Identification of a novel pathway essential for the immediate-early, interferon-independent antiviral response to enveloped virions

Viral infection elicits the activation of numerous cellular signal transduction pathways, leading to the induction of both innate and adaptive immunity. Previously we showed that entry of virion particles from a diverse array of enveloped virus families was capable of eliciting an interferon regulatory factor 3 (IRF-3)-mediated antiviral state in human fibroblasts in the absence of interferon production. Here we show that extracellular regulated kinase 1/2, p38 mitogen-activated protein kinase, and Jun N-terminal kinase/stress-activated protein kinase activities are not required for antiviral state induction. In contrast, treatment of cells with LY294002, an inhibitor of the phosphoinositide 3-kinase (PI3 kinase) family, prevents the induction of interferon-stimulated gene 56 (ISG56) and an antiviral response upon entry of virus particles. However, the prototypic class I p85/p110 PI3 kinase and its downstream effector Akt/PKB are dispensable for ISG and antiviral state induction. Furthermore, DNA-PK and PAK1, LY294002-sensitive members of the PI3 kinase family shown previously to be involved in IRF-3 activation, are also dispensable for ISG and antiviral state induction. The LY294002 inhibitor fails to prevent IRF-3 homodimerization or nuclear translocation upon virus particle entry. Together, these data suggest that virus entry triggers an innate antiviral response that requires the activity of a novel PI3 kinase family member.     

A single intranasal inoculation with a paramyxovirus-vectored vaccine protects guinea pigs against a lethal-dose Ebola virus challenge

To determine whether intranasal inoculation with a paramyxovirus-vectored vaccine can induce protective immunity against Ebola virus (EV), recombinant human parainfluenza virus type 3 (HPIV3) was modified to express either the EV structural glycoprotein (GP) by itself (HPIV3/EboGP) or together with the EV nucleoprotein (NP) (HPIV3/EboGP-NP). Expression of EV GP by these recombinant viruses resulted in its efficient incorporation into virus particles and increased cytopathic effect in Vero cells. HPIV3/EboGP was 100-fold more efficiently neutralized by antibodies to EV than by antibodies to HPIV3. Guinea pigs infected with a single intranasal inoculation of 10(5.3) PFU of HPIV3/EboGP or HPIV3/EboGP-NP showed no apparent signs of disease yet developed a strong humoral response specific to the EV proteins. When these animals were challenged with an intraperitoneal injection of 10(3) PFU of EV, there were no outward signs of disease, no viremia or detectable EV antigen in the blood, and no evidence of infection in the spleen, liver, and lungs. In contrast, all of the control animals died or developed severe EV disease following challenge. The highly effective immunity achieved with a single vaccine dose suggests that intranasal immunization with live vectored vaccines based on recombinant respiratory viruses may be an advantageous approach to inducing protective responses against severe systemic infections, such as those caused by hemorrhagic fever agents.     

Green tea polyphenols modulate insulin secretion by inhibiting glutamate dehydrogenase

Insulin secretion by pancreatic beta-cells is stimulated by glucose, amino acids, and other metabolic fuels. Glutamate dehydrogenase (GDH) has been shown to play a regulatory role in this process. The importance of GDH was underscored by features of hyperinsulinemia/hyperammonemia syndrome, where a dominant mutation causes the loss of inhibition by GTP and ATP. Here we report the effects of green tea polyphenols on GDH and insulin secretion. Of the four compounds tested, epigallocatechin gallate (EGCG) and epicatechin gallate were found to inhibit GDH with nanomolar ED(50) values and were therefore found to be as potent as the physiologically important inhibitor GTP. Furthermore, we have demonstrated that EGCG inhibits BCH-stimulated insulin secretion, a process that is mediated by GDH, under conditions where GDH is no longer inhibited by high energy metabolites. EGCG does not affect glucose-stimulated insulin secretion under high energy conditions where GDH is probably fully inhibited. We have further shown that these compounds act in an allosteric manner independent of their antioxidant activity and that the beta-cell stimulatory effects are directly correlated with glutamine oxidation. These results demonstrate that EGCG, much like the activator of GDH (BCH), can facilitate dissecting the complex regulation of insulin secretion by pharmacologically modulating the effects of GDH.     

Glycosaminoglycan Sulphation Affects the Seeded Misfolding of a Mutant Prion Protein

Background

The accumulation of protease resistant conformers of the prion protein (PrP^res) is a key pathological feature of prion diseases. Polyanions, including RNA and glycosaminoglycans have been identified as factors that contribute to the propagation, transmission and pathogenesis of prion disease. Recent studies have suggested that the contribution of these cofactors to prion propagation may be species specific.

Methodology/Principal Finding

In this study a cell-free assay was used to investigate the molecular basis of polyanion stimulated PrP^res formation using brain tissue or cell line derived murine PrP. Enzymatic depletion of endogenous nucleic acids or heparan sulphate (HS) from the PrP^C substrate was found to specifically prevent PrP^res formation seeded by mouse derived PrP^Sc. Modification of the negative charge afforded by the sulphation of glycosaminoglycans increased the ability of a familial PrP mutant to act as a substrate for PrP^res formation, while having no effect on PrP^res formed by wildtype PrP. This difference may be due to the observed differences in the binding of wild type and mutant PrP for glycosaminoglycans.

Conclusions/Significance

Cofactor requirements for PrP^res formation are host species and prion strain specific and affected by disease associated mutations of the prion protein. This may explain both species and strain dependent propagation characteristics and provide insights into the underlying mechanisms of familial prion disease. It further highlights the challenge of designing effective therapeutics against a disease which effects a range of mammalian species, caused by range of aetiologies and prion strains.     

