The Drosophila DIAP1 protein is required to prevent accumulation of a continuously generated, processed form of the apical caspase DRONC

Although loss of the inhibitor of apoptosis (LAP) protein DIAP1 has been shown to result in caspase activation and spontaneous cell death in Drosophila cells and embryos, the point at which DIAP1 normally functions to inhibit caspase activation is unknown. Depletion of the DIAP1 protein in Drosophila S2 cells or the Sf-IAP protein in Spodoptera frugiperda Sf21 cells by RNA interference (RNAi) or cycloheximide treatment resulted in rapid and widespread caspase-dependent apoptosis. Co-silencing of dronc or dark largely suppressed this apoptosis, indicating that DIAP1 is normally required to inhibit an activity dependent on these proteins. Silencing of dronc also inhibited DRICE processing following stimulation of apoptosis, demonstrating that DRONC functions as an apical caspase in S2 cells. Silencing of diap1 or treatment with UV light induced DRONC processing, which occurred in two steps. The first step appeared to occur continuously even in the absence of an apoptotic signal and to be dependent on DARK because full-length DRONC accumulated when dark was silenced in non-apoptotic cells. In addition, treatment with the proteasome inhibitor MG132 resulted in accumulation of this initially processed form of DRONC, but not full-length DRONC, in non-apoptotic cells. The second step in DRONC processing was observed only in apoptotic cells. These results indicate that the initial step in DRONC processing occurs continuously via a DARK-dependent mechanism in Drosophila cells and that DIAP1 is required to prevent excess accumulation of this first form of processed DRONC, presumably through its ability to act as a ubiquitin-protein ligase

Cleavage of the apoptosis inhibitor DIAP1 by the apical caspase DRONC in both normal and apoptotic drosophila cells

In Drosophila S2 cells, the apical caspase DRONC undergoes a low level of spontaneous autoprocessing. Unintended apoptosis is prevented by the inhibitor of apoptosis DIAP1, which targets the processed form of DRONC for degradation through its E3 ubiquitin protein ligase activity. Recent reports have demonstrated that shortly after the initiation of apoptosis in S2 cells, DIAP1 is cleaved following aspartate residue Asp-20 by the effector caspase DrICE. Here we report a novel caspase-mediated cleavage of DIAP1 in S2 cells. In both living and dying S2 cells, DIAP1 is cleaved by DRONC after glutamate residue Glu-205, located between the first and second BIR domains. The mutation of Glu-205 prevented the interaction of DIAP1 and processed DRONC but had no effect on the interaction with full-length DRONC. The mutation of Glu-205 also had a negative effect on the ability of overexpressed DIAP1 to prevent apoptosis stimulated by the proapoptotic protein Reaper or by UV light. These results expand our knowledge of the events that occur in the Drosophila apoptosome prior to and after receiving an apoptotic signal

The Drosophila Inhibitor of Apoptosis (IAP) DIAP2 Is Dispensable for Cell Survival, Required for the Innate Immune Response to Gram-negative Bacterial Infection, and Can Be Negatively Regulated by the Reaper/Hid/Grim Family of IAP-binding Apoptosis Inducers

Many inhibitor of apoptosis (IAP) family proteins inhibit apoptosis. IAPs contain N-terminal baculovirus IAP repeat domains and a C-terminal RING ubiquitin ligase domain. Drosophila IAP DIAP1 is essential for the survival of many cells, protecting them from apoptosis by inhibiting active caspases. Apoptosis initiates when proteins such as Reaper, Hid, and Grim bind a surface groove in DIAP1 baculovirus IAP repeat domains via an N-terminal IAP-binding motif. This evolutionarily conserved interaction disrupts DIAP1-caspase interactions, unleashing apoptosis-inducing caspase activity. A second Drosophila IAP, DIAP2, also binds Rpr and Hid and inhibits apoptosis in multiple contexts when overexpressed. However, due to a lack of mutants, little is known about the normal functions of DIAP2. We report the generation of diap2 null mutants. These flies are viable and show no defects in developmental or stress-induced apoptosis. Instead, DIAP2 is required for the innate immune response to Gram-negative bacterial infection. DIAP2 promotes cytoplasmic cleavage and nuclear translocation of the NF-{kappa}B homolog Relish, and this requires the DIAP2 RING domain. Increasing the genetic dose of diap2 results in an increased immune response, whereas expression of Rpr or Hid results in down-regulation of DIAP2 protein levels. Together these observations suggest that DIAP2 can regulate immune signaling in a dose-dependent manner, and this can be regulated by IBM-containing proteins. Therefore, diap2 may identify a point of convergence between apoptosis and immune signaling pathways

