lengthier than those of adult Ae. aegypti (blood-feeding). This suggests, for the first time, that type 1 and type 2 PM may have variable molecular architectures determined by different peritrophins and/or ancillary proteins, which may be partly modulated by diet.

Conformational Properties of Angiotensin II and Its Active and Inactive TOAC-Labeled Analogs in the Presence of Micelles. Electron Paramagnetic Resonance, Fluorescence, and Circular Dichroism Studies

The interaction between angiotensin II (AII, DRVYIHPF) and its analogs carrying 2,2,6,6-tetramethylpiperidine-1-oxyl-4-amino-4-carboxylic acid (TOAC) and detergents-negatively charged sodium dodecyl sulfate (SDS) and zwitterionic N-hexadecyl-N,N-dimethyl-3-ammonio-1-propanesulfonate (HPS)-was examined by means of EPR, CD, and fluorescence. EPR spectra of partially active TOAC(1)-AII and inactive TOAC(3)-AII in aqueous solution indicated fast tumbling, the freedom of motion being greater at the N-terminus. Line broadening occurred upon interaction with micelles. Below SDS critical micelle concentration, broader lines indicated complex formation with tighter molecular packing than in micelles. Small changes in hyperfine splittings evinced TOAC location at the micelle-water interface. The interaction with anionic micelles was more effective than with zwitterionic micelles. Peptide-micelle interaction caused fluorescence increase. The TOAC-promoted intramolecular fluorescence quenching was more, pronounced for TOAC(3)-AII because of the proximity between the nitroxide and Tyr(4). CD spectra showed that although both AII and TOAC(1)-AII presented flexible conformations in water, TOAC(3)-AII displayed conformational restriction because of the TOAC-imposed bend (Schreier et al., Biopolymers 2004, 74, 389). In HPS, conformational changes were observed for the labeled peptides at neutral and basic pH. In SDS, all peptides underwent pH-dependent conformational changes. Although the spectra suggested similar folds for All and TOAC(1)-AII, different conformations were acquired by TOAC(3)-AII. The membrane environment has been hypothesized to shift conformational equilibria so as to stabilize the receptor-bound conformation of ligands. The fact that TOAC(3)-AII is unable to acquire conformations similar to those of native AII and partially active TOAC(1)-AII is probably the explanation for its lack of biological activity. (C) 2009 Wiley Periodicals, Inc. Biopolymers (Pept Sci) 92: 525-537, 2009.

Protein Adsorption onto Polyelectrolyte Layers: Effects of Protein Hydrophobicity and Charge Anisotropy

Ellipsometry was used to investigate the influence of ionic strength (I) and pH on the adsorption of bovine serum albumin (BSA) or beta-lactoglobulin (BLG) onto preabsorbed layers of two polycations: poly(diallyldimethylammonium chloride) (PDADMAC) or poly(4-vinylpyridine bromide) quaternized with linear aliphatic chains of two (QPVP-C2) or five (QPVP-C5) carbons. Comparisons among results for the three polycations reveal hydrophobic interactions, while comparisons between BSA and BLG-proteins of very similar isoelectric points (pI)-indicate the importance of protein charge anisotropy. At pH close to pI, the ionic strength dependence of the adsorbed amount of protein (Gamma) displayed maxima in the range 10 < I < 25 mM corresponding to Debye lengths close to the protein radii. Visualization of protein charge by Delphi suggested that these ionic strength conditions corresponded to suppression of long-range repulsion between polycations and protein positive domains, without diminution of short-range attraction between polycation segments and locally negative protein domains, in a manner similar to the behavior of PE-protein complexes in solution.(1-4) This description was consistent with the disappearance of the maxima at pH either above or below pI. In the former case, Gamma values decrease exponentially with I(1/2), due to screening of attractions, while in the latter case adsorption of both proteins decreased at low I due to strong repulsion. Close to or below pI both proteins adsorbed more strongly onto QPVP-C5 than onto QPVP-C2 or PDADMAC due to hydrophobic interactions with the longer alkyl group. Above pI, the adsorption was more pronounced with PDADMAC because these chains may assume more loosely bound layers due to lower linear charge density.

