Protein crystals in adenovirus type 5-infected cells: requirements for intranuclear crystallogenesis, structural and functional analysis

Intranuclear crystalline inclusions have been observed in the nucleus of epithelial cells infected with Adenovirus serotype 5 (Ad5) at late steps of the virus life cycle. Using immuno-electron microscopy and confocal microscopy of cells infected with various Ad5 recombinants modified in their penton base or fiber domains, we found that these inclusions represented crystals of penton capsomers, the heteromeric capsid protein formed of penton base and fiber subunits. The occurrence of protein crystals within the nucleus of infected cells required the integrity of the fiber knob and part of the shaft domain. In the knob domain, the region overlapping residues 489–492 in the FG loop was found to be essential for crystal formation. In the shaft, a large deletion of repeats 4 to 16 had no detrimental effect on crystal inclusions, whereas deletion of repeats 8 to 21 abolished crystal formation without altering the level of fiber protein expression. This suggested a crucial role of the five penultimate repeats in the crystallisation process. Chimeric pentons made of Ad5 penton base and fiber domains from different serotypes were analyzed with respect to crystal formation. No crystal was found when fiber consisted of shaft (S) from Ad5 and knob (K) from Ad3 (heterotypic S5-K3 fiber), but occurred with homotypic S3K3 fiber. However, less regular crystals were observed with homotypic S35-K35 fiber. TB5, a monoclonal antibody directed against the Ad5 fiber knob was found by immunofluorescence microscopy to react with high efficiency with the intranuclear protein crystals in situ. Data obtained with Ad fiber mutants indicated that the absence of crystalline inclusions correlated with a lower infectivity and/or lower yields of virus progeny, suggesting that the protein crystals might be involved in virion assembly. Thus, we propose that TB5 staining of Ad-infected 293 cells can be used as a prognostic assay for the viability and productivity of fiber-modified Ad5 vectors

PTSD-8: A Short PTSD Inventory

Traumatic events pose great challenges on mental health services in scarcity of specialist trauma clinicians and services. Simple short screening instruments for detecting adverse psychological responses are needed. Several brief screening instruments have been developed. However, some are limited, especially in relation to reflecting the posttraumatic stress disorder (PTSD) diagnosis. Recently, several studies have challenged pre-existing ideas about PTSD’s latent structure. Factor analytic research currently supports two four factor models. One particular model contains a dysphoria factor which has been associated with depression and anxiety. The symptoms in this factor have been hailed as less specific to PTSD. The scope of this article is therefore to present a short screening instrument, based on this research; Posttraumatic Stress Disorder (PTSD) – 8 items. The PTSD-8 is shown to have good psychometric properties in three independent samples of whiplash patients (n=1710), rape victims (n=305), and disaster victims (n=516). Good test-rest reliability is also shown in a pilot study of young adults from families with alcohol problems (n=56)

Description et évaluation d'un réseau d'épidémiosurveillance des pathologies porcines mis en place dans un district du Nord Vietnam

Background and objective: Early menarche is increasing in prevalence worldwide, prompting clinical andpublic health interest on its links with pulmonary function. We aimed to investigate the relationship betweenearly menarche and lung function in middle age.Methods: The population-based Tasmanian LongitudinalHealth Study (born 1961; n = 8583), was initiated in 1968.The 5th Decade follow-up data (mean age: 45 years)included age at menarche and complex lung function testing. The 6th Decade follow-up (age: 53 years) repeated spirometry and gas transfer factor. Multiple linear regressionand mediation analyses were performed to determine theassociation between age at menarche and adult lung function and investigate biological pathways, including the proportion mediated by adult-attained height.Results: Girls reporting an early menarche (Conclusion:Early menarche was associated withreduced adult lung function. This is the first study toinvestigate post-BD outcomes and quantify the partialrole of adult height in this association

Overexpression of human kynurenine-3-monooxygenase protects against 3-hydroxykynurenine-mediated apoptosis through bidirectional non-linear feedback

Kynurenine 3-monooxygenase (KMO) is a critical regulator of inflammation. The preferred KMO substrate, kynurenine, is converted to 3-hydroxykynurenine (3HK), and this product exhibits cytotoxicity through mechanisms that culminate in apoptosis. Here, we report that overexpression of human KMO with orthotopic localisation to mitochondria creates a metabolic environment during which the cell exhibits increased tolerance for exogenous 3HK-mediated cellular injury. Using the selective KMO inhibitor Ro61-8048, we show that KMO enzyme function is essential for cellular protection. Pan-caspase inhibition with Z-VAD-FMK confirmed apoptosis as the mode of cell death. By defining expression of pathway components upstream and downstream of KMO, we observed alterations in other key kynurenine pathway components, particularly tryptophan-2,3-dioxygenase upregulation, through bidirectional nonlinear feedback. KMO overexpression also increased expression of inducible nitric oxide synthase (iNOS). These changes in gene expression are functionally relevant, because siRNA knockdown of the pathway components kynureninase and quinolinate phosphoribosyl transferase caused cells to revert to a state of susceptibility to 3HK-mediated apoptosis. In summary, KMO overexpression, and importantly KMO activity, have metabolic repercussions that fundamentally affect resistance to cell stress