Genetic reduction of mTOR extends lifespan in a mouse model of Hutchinson‐Gilford Progeria syndrome

Hutchinson‐Gilford progeria syndrome (HGPS) is a rare accelerated aging disorder most notably characterized by cardiovascular disease and premature death from myocardial infarction or stroke. The majority of cases are caused by a de novo single nucleotide mutation in the LMNA gene that activates a cryptic splice donor site, resulting in production of a toxic form of lamin A with a 50 amino acid internal deletion, termed progerin. We previously reported the generation of a transgenic murine model of progeria carrying a human BAC harboring the common mutation, G608G, which in the single‐copy state develops features of HGPS that are limited to the vascular system. Here, we report the phenotype of mice bred to carry two copies of the BAC, which more completely recapitulate the phenotypic features of HGPS in skin, adipose, skeletal, and vascular tissues. We further show that genetic reduction of the mechanistic target of rapamycin (mTOR) significantly extends lifespan in these mice, providing a rationale for pharmacologic inhibition of the mTOR pathway in the treatment of HGPS.     

A test of two mechanisms proposed to optimize grassland aboveground primary productivity in response to grazing

Aims Mesic grasslands have a long evolutionary history of grazing by large herbivores and as a consequence, grassland species have numerous adaptations allowing them to respond favourably to grazing. Although empirical evidence has been equivocal, theory predicts that such adaptations combined with alterations in resources can lead to grazing-induced overcompensation in aboveground net primary production (ANPP; grazed ANPP> ungrazed ANPP) under certain conditions. We tested two specific predictions from theory. First, overcompensation is more likely to occur in annually burned grasslands because limiting nutrients that would be lost with frequent fires are recycled through grazers and stimulate ANPP. Second, overcompensation of biomass lost to grazers is more likely to occur in unburned sites where grazing has the greatest effect on increasing light availability through alterations in canopy structure.Methods We tested these nutrient versus light-based predictions in grazed grasslands that had been annually burned or protected from fire for>20 years. We assessed responses in ANPP to grazing by large ungulates using both permanent and moveable grazing exclosures (252 exclosures from which biomass was harvested from 3192 quadrats) in a 2-year study. Study sites were located at the Konza Prairie Biological Station (KPBS) in North America and at Kruger National Park (KNP) in South Africa. At KPBS, sites were grazed by North American bison whereas in KNP sites were grazed either by a diverse suite of herbivores (e.g. blue wildebeest, Burchell's zebra, African buffalo) or by a single large ungulate (African buffalo).Important findings We found no evidence for overcompensation in either burned or unburned sites, regardless of grazer type. Thus, there was no support for either mechanism leading to overcompensation. Instead, complete compensation of total biomass lost to grazers was the most common response characterizing grazing–ANPP relationships with, in some cases, undercompensation of grass ANPP being offset by increased ANPP of forbs likely due to competitive release. The capability of these very different grass-dominated systems to maintain ANPP while being grazed has important implications for energy flow, ecosystem function and the trophic dynamics of grasslands.     

Managed Wildfire Effects on Forest Resilience and Water in the Sierra Nevada

Fire suppression in many dry forest types has left a legacy of dense, homogeneous forests. Such landscapes have high water demands and fuel loads, and when burned can result in catastrophically large fires. These characteristics are undesirable in the face of projected warming and drying in the western US. Alternative forest and fire treatments based on managed wildfire—a regime in which fires are allowed to burn naturally and only suppressed under defined management conditions—offer a potential strategy to ameliorate the effects of fire suppression. Understanding the long-term effects of this strategy on vegetation, water, and forest resilience is increasingly important as the use of managed wildfire becomes more widely accepted. The Illilouette Creek Basin in Yosemite National Park has experienced 40 years of managed wildfire, reducing forest cover by 22%, and increasing meadow areas by 200% and shrublands by 24%. Statistical upscaling of 3300 soil moisture observations made since 2013 suggests that large increases in wetness occurred in sites where fire caused transitions from forests to dense meadows. The runoff ratio (ratio of annual runoff to precipitation) from the basin appears to be increasing or stable since 1973, compared to declines in runoff ratio for nearby, unburned watersheds. Managed wildfire appears to increase landscape heterogeneity, and likely improves resilience to disturbances, such as fire and drought, although more detailed analysis of fire effects on basin-scale hydrology is needed.     

Evolution of the claustrum in Cnidaria: comparative anatomy reveals that it is exclusive to some species of Staurozoa and absent in Cubozoa

The claustrum in Cnidaria is a tissue in the gastrovascular cavity delimited by a central layer of mesoglea surrounded by gastrodermis (i.e., gastrodermis-mesoglea-gastrodermis), without communication with epidermis. By dividing the gastrovascular cavity, the four claustra provide an additional level of complexity. The presence of claustra in Cubozoa and Staurozoa has been used as evidence supporting a close relationship between these two cnidarian classes. However, the detailed anatomy of the claustrum has never been comparatively analyzed, rendering the evolution of this character among Cnidaria and its homology in Staurozoa and Cubozoa uncertain. This study provides a comparative investigation of the internal anatomy of the claustrum in Staurozoa and Cubozoa, addressing its evolutionary history based on recent phylogenetic hypotheses for Cnidaria. We conclude that the claustrum is a character exclusive to some species of Staurozoa, with a homoplastic evolution in the class, and that the structure called the “claustrum” in Cubozoa corresponds to the valve of gastric ostium, a structure at the base of the manubrium, which is also present in Staurozoa with and without claustrum. Thus, the claustrum cannot be a synapomorphy of a hypothetical clade uniting Staurozoa and Cubozoa, nor can its hypothetical presence in enigmatic fossils be used to support cubozoan affinities.