Adsorption of 4-aminopyridine on Co and Ag electrodes probed by SERS

The adsorption of 4-aminopyridine (4-AP) on Co and Ag electrodes in acid or alkaline solutions of KCl and KI electrolyte salts were monitored by the Surface-enhanced Raman Spectroscopy (SERS) technique. The SERS intensity for the Ag electrode was in 2 orders of magnitude higher than for the Co electrode, due to the enhancement of the Raman cross-section on Ag by the surface-plasmon excitation. In acidic chloride medium (pH 4), the SERS results for Ag electrodes indicate that the protonated form of 4-AP (4-APH(+)) adsorbs in the potential range of -0.1 to -0.6 V (Ag broken vertical bar AgCl broken vertical bar KCl sat) through hydrogen-bonding between 4-APH(+) and Cl(-) adsorbed on the electrode surface: at more negative potentials the neutral form 4-AP is the predominant adsorbed species. For Co electrode in the same medium, only bands due to neutral 4-AP were observed in the spectra at -0.8 and -0.9 V. For more negative potentials bands assigned to both 4-AP and 4-AP surface complex are observed, with the lasts being enhanced, as the potentials are turned more negative. In alkaline chloride medium (pH 13), for less negative potentials the bands assigned to free 4-AP were observed in the spectra of both Ag and Co surfaces. For more negative potentials, only bands assigned to the 4-AP surface complex were observed. For 0.1 mol L(-1) KI acidic or alkaline solutions, bands assigned to 4-AP and 4-APH(+) were observed in a wider potential range than in chloride solutions. An adsorption scheme of 4-AP on Ag and Co is proposed for acidic and alkaline solutions. (C) 2010 Elsevier B.V. All rights reserved.

Expression, Purification, Bioactivity, and Partial Characterization of a Recombinant Human Bone Morphogenetic Protein-7 Produced in Human 293T Cells

Bone morphogenetic protein-7 (BMP-7) is a secreted multifunctional growth factor of the TGF-beta superfamily, which is predominantly known for its osteoinductive properties and emerging potential for treatment of kidney diseases. The mature 34-38 kDa disulfide-linked homodimer protein plays a key role in the differentiation of mesenchymal cells into bone and cartilage. In this study, the full-length sequence of hBMP-7 was amplified and, then, cloned, expressed, and purified from the conditioned medium of 293T cells stably transfected with a lentiviral vector. The mature protein dimer form was properly secreted and recognized by anti-BMP-7 antibodies, and the protein was shown to be glycosilated by treatment with exoglycosidase, followed by western blotting. Moreover, the activity of the purified protein was demonstrated both in vitro, by alkaline phosphatase activity in C2C12 cells, and in vivo by induction of ectopic bone formation in Balb/c Nude mice after 21 days, respectively. This recombinant protein platform may be very useful for expression of different human cytokines and other proteins for medical applications.

Sdo1p, the yeast orthologue of Shwachman-Bodian-Diamond syndrome protein, binds RNA and interacts with nuclear rRNA-processing factors

The Shwachman-Bodian-Diamond syndrome protein (SBDS) is a member of a highly conserved protein family of not well understood function, with putative orthologues found in different organisms ranging from Archaea, yeast and plants to vertebrate animals. The yeast orthologue of SBDS, Sdo1p, has been previously identified in association with the 60S ribosomal subunit and is proposed to participate in ribosomal recycling. Here we show that Sdo1p interacts with nucleolar rRNA processing factors and ribosomal proteins, indicating that it might bind the pre-60S complex and remain associated with it during processing and transport to the cytoplasm. Corroborating the protein interaction data, Sdo1p localizes to the nucleus and cytoplasm and co-immunoprecipitates precursors of 60S and 40S subunits, as well as the mature rRNAs. Sdo1p binds RNA directly, suggesting that it may associate with the ribosomal subunits also through RNA interaction. Copyright (C) 2009 John Wiley & Sons, Ltd.