The Drosophila caspase Ice is important for many apoptotic cell deaths and for spermatid individualization, a nonapoptotic process

Caspase family proteases play important roles in the regulation of apoptotic cell death. Initiator caspases are activated in response to death stimuli, and they transduce and amplify these signals by cleaving and thereby activating effector caspases. In Drosophila, the initiator caspase Nc (previously Dronc) cleaves and activates two short-prodomain caspases, Dcp-1 and Ice (previously Drice), suggesting these as candidate effectors of Nc killing activity. dcp-1-null mutants are healthy and possess few defects in normally occurring cell death. To explore roles for Ice in cell death, we generated and characterized an Ice null mutant. Animals lacking Ice show a number of defects in cell death, including those that occur during embryonic development, as well as during formation of adult eyes, arista and wings. Ice mutants exhibit subtle defects in the destruction of larval tissues, and do not prevent destruction of salivary glands during metamorphosis. Cells from Ice animals are also markedly resistant to several stresses, including X-irradiation and inhibition of protein synthesis. Mutations in Ice also suppress cell death that is induced by expression of Rpr, Wrinkled (previously Hid) and Grim. These observations demonstrate that Ice plays an important non-redundant role as a cell death effector. Finally, we demonstrate that Ice participates in, but is not absolutely required for, the non-apoptotic process of spermatid differentiation

The air-rail intermodality of tourism in the city of Toledo

Con la apertura de la nueva infraestructura en Madrid que comunica las estaciones de Atocha y Chamartín con el Aeropuerto de Madrid, la ciudad de Toledo se encuentra directamente conectada con el aeropuerto internacional de Madrid, lo que abre nuevas posibilidades de oferta turística con la capacidad de fomentar el turismo internacional para la ciudad de Toledo. Teniendo en cuenta la importancia de la conexión con el aeropuerto internacional para la ciudad de Toledo, unido a la nueva tendencia de creación de líneas de autobuses directos al aeropuerto desde diferentes ciudades y a las recientes ofertas de billetes combinados TAV-Avión en otros corredores, este estudio se centra en la movilidad intermodal, en concreto aéreo-ferroviaria, del turista que accede a la ciudad de Toledo. La metodología utilizada es el análisis descriptivo de los datos obtenidos a través de una encuesta realizada en el corredor, que permite el análisis del modo de transporte utilizado por los turistas, en el acceso a la ciudad de Toledo y en particular desde el aeropuerto de Madrid.With the opening of the new infrastructure in Madrid connecting the Atocha and Chamartin stations with the airport, the city of Toledo has become directly connected to the international airport of Madrid. This provides new possibilities for tourism due to the ability to attract international tourism to the city of Toledo. Given the importance of connecting the international airport with the city of Toledo, together with the new trend of creating direct bus lines to the airport from different cities and the recent offers in combined TAV-Plan tickets for other routes, this study focuses on the intermodal mobility, in particular air-rail, of tourists visiting the city of Toledo. The methodology is based on a descriptive analysis of the data obtained through a survey conducted in the corridor that allows an analysis to be made of the mode of transport used by tourists to access the city of Toledo, particularly from Madrid airport.Este Trabajo se ha desarrollado en el marco del Proyecto “Integración del transporte aéreo y la alta velocidad ferroviaria: impactos sobre accesibilidad y medio ambiente (AERO-AVE)”, concedido por el Ministerio de Fomento (Orden FOM/3836/2008), en la Convocatoria de 2008 de Ayudas a proyectos de I+D del Subprograma para la Movilidad sostenible y el cambio modal en el transporte. Acción Estratégica Energía y Cambio climático, en el marco del Plan Nacional de Investigación Científica, Desarrollo e Innovación Tecnológica 2008-2011

Avaliação do método centróide para estudo de adaptabilidade ao ambiente de clones de <i>Eucalyptus grandis</i>.

