Interferon induction by avian reovirus

We have previously shown that the replication of avian reovirus (ARV) in chicken embryo fibroblasts (CEF) is more resistant to the antiviral action of interferon (IFN) than the replication of vesicular stomatitis virus (VSV) or vaccinia virus (VV). In this study we examined the capacity of these three viruses to induce the expression of IFN when infecting avian cells. Efficient expression of both type-α and type-β IFNs, as well as of the double-stranded RNA (dsRNA)-activated protein kinase (PKR), takes place in ARV-infected CEF, but not in cells infected with VSV or VV. PKR expression is not directly induced by ARV infection, but by the IFN secreted by ARV-infected cells. IFN induction in ARV-infected cells requires viral uncoating, but not viral gene expression, a situation similar to that reported for apoptosis induction by ARV-infected cells. However, our results demonstrate that IFN induction by ARV-infected CEF occurs by a caspase-independent mechanismThis work was funded by grants from the Spanish Ministerio de Economia y Competitividad (BFU2010-22228 and BFU2013-43513-R) and from the Xunta de Galicia (CN 2012/018). I.L.S. was a recipient of a predoctoral FPU fellowship (Ministerio de Educación y Ciencia) and a Research Fellowship (Bolsa de Investigación) (Deputación Provincial da Coruña)S

Response of Three Different Viruses to Interferon Priming and Dithiothreitol Treatment of Avian Cells

We have previously shown that the replication of avian reovirus (ARV) in chicken cells is much more resistant to interferon (IFN) than the replication of vesicular stomatitis virus (VSV) or vaccinia virus (VV). In this study, we have investigated the role that the double-stranded RNA (dsRNA)-activated protein kinase (PKR) plays in the sensitivity of these three viruses toward the antiviral action of chicken interferon. Our data suggest that while interferon priming of avian cells blocks vaccinia virus replication by promoting PKR activation, the replication of vesicular stomatitis virus appears to be blocked at a pretranslational step. Our data further suggest that the replication of avian reovirus in chicken cells is quite resistant to interferon priming because this virus uses strategies to downregulate PKR activation and also because translation of avian reovirus mRNAs is more resistant to phosphorylation of the alpha subunit of initiation factor eIF2 than translation of their cellular counterparts. Our results further reveal that the avian reovirus protein sigmaA is able to prevent PKR activation and that this function is dependent on its double-stranded RNA-binding activity. Finally, this study demonstrates that vaccinia virus and avian reovirus, but not vesicular stomatitis virus, express/induce factors that counteract the ability of dithiothreitol to promote eIF2 phosphorylation. Our data demonstrate that each of the three different viruses used in this study elicits distinct responses to interferon and to dithiothreitol-induced eIF2 phosphorylation when infecting avian cellsThe Spanish Ministerio de Economía y Competitividad provided funding to Javier Benavente and José Martínez-Costas under grant numbers BFU2010-22228 and BFU2013-43513-R. Support was also provided by the Xunta de Galicia (CN 2012/018). Irene Lostalé-Seijo was a recipient of a predoctoral FPU fellowship (Ministerio de Educación y Ciencia) and a Research Fellowship (Bolsa de Investigación; Deputación Provincial da Coruña)S

Avian reovirus-triggered apoptosis enhances both virus spread and the processing of the viral nonstructural muNS protein

Avian reovirus non-structural protein muNS is partially cleaved in infected chicken embryo fibroblast cells to produce a 55-kDa carboxyterminal protein, termed muNSC, and a 17-kDa aminoterminal polypeptide, designated muNSN. In this study we demonstrate that muNS processing is catalyzed by a caspase 3-like protease activated during the course of avian reovirus infection. The cleavage site was mapped by site directed mutagenesis between residues Asp-154 and Ala-155 of the muNS sequence. Although muNS and muNSC, but not muNSN, are able to form inclusions when expressed individually in transfected cells, only muNS is able to recruit specific ARV proteins to these structures. Furthermore, muNSC associates with ARV factories more weakly than muNS, sigmaNS and lambdaA. Finally, the inhibition of caspase activity in ARV-infected cells does not diminish ARV gene expression and replication, but drastically reduces muNS processing and the release and dissemination of progeny viral particlesThis work was funded by grants from the Ministerio de Economia Competitividad (BFU2010-22228) and from the Xunta de Galicia (CN 2012/018)S

Different intracellular distribution of avian reovirus core protein sigmaA in cells of avian and mammalian origin