Peritrophic matrix of Phlebotomus duboscqi and its kinetics during Leishmania major development

Light microscopy of native preparations, histology, and electron microscopy have revealed that Phlebotomus duboscqi belongs to a class of sand fly species with prompt development of the peritrophic matrix (PM). Secretion of electron-lucent fibrils, presumably chitin, starts immediately after the ingestion of a blood meal and, about 6 h later, is followed by secretion of amorphous electron-dense components, presumably proteins and glycoproteins. The PM matures in less than 12 h and consists of a thin laminar outer layer and a thick amorphous inner layer. No differences have been found in the timing of the disintegration of the PM in females infected with Leishmania major. In both groups of females (infected and uninfected), the disintegration of the PM is initiated at the posterior end. Although parasites are present at high densities in the anterior part of the blood meal bolus, they escape from the PM at the posterior end only. These results suggest that L. major chitinase does not have an important role in parasite escape from the PM. Promastigotes remain in the intraperitrophic space until the PM is broken down by sand-fly-derived chitinases and only then migrate anteriorly. Disintegration of the PM occurs simultaneously with the morphological transformation of parasites from procyclic forms to long nectomonads. A novel role is ascribed to the anterior plug, a component of the PM secreted by the thoracic midgut; this plug functions as a temporary barrier to stop the forward migration of nectomonads to the thoracic midgut

Should I Stay or Should I Go? A Habitat-Dependent Dispersal Kernel Improves Prediction of Movement

The analysis of animal movement within different landscapes may increase our understanding of how landscape features affect the perceptual range of animals. Perceptual range is linked to movement probability of an animal via a dispersal kernel, the latter being generally considered as spatially invariant but could be spatially affected. We hypothesize that spatial plasticity of an animal's dispersal kernel could greatly modify its distribution in time and space. After radio tracking the movements of walking insects (Cosmopolites sordidus) in banana plantations, we considered the movements of individuals as states of a Markov chain whose transition probabilities depended on the habitat characteristics of current and target locations. Combining a likelihood procedure and pattern-oriented modelling, we tested the hypothesis that dispersal kernel depended on habitat features. Our results were consistent with the concept that animal dispersal kernel depends on habitat features. Recognizing the plasticity of animal movement probabilities will provide insight into landscape-level ecological processes

Wnt, Hedgehog and Junctional Armadillo/β-Catenin Establish Planar Polarity in the Drosophila Embryo

To generate specialized structures, cells must obtain positional and directional information. In multi-cellular organisms, cells use the non-canonical Wnt or planar cell polarity (PCP) signaling pathway to establish directionality within a cell. In vertebrates, several Wnt molecules have been proposed as permissible polarity signals, but none has been shown to provide a directional cue. While PCP signaling components are conserved from human to fly, no PCP ligands have been reported in Drosophila. Here we report that in the epidermis of the Drosophila embryo two signaling molecules, Hedgehog (Hh) and Wingless (Wg or Wnt1), provide directional cues that induce the proper orientation of Actin-rich structures in the larval cuticle. We further find that proper polarity in the late embryo also involves the asymmetric distribution and phosphorylation of Armadillo (Arm or β-catenin) at the membrane and that interference with this Arm phosphorylation leads to polarity defects. Our results suggest new roles for Hh and Wg as instructive polarizing cues that help establish directionality within a cell sheet, and a new polarity-signaling role for the membrane fraction of the oncoprotein Arm

Characterization of the Modes of Binding between Human Sweet Taste Receptor and Low-Molecular-Weight Sweet Compounds

One of the most distinctive features of human sweet taste perception is its broad tuning to chemically diverse compounds ranging from low-molecular-weight sweeteners to sweet-tasting proteins. Many reports suggest that the human sweet taste receptor (hT1R2–hT1R3), a heteromeric complex composed of T1R2 and T1R3 subunits belonging to the class C G protein–coupled receptor family, has multiple binding sites for these sweeteners. However, it remains unclear how the same receptor recognizes such diverse structures. Here we aim to characterize the modes of binding between hT1R2–hT1R3 and low-molecular-weight sweet compounds by functional analysis of a series of site-directed mutants and by molecular modeling–based docking simulation at the binding pocket formed on the large extracellular amino-terminal domain (ATD) of hT1R2. We successfully determined the amino acid residues responsible for binding to sweeteners in the cleft of hT1R2 ATD. Our results suggest that individual ligands have sets of specific residues for binding in correspondence with the chemical structures and other residues responsible for interacting with multiple ligands