A acentuada interação genótipo x ambiente presente em muitas culturas faz com que estudos de adaptabilidade a ambientes específicos sejam parte integrante dos programas de melhoramento vegetal. A resposta diferenciada dos genótipos a ambientes favoráveis e desfavoráveis pode ser estudada utilizando metodologias baseadas em três estratégias principais: análise de variância, regressão linear e em estatísticas não-paramétricas. Este trabalho apresenta uma nova metodologia para estudo de interação genótipo x ambiente que se baseia na comparação de valores de distância cartesiana entre os genótipos e quatro referências ideais em uma dispersão de componentes principais, visando a facilitar a recomendação de genótipos. Esse método, chamado de método centróide, difere em relação aos métodos baseados em análise de variância por permitir o direcionamento dos cultivares em relação à variação ambiental e pela facilidade de identificação dos genótipos, dispensando a análise simultânea de vários parâmetros como nos métodos baseados em regressão. Para exemplificar sua aplicação, foram avaliados 25 genótipos provenientes de testes clonais de Eucalyptus grandis aos 74 meses de idade plantados em quatro ambientes em modelo fatorial e delineamento em blocos ao acaso com seis repetições, sendo que os efeitos de genótipos foram considerados fixos e os efeitos de blocos e ambientes aleatórios. Foram identificados quatro clones de boa adaptabilidade geral além de outros de adaptação específica a grupo de ambientes que também podem ser recomendados visando a capitalizar o efeito da interação. Os resultados foram comparados com os obtidos pela metodologia de regressão proposta por Eberhart e Russel (1966) e pelo método proposto por Lin e Binns (1988) e permitem concluir que o método centróide foi eficiente na identificação dos clones de Eucalyptus grandis avaliados de comportamento diferenciado entre os ambientes; associado à facilidade de recomendação e ordenamento dos genótipos a grupos de adaptabilidade específicos.</p