Genomic organization and transcription analysis of the 195-bp satellite DNA in Trypanosoma cruzi

The 195-bp satellite DNA is the most abundant Trypanosoma cruzi repetitive sequence. Here we show by RNA blotting and RT-PCR that 195 SAT is intensely transcribed. We observed a positive correlation between the level of satellite RNA and the abundance of the satellite copies in the genome of T cruzi strains and that the satellite expression is not developmentally regulated. By analyzing CL Brener individual reads, we estimated that 195 SAT corresponds to approximately 5% of the CL Brener genome. 195 SAT elements were found in only 37 annotated contigs, indicating that a large number of satellite copies were not incorporated into the assembled data. The assembled satellite units are distributed in non-syntenic regions with Trypanosoma brucei and Leishmania major genomes, enriched with surface proteins, retroelements, RHS and hypothetical proteins. Satellite repeats were not observed in annotated subtelomeric regions. We report that 12 satellite sequences are truncated by the retroelement VIPER. (C) 2008 Elsevier B.V. All rights reserved.

The functions of Japanese Sound Symbolic Words in Different Types of Texts and Their Translation

Though sound symbolic words (onomatopoeia and mimetic words, or giongo and gitaigo in Japanese) exist in other languages, it would not be so easy to compare them to those in Japanese. This is because unlike in Japanese, in many other languages (here we see English and Spanish) sound symbolic words do not have distinctive forms that separate them immediately from the rest of categories of words. In Japanese, a sound symbolic word has a radical (that is based on the elaborated Japanese sound symbolic system), and often a suffix that shows subtle nuance. Together they give the word a distinctive form that differentiates it from other categories of words, though its grammatical functions could vary, especially in the case of mimetic words (gitaigo). Without such an obvious feature, in other languages, it would not be always easy to separate sound symbolic words from the rest. These expressions are extremely common and used in almost all types of text in Japanese, but their elaborated sound symbolic system and possibly their various grammatical functions are making giongo and gitaigo one of the most difficult challenges for the foreign students and translators. Studying the translation of these expressions into other languages might give some indication related to the comparison of Japanese sound symbolic words and those in other languages. Though sound symbolic words are present in many types of texts in Japanese, their functions in traditional forms of text (letters only) and manga (Japanese comics)are different and they should be treated separately. For example, in traditional types of text such as novels, the vast majority of the sound symbolic words used are mimetic words (gitaigo) and most of them are used as adverbs, whereas in manga, the majority of the sound symbolic words used (excluding those appear within the speech bubbles) are onomatopoeias (giongo) and often used on their own (i.e. not as a part of a sentence). Naturally, the techniques used to translate these expressions in the above two types of documents differ greatly. The presentation will focus on i) grammatical functions of Japanese sound symbolic words in traditional types of texts (novels/poems) and in manga works, and ii) whether their features and functions are maintained (i.e. whether they are translated as sound symbolic words) when translated into other languages (English and Spanish). The latter point should be related to a comparison of sound symbolic words in Japanese and other languages, which will be also discussed.

Capability outcomes from educational and ICT capability inputs : an analysis of ICT use in informal education in Kenya

In the field of Information and Communication Technologies for Development (ICT4D) ICT use in education is well studied. Education is often seen as a pre-requisite for development and ICTs are believed to aid in education, e.g. to make it more accessible and to increase its quality. In this paper we study the access and use of ICT in a study circle (SC) education program in the south coast of Kenya. The study is qualitative reporting results based on interviews and observations with SC participants, government officers and SC coordinators and teachers. The study builds on the capability approach perspective of development where individuals’ opportunities and ability to live a life that they value are focused. The aim of the study is to investigate the capability outcomes enabled through the capability inputs access and use of ICT in education as well as the factors that enabled and/or restricted the outcomes. Findings show that many opportunities have been enabled such as an increase in the ability to generate an income, learning benefits, community development and basic human development (e.g. literacy and self-confidence). However, conversion factors such as a poorly developed infrastructure and poor IT literacy prevent many of the individuals from taking full advantage of the ICT and the opportunities it enables. 

