Arenavirus Nucleoprotein Targets Interferon Regulatory Factor-Activating Kinase IKKε.

Arenaviruses perturb innate antiviral defense by blocking induction of type I interferon (IFN) production. Accordingly, the arenavirus nucleoprotein (NP) was shown to block activation and nuclear translocation of interferon regulatory factor 3 (IRF3) in response to virus infection. Here, we sought to identify cellular factors involved in innate antiviral signaling targeted by arenavirus NP. Consistent with previous studies, infection with the prototypic arenavirus lymphocytic choriomeningitis virus (LCMV) prevented phosphorylation of IRF3 in response to infection with Sendai virus, a strong inducer of the retinoic acid-inducible gene I (RIG-I)/mitochondrial antiviral signaling (MAVS) pathway of innate antiviral signaling. Using a combination of coimmunoprecipitation and confocal microscopy, we found that LCMV NP associates with the IκB kinase (IKK)-related kinase IKKε but that, rather unexpectedly, LCMV NP did not bind to the closely related TANK-binding kinase 1 (TBK-1). The NP-IKKε interaction was highly conserved among arenaviruses from different clades. In LCMV-infected cells, IKKε colocalized with NP but not with MAVS located on the outer membrane of mitochondria. LCMV NP bound the kinase domain (KD) of IKKε (IKBKE) and blocked its autocatalytic activity and its ability to phosphorylate IRF3, without undergoing phosphorylation. Together, our data identify IKKε as a novel target of arenavirus NP. Engagement of NP seems to sequester IKKε in an inactive complex. Considering the important functions of IKKε in innate antiviral immunity and other cellular processes, the NP-IKKε interaction likely plays a crucial role in arenavirus-host interaction.

Expression of a functional single chain antibody on the surface of extracellular enveloped vaccinia virus as a step towards selective tumour cell targeting.

Recombinant vaccinia virus with tumour cell specificity may provide a versatile tool either for direct lysis of cancer cells or for the targeted transfer of genes encoding immunomodulatory molecules. We report the expression of a single chain antibody on the surface of extracellular enveloped vaccinia virus. The wild-type haemagglutinin, an envelope glycoprotein which is not required for viral infection and replication, was replaced by haemagglutinin fusion molecules carrying a single chain antibody directed against the tumour-associated antigen ErbB2. ErbB2 is an epidermal growth factor receptor-related tyrosine kinase overexpressed in a high percentage of human adenocarcinomas. Two fusion proteins carrying the single chain antibody at different NH2-terminal positions were expressed and exposed at the envelope of the corresponding recombinant viruses. The construct containing the antibody at the site of the immunoglobulin-like loop of the haemagglutinin was able to bind solubilized ErbB2. This is the first report of replacement of a vaccinia virus envelope protein by a specific recognition structure and represents a first step towards modifying the host cell tropism of the virus.

The SOCS3-independent expression of IDO2 supports the homeostatic generation of T regulatory cells by human dendritic cells.

Dendritic cells (DCs) are professional APCs that have a role in the initiation of adaptive immune responses and tolerance. Among the tolerogenic mechanisms, the expression of the enzyme IDO1 represents an effective tool to generate T regulatory cells. In humans, different DC subsets express IDO1, but less is known about the IDO1-related enzyme IDO2. In this study, we found a different pattern of expression and regulation between IDO1 and IDO2 in human circulating DCs. At the protein level, IDO1 is expressed only in circulating myeloid DCs (mDCs) and is modulated by PGE2, whereas IDO2 is expressed in both mDCs and plasmacytoid DCs and is not modulated by PGE2. In healthy subjects, IDO1 expression requires the presence of PGE2 and needs continuous transcription and translation, whereas IDO2 expression is constitutive, independent from suppressor of cytokine signaling 3 activity. Conversely, in patients suffering from inflammatory arthritis, circulating DCs express both IDO1 and IDO2. At the functional level, both mDCs and plasmacytoid DCs generate T regulatory cells through an IDO1/IDO2-dependent mechanism. We conclude that, in humans, whereas IDO1 provides an additional mechanism of tolerance induced by proinflammatory mediators, IDO2 is stably expressed in steady-state conditions and may contribute to the homeostatic tolerogenic capacity of DCs.

Description of Trichuris travassosi n. sp. (Nematoda:Trichurinae) from a brasilian rodent, by light and scanning electron microscopy

A new species of a trichurid nematode Trichuris travassosi n. sp., recovered from a wild rodent in the State of Rio Grande do Sul, Brazil, is described and compared to T. myocastoris (Enigk, 1933) and their differentiation was on the basis of detailed morphometrical study. Oryzomys nigripes (Olfers, 1818) is a new host record for the genus. The denomination spicular prepuce is proposed to designate the structure previously named spicular sheath and, conversely, spicular sheath to indicate the cuticle that convers the spicule.

Alpha beta lineage-committed thymocytes can be rescued by the gamma delta T cell receptor (TCR) in the absence of TCR beta chain.