EVALUATION OF THE CENTROID METHOD FOR STUDY OF ENVIRONMENT ADAPTABILITY OF CLONES OF Eucalyptus grandis

A acentuada intera\ue7\ue3o gen\uf3tipo x ambiente presente em muitas culturas faz com que estudos de adaptabilidade a ambientes espec\uedficos sejam parte integrante dos programas de melhoramento vegetal. A resposta diferenciada dos gen\uf3tipos a ambientes favor\ue1veis e desfavor\ue1veis pode ser estudada utilizando metodologias baseadas em tr\ueas estrat\ue9gias principais: an\ue1lise de vari\ue2ncia, regress\ue3o linear e em estat\uedsticas n\ue3o-param\ue9tricas. Este trabalho apresenta uma nova metodologia para estudo de intera\ue7\ue3o gen\uf3tipo x ambiente que se baseia na compara\ue7\ue3o de valores de dist\ue2ncia cartesiana entre os gen\uf3tipos e quatro refer\ueancias ideais em uma dispers\ue3o de componentes principais, visando a facilitar a recomenda\ue7\ue3o de gen\uf3tipos. Esse m\ue9todo, chamado de m\ue9todo centr\uf3ide, difere em rela\ue7\ue3o aos m\ue9todos baseados em an\ue1lise de vari\ue2ncia por permitir o direcionamento dos cultivares em rela\ue7\ue3o \ue0 varia\ue7\ue3o ambiental e pela facilidade de identifica\ue7\ue3o dos gen\uf3tipos, dispensando a an\ue1lise simult\ue2nea de v\ue1rios par\ue2metros como nos m\ue9todos baseados em regress\ue3o. Para exemplificar sua aplica\ue7\ue3o, foram avaliados 25 gen\uf3tipos provenientes de testes clonais de Eucalyptus grandis aos 74 meses de idade plantados em quatro ambientes em modelo fatorial e delineamento em blocos ao acaso com seis repeti\ue7\uf5es, sendo que os efeitos de gen\uf3tipos foram considerados fixos e os efeitos de blocos e ambientes aleat\uf3rios. Foram identificados quatro clones de boa adaptabilidade geral al\ue9m de outros de adapta\ue7\ue3o espec\uedfica a grupo de ambientes que tamb\ue9m podem ser recomendados visando a capitalizar o efeito da intera\ue7\ue3o. Os resultados foram comparados com os obtidos pela metodologia de regress\ue3o proposta por Eberhart e Russel (1966) e pelo m\ue9todo proposto por Lin e Binns (1988) e permitem concluir que o m\ue9todo centr\uf3ide foi eficiente na identifica\ue7\ue3o dos clones de Eucalyptus grandis avaliados de comportamento diferenciado entre os ambientes; associado \ue0 facilidade de recomenda\ue7\ue3o e ordenamento dos gen\uf3tipos a grupos de adaptabilidade espec\uedficos.The intense genotype x environment interaction present in many cultures make studies of adaptability to specific environments an important part of vegetal improvement programs. The different response of genotypes to favorable and unfavorable environments can be studied using methodologies based on three main strategies: analysis of variance, linear regression and non-parametric statistics. This work presents a new methodology for the study of genotype environment interaction, using principal components analysis, in order to facilitate the recommendation of specific genotypes to certain environments. This methodology, called centroid method, consists in the comparison of cartesian distance values between the genotypes and four ideal references using principal components. It differs from methods based in analysis of variance due to the aiming of genotypes according to environment variation and the facility of genotype identification, dispensing the analysis of several parameters as in methods based on regression. In order to exemplify the use of this method an experiment of Eucalyptus grandis planted in random blocks design and four environments was studied. Four clones of general adaptability were identified besides others of specific adaptation that can also be recommended aiming to capitalize the interaction effect. The results of this study were compared with the ones obtained through other methodologies by Eberhart e Russel (1966) and by Lin and Binns (1988) and allows to conclude that the centroid method was efficient in the identification of differentiated performance E. grandis clones; associated to the easiness of recommendation and ordering of the genotypes into specific adaptability groups

Practical Pharmacist-Led Interventions to Improve Antimicrobial Stewardship in Ghana, Tanzania, Uganda and Zambia.

The World Health Organisation (WHO) and others have identified, as a priority, the need to improve antimicrobial stewardship (AMS) interventions as part of the effort to tackle antimicrobial resistance (AMR). An international health partnership model, the Commonwealth Partnerships for Antimicrobial Stewardship (CwPAMS) programme, was established between selected countries in Africa (Ghana, Tanzania, Zambia and Uganda) and the UK to support AMS. This was funded by UK aid under the Fleming Fund and managed by the Commonwealth Pharmacists Association (CPA) and Tropical Health and Education Trust (THET). The primary aims were to develop local AMS teams and generate antimicrobial consumption surveillance data, quality improvement initiatives, infection prevention and control (IPC) and education/training to reduce AMR. Education and training were key components in achieving this, with pharmacists taking a lead role in developing and leading AMS interventions. Pharmacist-led interventions in Ghana improved access to national antimicrobial prescribing guidelines via the CwPAMS mobile app and improved compliance with policy from 18% to 70% initially for patients with pneumonia in one outpatient clinic. Capacity development on AMS and IPC were achieved in both Tanzania and Zambia, and a train-the-trainer model on the local production of alcohol hand rub in Uganda and Zambia. The model of pharmacy health partnerships has been identified as a model with great potential to be used in other low and middle income countries (LMICs) to support tackling AMR