O behaviorismo radical de B. F. Skinner e sua aplicabilidade socialmente relevante

o presente trabalho tem como objetivo a avaliação da capacidade do behaviorismo radical skinneriano'para fundamentar aplicações práticas dotadas de efetiva relevância humana e social, assim definidas em' função do caráter politicamente progressista e democratizante que se lhes possa imprimir. Essa avaliação é desenvolvida através de dois distintos, mas interrelacionados, procedimentos acadêmicos: uma argumentação discursiva baseada em pesquisa bibliográfica pertinente, que se complementapor um estudo empírico de campo a nível exploratório. A apreciação discursiva do behaviorismo radical é inicialmente conduzida sob uma perspectiva histórico-acadêmica, cujo foco contínuo de referência é o próprio agente individual basicamente responsável pelo gradativo processo de constituição desse sistema científico - B. F. Skinner. Em uma da etapa, o behaviorismo radical vem a ser discutido deforma mais impessoalizada, contra os sucessivos panos de fundo da filosofia da ciência, das relações sociais humanas e da ética implicada nas intervenções práticas sobre tais relações. O estudo exploratório, que então-se segue, consiste no julgamento da utilidade potencial de um instrumento de educação política popular, elaborado previamente a partir dos princípios behavioristas radicais básicos e seus desdobramentos psicossociológicos, e destinado a uma clientela sócio-economicamente desprivi1egiada, constituída de indivíduos adultos, de ambos os sexos, com escolaridade 'em torno do 19 grau completo. A avaliação dessa Cartilha de Contracontole Social através de entrevistas semi-estruturadas e um breve questionário objetivo, por quatorze lnformantes - qualificados em termos de sua 1iderança e/ou militância em associações de moradores de bairros e de favelas, sindicatos de trabalhadores, núcleos comunitários re1igiosos, entidades assistenciais autônomas e associações de servidores em organizações de trabalho - vem a apresentar, em linhas gerais, resultados muito positivos. A iniciativa é considerada extremamente séria e responsável; e o conteúdo da Carti1ha, bastante compatível com os objetivos desses movimentos populares, assim como muito útil para dotar as suas atividades de uma maior eficácia social. Evidencia-se, não obstante, uma restrição generalizada quanto à possibilidade de compreensão do texto por parte da população-alvo; o que deve, portanto, ensejar sua reformu1ação para o emprego em futuros projetos de prosseguimento dessa linha de pesquisa. Conclui-se, enfim, que a tese em seu todo parece apta a contribuir, se suficientemente divulgada, para a promoção de um maior interesse pelo behaviorismo de Skinner no âmbito acadêmico do Rio de Janeiro, bem como para a produção de um exercício mais eficaz de contracontro1e sobre as atuais práticas aversivas e/ou exploratórias de controle social.

Patentes como instrumento de aumento de dependência tecnológica e econômica: estudo da indústria farmacêutica