Commitment of the alpha beta and gamma delta T cell lineages within the thymus has been studied in T cell receptor (TCR)-transgenic and TCR mutant murine strains. TCR gamma delta-transgenic or TCR beta knockout mice, both of which are unable to generate TCR alpha beta-positive T cells, develop phenotypically alpha beta-like thymocytes in significant proportions. We provide evidence that in the absence of functional TCR beta protein, the gamma delta TCR can promote the development of alpha beta-like thymocytes, which, however, do not expand significantly and do not mature into gamma delta T cells. These results show that commitment to the alpha beta lineage can be determined independently of the isotype of the TCR, and suggest that alpha beta versus gamma delta T cell lineage commitment is principally regulated by mechanisms distinct from TCR-mediated selection. To accommodate our data and those reported previously on the effect of TCR gamma and delta gene rearrangements on alpha beta T cell development, we propose a model in which lineage commitment occurs independently of TCR gene rearrangement.

Absolute requirement for the pre-T cell receptor alpha chain during NK1.1+ TCRalphabeta cell development.

Most natural killer T (NKT) cells express a highly skewed alphabeta TCR repertoire, consisting of an invariant V alpha14-J alpha281 chain paired preferentially with a polyclonal Vbeta8.2 chain. This repertoire is positively selected by the monomorphic CD1d molecule expressed on cells of hematopoietic origin. The origin of NKT cells and their lineage relationship to conventional T cells is controversial. We show here that the development of NKT cells is absolutely dependent on expression of the pre-TCRalpha chain, in marked contrast to conventional T cells which arise in significant numbers even in the absence of a functional pre-TCR. Distinct developmental requirements for pre-TCR expression in the NKT and T cell lineages may reflect differences in the ability of the TCRalphabeta to substitute functionally for the pre-TCR in immature precursor cells.

The Global Diffusion of Regulatory Agencies: Channels of Transfer and Stages of Diffusion

The autonomous regulatory agency has recently become the ‘appropriate model’ of governance across countries and sectors. The dynamics of this process is captured in our data set, which covers the creation of agencies in 48 countries and 16 sectors since the 1920s. Adopting a diffusion approach to explain this broad process of institutional change, we explore the role of countries and sectors as sources of institutional transfer at different stages of the diffusion process. We demonstrate how the restructuring of national bureaucracies unfolds via four different channels of institutional transfer. Our results challenge theoretical approaches that overemphasize the national dimension in global diffusion and are insensitive to the stages of the diffusion process. Further advance in study of diffusion depends, we assert, on the ability to apply both cross-sectoral and cross-national analysis to the same research design and to incorporate channels of transfer with different causal mechanisms for different stages of the diffusion process.

Pharmacokinetics and posterior segment biodistribution of ESBA105, an anti-TNF-alpha single-chain antibody, upon topical administration to the rabbit eye.

PURPOSE: The purpose of this study was to characterize local distribution and systemic absorption of the tumor necrosis factor (TNF)-alpha inhibitory single-chain antibody fragment (scFv) ESBA105 following topical administration to the eye in vivo. METHODS: Rabbits received ESBA105 as topical eye drops in two dosing regimens. First, pharmacokinetics after the topical route of administration was compared to the intravenous (i.v.) route by means of applying the identical cumulative daily dose of ESBA105. In a second study rabbits received five eye drops daily for six consecutive days in a lower frequency topical dosing regimen. Kinetics and biodistribution of ESBA105 in ocular tissues and fluids as well as in sera were determined in all animals. RESULTS: After topical administration to the eye, ESBA105 quickly reaches therapeutic concentrations in all ocular compartments. Systemic exposure after topical administration is 25,000-fold lower than exposure after i.v. injection of the identical cumulative daily dose. ESBA105 levels in vitreous humor and neuroretina are significantly higher on topical administration than after i.v. injection. Absolute and relative intraocular biodistribution of ESBA105 is different with topical and systemic delivery routes. Compared to its terminal half-life in circulation (7 hours), the vitreal half-life of ESBA105 is significantly enhanced (16-24 hours). CONCLUSIONS: On topical administration, ESBA105 is efficiently absorbed and distributed to all compartments of the eye, whereby systemic drug exposure is very low. Based on its unique intraocular biodistribution and pharmacokinetics and the absolute intraocular levels reached, topical ESBA105 appears highly attractive for treatment of various ophthalmological disorders.

Defining the site of light perception and initiation of phototropism in Arabidopsis.

Phototropism is an adaptive response allowing plants to optimize photosynthetic light capture. This is achieved by asymmetric growth between the shaded and lit sides of the stimulated organ. In grass seedlings, the site of phototropin-mediated light perception is distinct from the site of bending; however, in dicotyledonous plants (e.g., Arabidopsis), spatial aspects of perception remain debatable. We use morphological studies and genetics to show that phototropism can occur in the absence of the root, lower hypocotyl, hypocotyl apex, and cotyledons. Tissue-specific expression of the phototropin1 (phot1) photoreceptor demonstrates that light sensing occurs in the upper hypocotyl and that expression of phot1 in the hypocotyl elongation zone is sufficient to enable a normal phototropic response. Moreover, we show that efficient phototropism occurs when phot1 is expressed from endodermal, cortical, or epidermal cells and that its local activation rapidly leads to a global response throughout the seedling. We propose that spatial aspects in the steps leading from light perception to growth reorientation during phototropism differ between grasses and dicots. These results are important to properly interpret genetic experiments and establish a model connecting light perception to the growth response, including cellular and morphological aspects.