A comparative analysis of the intracellular distribution of avian reovirus (ARV) core protein sigmaA in cells of avian and mammalian origin revealed that, whereas the viral protein accumulates in the cytoplasm and nucleolus of avian cells, most sigmaA concentrates in the nucleoplasm of mammalian cells in tight association with the insoluble nuclear matrix fraction. Our results further showed that sigmaA becomes arrested in the nucleoplasm of mammalian cells via association with mammalian cell-specific factors and that this association prevents nucleolar targeting. Inhibition of RNA polymerase II activity, but not of RNA polymerase I activity, in infected mammalian cells induces nucleus-to-cytoplasm sigmaA translocation through a CRM1- and RanGTP-dependent mechanism, yet a heterokaryon assay suggests that sigmaA does not shuttle between the nucleus and cytoplasm. The scarcity of sigmaA in cytoplasmic viral factories of infected mammalian cells could be one of the factors contributing to limited ARV replication in mammalian cellsThis research was supported by grants from the Spanish Ministerio de Ciencia y Tecnología (BFU2007-61330/BMC) and from the Xunta de Galicia (08CSA009203PR). L. V-I. and I. L-S. were recipients of predoctoral fellowships from the FPI and FPU programs of the Spanish Ministerio de Ciencia y TecnologíaS

IC-Tagging methodology applied to the expression of viral glycoproteins and the difficult-to-express membrane-bound IGRP autoantigen

We have previously developed a methodology to produce protein microspheres (MS) that can be loaded with proteins of interest in living cells through their C or N-terminal tagging with the so-called IC-Tag. The IC-Tagging method has many applications ranging from the production of immobilized enzymes for industrial use to the production of subunit vaccines due to its intrinsic adjuvancy. Here we show the adaptation of the IC-Tagging to work inside the endoplasmic reticulum and bacteria, allowing us to produce properly modified viral glycoproteins. Additionally, we were able to express the Islet-specific glucose-6-phosphatase catalytic subunit-related protein (IGRP), whose expression remained elusive to date possibly due to its toxicity when over-expressed. IGRP is an antigen of enormous pharmaceutical interest as it is specifically targeted during the autoimmune response taking place in both the Non-Obese Diabetic (NOD) mice and type 1 diabetes (T1D) patients leading to the destruction of insulin-producing beta cellsThis work was financed by the Spanish Ministerio de Economía y Competitividad, grant BFU2013-43513-R. Financial support from the Xunta de Galicia (Centro singular de investigación de Galicia accreditation 2016–2019) and the European Union (European Regional Development Fund - ERDF), is gratefully acknowledged. Irene Lostalé-Seijo was a recipient of a predoctoral FPU fellowship (Ministerio de Educación y Ciencia) and a Research Fellowship (Bolsa de Investigación; Deputación Provincial da Coruña)S

Microspheres-prime/rMVA-boost vaccination enhances humoral and cellular immune response in IFNAR(−/−) mice conferring protection against serotypes 1 and 4 of bluetongue virus

Bluetongue virus (BTV) is the causative agent of bluetongue disease (BT), which affects domestic and wild ruminants. At the present, 27 different serotypes have been documented. Vaccination has been demonstrated as one of the most effective methods to avoid viral dissemination. To overcome the drawbacks associated with the use of inactivated and attenuated vaccines we engineered a new recombinant BTV vaccine candidate based on proteins VP2, VP7, and NS1 of BTV-4 that were incorporated into avian reovirus muNS-Mi microspheres (MS-VP2/VP7/NS1) and recombinant modified vaccinia virus Ankara (rMVA). The combination of these two antigen delivery systems in a heterologous prime-boost vaccination strategy generated significant levels of neutralizing antibodies in IFNAR(−/−) mice. Furthermore, this immunization strategy increased the ratio of IgG2a/IgG1 in sera, indicating an induction of a Th1 response, and elicited a CD8 T cell response. Immunized mice were protected against lethal challenges with the homologous serotype 4 and the heterologous serotype 1 of BTV. All these results support the strategy based on microspheres in combination with rMVAs as a promising multiserotype vaccine candidate against BTVThis work was supported by grants from the Spanish Ministerio de Economía y Competitividad (AGL2011-23506, AGL-2014-57430-R and BFU2013-43513-R). Financial support from the Consellería de Cultura, Educación e Ordenación Universitaria (Centro singular de investigación de Galicia accreditation 2016–2019, ED431G/09) and the European Regional Development Fund (ERDF), is also gratefully acknowledgedS

Diseño de la asignatura Trabajo Fin de Grado en Geología (Facultad de Ciencias, UA)