A dependência brasileira, na indústria farmacêutica/farmoquÍmica, em relação aos interesses e capitais estrangeiros é extremamente elevada e incompatível com as dimensões de mercado que o país representa e com os interesses nacionais. Por outro lado, é praticamente, nulo o trabalho de pesquisa básica no setor, aqui realizado, seja por empresas nacionais ou, principalmente, estrangeiras. Comumente,alega-se que a falta de pesquisa no setor' é decorrência do não reconhecimento de patentes de produtos e respectivos processos de produção. Este trabalho procura demonstrar que a causa básica da dependência tecnológica e, consequentemente, econômica no setor não tem relação significativa com a legislação que não reconhece tais patentes. Ao contrário, o sistema de patentes pode conduzir a um maior grau de dependência para os países em desenvolvimento. Além disto, constata-se que, apesar de o Brasil ser o principal país sob o processo de desenvolvimento dependente, no setor farmacêutico/farmacoquímico nos encontramos ainda num estágio de total dependência, sem desenvolvimento significativo, porque os capitais estrangeiros são para cá dirigidos com o fim de simplesmente explorarem o mercado local, sem o intuito de transferir "know-how" e equipar tecnologicamente a base industrial a qui existente. Tais aspectos, abordados à luz dos conceitos da Teoria Geral do Imperialismo e da Teoria do Desenvolvimento Dependente, são analisados conjuntamente com a evolução histórica do setor no Brasil e com os fatores determinantes dessa atual situação de dependência econômica e tecnológica.

Public debt management, monetary policy and financial institutions

Despite the large size of the Brazilian debt market, as well the large diversity of its bonds, the picture that emerges is of a market that has not yet completed its transition from the role it performed during the megainflation years, namely that of providing a liquid asset that provided positive real returns. This unfinished transition is currently placing the market under severe stress, as fears of a possible default from the next administration grow larger. This paper analyzes several aspects pertaining to the management of the domestic public debt. The causes for the extremely large and fast growth ofthe domestic public debt during the seven-year period that President Cardoso are discussed in Section 2. Section 3 computes Value at Risk and Cash Flow at Risk measures for the domestic public debt. The rollover risk is introduced in a mean-variance framework in Section 4. Section 5 discusses a few issues pertaining to the overlap between debt management and monetary policy. Finally, Section 6 wraps up with policy discussion and policy recommendations.

Análise da Expressão de APE1 após Estresse Oxidativo Induzido em Células Proficientes e Deficientes na Via de Reparo por Excisão de Nucleotídeos

Riboflavin is a vitamin very important in aerobic organisms, as a precursor of many coenzymes involved in the electron transporter chain. However, after photosensitization of riboflavin with UV or visible light, it generates reactive oxygen species (ROS), which can oxidize the DNA. The repair of oxidative lesions on DNA occurs through the base excision repair pathway (BER), where APE1 endonuclease plays a central role. On the other hand, the nucleotide excision repair pathway (NER) repairs helix-distorting lesions. Recently, it was described the participation of NERproteins in the repair of oxidative damage and in stimulation of repair function fromAPE1. The aim of this research was to evaluate the cytotoxic effects of photosensitized riboflavin (RF*) in cells proficient and deficient in NER, correlating with APE1 expression. For this propose, the cells were treated with RF* and it was performed the cell viability assay, extraction of whole proteins, cells fractionation, immunoblotting, indirect immunofluorescence and analysis of polymorphisms of BER gens. The results evidenced that cells deficient in XPA and CSB proteins were more sensitive to RF*. However, XPC-deficient cells presented similar resistance to MRC5- SV cells, which is proficient in NER. These results indicate that XPA and CSB proteins have an important role on repair of oxidative lesions induced by RF*. Additionally, it was evidenced that single nucleotide polymorphisms (SNPs) in BER enzymes may influence in sensitivity of NER-deficient cell lines. Concerning the APE1 expression, the results showed that expression of this protein after treatment with RF* only changed in XPC-deficient cells. Though, it was observed that APE1 is recruited and is bound to chromatin in MRC5-SV and XPA cells after treatment with RF*. The results also showed the induction of DNA damage after treatment with RF*, through the analysis of-H2AX, since the treatment promoted an increase of endogenous levels of this phosphorylated protein, which acts signaling double strand-break on DNA. On the other hand, in XPC-deficient cells, regardless of resistance of RF*, the endogenous levels of APE1 are extremely reduced when compared with other cell lines and APE1 is not bound to chromatin after treatment with RF*. These results conclude that RF* was able to induce cell death in NERdeficient cells, where XPA and CSB cells were more sensitive when compared with MRC5-SV and XPC-deficient cells. This last result is potentially very interesting, since XPC-deficient cell line presents low levels of APE1. Additionally, the results evidenced that APE1 protein can be involved in the repair of oxidative damage induced by RF*, because APE1 is recruited and bound strongly to chromatin after treatment.