Con la implantación en el año académico 2013-14 del cuarto y último curso del grado en Geología en la Universidad de Alicante, se ha implementado la asignatura obligatoria de Trabajo Fin de Grado en Geología (TFGG) con el objetivo de que el estudiante demuestre las competencias adquiridas durante sus estudios de geología, tanto las específicas de la titulación, como las competencias generales o transversales, tal como son las ligadas a la búsqueda y organización de documentación, a la redacción clara y concisa de una memoria escrita que recoja un trabajo original del alumno y a la presentación en público de su trabajo de manera clara y adecuada. Siguiendo las directrices que se establecen, tanto en la Facultad de Ciencias, como en la Universidad de Alicante se ha diseñado una asignatura Trabajo Fin de Grado con varias modalidades o líneas de actividad docente. Se ha constituido una Comisión de Trabajo de Fin de Grado en Geología (CTFGG) con el propósito principal de organizar y supervisar la asignación, seguimiento y evaluación de los trabajos

Compétence en Technologie de l’Information Géographique (TIG) dans les études universitaires: réflexions et propositions participative

El objetivo de este trabajo es ofrecer una reflexión que conduzca a una asignación coherente de competencias en la enseñanza sobre Tecnologías de la Información Geográfica (Cartografía, Sistemas de Información Geográfica y Teledetección) que se imparte en la universidad. Tras analizar diversas propuestas al respecto, en España y a nivel internacional, y a través de una serie de talleres participativos, un grupo de académicos de la Universidad de Alcalá ha generado su propia propuesta sobre las competencias más importantes y necesarias a conseguir en cada nivel de la educación superior (grado, master y doctorado). La propuesta ha sido contrastada con los resultados de una encuesta realizada entre un conjunto de académicos universitarios españoles.The aim of this paper is to offer an insight on competences leading to their consistent allocation in the different courses on Geographic Information Technologies (Cartography, Geographic Information Systems - GIS and Remote Sensing) taught in universities. To do this, after analyzing the various proposals on the subject, both in Spain and internationally, and after conducting a series of participatory workshops, a group of academics from the University of Alcalá has generated its own proposal on the most important and necessary competences to acquire at each level of higher education (bachelor, master and doctorate). Furthermore, the proposal has been contrasted with the results of a survey conducted by a wide range of Spanish university academics.Le but de cet article est de proposer une réflexion conduisant à une allocation cohérente des compétences dans les différents cours sur les technologies d’information géographique (Cartographie, Système d’Information Géographique- SIG et télédétection) enseignées dans les universités. Pour ce faire, après avoir analysé les différentes propositions sur le sujet, à la fois en Espagne et à l’étranger, et après la réalisation d’une série d’ateliers participatifs, un groupe de l’Université de Alcalá a généré sa propre proposition sur les compétences les plus importants et nécessaires pour obtenir à chaque niveau de l’enseignement supérieur (licence, master et doctorat). En outre, la proposition a été contrastée avec les résultats d’une enquête menée par un large éventail d’universitaires espagnols

Occupational exposure to pesticides and endometrial cancer in the Screenwide case-control study

Background: Endometrial cancer is the most common gynaecological tumour in developed countries and disease burden is expected to increase over the years. Identifying modifiable risk factors may help developing strategies to reduce the expected increasing incidence of these neoplasms. Objective: This study evaluates the association between occupational exposure to pesticides and endometrial cancer using data from a recent case-control study in Spain. Methods: The analyses included data from 174 consecutive incident endometrial cancer cases and 216 hospital controls frequency-matched by age. Data were collected through structured epidemiological questionnaires and exposure to pesticides was assessed using a Spanish job-exposure matrix (MatEmESp). Results: Overall, 12% of controls and 18% of cases were occupationally exposed to pesticides. We observed a positive association between occupational exposure to pesticides and endometrial cancer (OR = 2.08; 95% CI = 1.13-3.88 compared to non-exposed). In general, exposures that occurred farther in the past were significantly associated with endometrial cancer. Exposure to insecticides, fungicides and herbicides were positively associated with endometrial cancer (OR = 2.08; 95% CI = 1.13-3.88, OR = 4.40; 95% CI = 1.65-13.33, and OR = 5.25; 95% CI = 1.84-17.67, respectively). The agricultural, poultry and livestock activities scenario was associated with endometrial cancer (OR = 4.16; 95% CI = 1.59-12.32), while the cleaning exposure scenario was not (OR = 1.22; 95% CI = 0.55-2.67). Conclusions: Assessment of occupational exposure to pesticides assessed using a Spanish job-exposure matrix revealed a positive association with endometrial cancer. The elucidation of the role of pesticide compounds on endometrial cancer should shed a light on the aetiology of this tumour