Functional XPB/RAD25 redundancy in Arabidopsis genome: characterization of AtXPB2 and expression analysis

The xeroderma pigmentosum complementation group B (XPB) protein is involved in both DNA repair and transcription in human cells. It is a component of the transcription factor IIH (TFIIH) and is responsible for DNA helicase activity during nucleotide (nt) excision repair (NER). Its high evolutionary conservation has allowed identification of homologous proteins in different organisms, including plants. In contrast to other organisms, Arabidopsis thaliana harbors a duplication of the XPB orthologue (AtXPB1 and AtXPB2), and the proteins encoded by the duplicated genes are very similar (95% amino acid identity). Complementation assays in yeast rad25 mutant strains suggest the involvement of AtXPB2 in DNA repair, as already shown for AtXPB1, indicating that these proteins may be functionally redundant in the removal of DNA lesions in A. thaliana. Although both genes are expressed in a constitutive manner during the plant life cycle, Northern blot analyses suggest that light modulates the expression level of both XPB copies, and transcript levels increase during early stages of development. Considering the high similarity between AtXPB1 and AtXPB2 and that both of predicted proteins may act in DNA repair, it is possible that this duplication may confer more flexibility and resistance to DNA damaging agents in thale cress. (C) 2004 Elsevier B.V. All rights reserved.

Avaliação de efeitos de um extrato aquoso de cinnamomum zeylanicum L. na marcação de constituintes sanguíneos com tecnécio-99m e na morfologia das hemácias de ratos wistar

Radiobiocomplexes are used to obtain images in nuclear medicine and employed in basic research. Blood constituents labeled with technetium-99m (99mTc) have also been employed as radiobiocomplexes and used also experimental model for evaluation of the biological effects of natural or synthetic drugs. The analysis of the morphology and the morphometrics parameters (perimeter/área ratio) can be used to evaluate the effects of drugs upon the structure of the membrane of red blood cells. Cinnamomum zeylanicum (cinnamon) is a spice used as herbal medicine to treat diseases. The aim of this study was to evaluate the effect of in vitro and in vivo treatment with an aqueous cinnamon extract on the labeling of blood constituents with 99mTc and on the morphology of red blood cells from Wistar rats. In the in vitro treatment, isolated blood sample from animals were incubated with cinnamon extract. In the in vivo treatment, blood samples were also withdrawn from animals treated with cinnamon extract. In both cases, the radiolabeling of blood constituents was done. The morphological analysis of red blood cells was also done. As control, blood or animals treated with NaCl 0.9%. The data obtained on the labeling of blood constituents with 99mTc experiments indicated that the in vitro treatment with cinnamon extract was capable to decrease signiicantly (p<0.05) the percentage of radioactivity in cellular compartments and on the fixation of cellular and plasma proteins. These effects were not observed on the in vivo treatment. The results obtained for the morphology of red blood cells suggest that the in vitro and in vivo treatments did not alter the morphology and the perimeter/area ratio. The in vitro treatment with aqueous cinnamon extract could affect the membrane structures related with the oxidation status of the stannous ion pertechnetate ion, altering the labeling of blood constituentes with 99mTc. This study was a multidisciplinary experimental research. It was developed with the contribution of the different Departments and Services of the Hospital Universitário Pedro Ernesto of the Universidade do Estado do Rio de Janeiro