The amino-terminal half of rotavirus SA114fM VP4 protein contains a hemagglutination domain and primes for neutralizing antibodies to the virus.

We have previously reported the synthesis in Escherichia coli of polypeptide MS2-VP8', which contains the amino-terminal half of the SA114fM VP4 protein fused to MS2 bacteriophage polymerase sequences (C. F. Arias, M. Lizano, and S. López, J. Gen. Virol. 68:633-642, 1987). In this work we have synthesized the carboxy-terminal half of the VP4 protein also fused to the MS2 polymerase. This protein, designated MS2-VP5', was recognized by sera to the complete virion and was able to induce antibodies to the virus when administered to mice; however, these antibodies had no neutralizing activity. The two chimeric polypeptides were tested for their ability to agglutinate erythrocytes and to prime the immune system of mice. Bacterial lysates enriched for the MS2-VP8' hybrid polypeptide, but not those enriched for the MS2-VP5' protein or those containing proteins from the host E. coli strain, had hemagglutinating activity. This hemagglutination was inhibited by sera to SA114fM rotavirus. In addition, a single dose of the MS2-VP8' polypeptide was able to prime the immune system of mice for an augmented neutralizing antibody response when the animals were subsequently immunized with purified SA114fM virus.     

Interaction of the Human Papillomavirus E6 Oncoprotein with Sorting Nexin 27 Modulates Endocytic Cargo Transport Pathways

A subset of high-risk Human Papillomaviruses (HPVs) are the causative agents of a large number of human cancers, of which cervical is the most common. Two viral oncoproteins, E6 and E7, contribute directly towards the development and maintenance of malignancy. A characteristic feature of the E6 oncoproteins from cancer-causing HPV types is the presence of a PDZ binding motif (PBM) at its C-terminus, which confers interaction with cellular proteins harbouring PDZ domains. Here we show that this motif allows E6 interaction with Sorting Nexin 27 (SNX27), an essential component of endosomal recycling pathways. This interaction is highly conserved across E6 proteins from multiple high-risk HPV types and is mediated by a classical PBM-PDZ interaction but unlike many E6 targets, SNX27 is not targeted for degradation by E6. Rather, in HPV-18 positive cell lines the association of SNX27 with components of the retromer complex and the endocytic transport machinery is altered in an E6 PBM-dependent manner. Analysis of a SNX27 cargo, the glucose transporter GLUT1, reveals an E6-dependent maintenance of GLUT1 expression and alteration in its association with components of the endocytic transport machinery. Furthermore, knockdown of E6 in HPV-18 positive cervical cancer cells phenocopies the loss of SNX27, both in terms of GLUT1 expression levels and its vesicular localization, with a concomitant marked reduction in glucose uptake, whilst loss of SNX27 results in slower cell proliferation in low nutrient conditions. These results demonstrate that E6 interaction with SNX27 can alter the recycling of cargo molecules, one consequence of which is modulation of nutrient availability in HPV transformed tumour cells.     

Fifty years of geomorphologic change in Damas Island, Quepos, Costa Rica

Fifty years of geomorphologic change in Damas Island, Quepos, Costa Rica, were studied from a photographic record that is available since 1947. Coastal dynamics were accelerated by the El Ni?o Phenomenon in 1997 which was simultaneous with the August-September astronomical tide, one of the highest in the 4-5 year cycle. Additionally, waves with high energy were present in some periods of these months. Processes were enough to break the island in two blocks and to initialize erosion and transport sediment that continues to date. The frequency of tropical storms and the wave energy will be greater in the next years increasing sediment instability processes in parts of the island. Two topographic profiles have shown that the island is not in equilibrium and that adding all the possible mareographic components it will be prone to continued erosion. The marine habitats around the island should be changing because the fresh and salt water input has been modified, specially because alteration in the Parrita and Paquita hydrological river basins, and its effects on the sediments of this system.     

Activation of heterotrimeric G-protein signaling by a ras-related protein. Implications for signal integration

Utilizing a functional screen in the yeast Saccharomyces cerevisiae we identified mammalian proteins that activate heterotrimeric G-protein signaling pathways in a receptor-independent fashion. One of the identified activators, termed AGS1 (for activator of G-protein signaling), is a human Ras-related G-protein that defines a distinct subgroup of the Ras superfamily. Expression of AGS1 in yeast and in mammalian cells results in specific activation of Galpha(i)/Galpha(o) heterotrimeric signaling pathways. In addition, the in vivo and in vitro properties of AGS1 are consistent with it functioning as a direct guanine nucleotide exchange factor for Galpha(i)/Galpha(o). AGS1 thus presents a unique mechanism for signal integration via heterotrimeric G-protein signaling pathways.     

Effect of preservatives on IgG aggregation, complement-activating effect and hypotensive activity of horse polyvalent antivenom used in snakebite envenomation.

Intravenous administration of antivenoms is associated with early adverse reactions in a number of cases, but the causes of this phenomenon are still unclear. The effect of preservatives (phenol and thimerosal) on IgG aggregate and dimer formation, in vitro complement-activating effect and hypotensive activity of a whole IgG horse liquid polyvalent antivenom, produced by caprylic acid fractionation, was assessed. These parameters were studied since they have been associated with the development of early adverse reactions to the administration of antivenoms and human immunoglobulins. After a three-year storage period at 4 degrees C, antivenoms with preservatives had an increased content of IgG aggregates and dimers when compared with antivenom devoid of phenol and thimerosal. These observations correlate with a slight increment in the turbidity of preservative-containing antivenoms. The three antivenoms studied (formulation: no preservatives; with phenol and thimerosal; with thimerosal alone) activated human complement in vitro, with only minor quantitative differences among them. When antivenoms were administered as a bolus intravenous injection in rats, a rapid and prominent hypotension of short duration was observed after injection of phenol-containing antivenom, whereas such an effect was absent in antivenom free of preservative and in the one containing only thimerosal. Bolus injection of saline solution with phenol resulted in a similar hypotension, indicating that the effect is due to phenol. However, when phenol-containing antivenom was diluted 1:5 with saline solution before infusion, as occurs in the clinical use of this product, no hypotension was observed. Our results stress the need to evaluate the effects of preservatives on the physicochemical and pharmacological characteristics of antivenoms.     

Probing cytochrome P450 bioactivation and fluorescent properties with morpholinyl-tethered anthraquinones

Structural features from the anticancer prodrug nemorubicin (MMDX) and the DNA-binding molecule DRAQ5? were used to prepare anthraquinone-based compounds, which were assessed for their potential to interrogate cytochrome P450 (CYP) functional activity and localisation. 1,4-disubstituted anthraquinone 8 was shown to be 5-fold more potent in EJ138 bladder cancer cells after CYP1A2 bioactivation. In contrast, 1,5-bis((2-morpholinoethyl)amino) substituted anthraquinone 10 was not CYP-bioactivated but was shown to be fluorescent and subsequently photo-activated by a light pulse (at a bandwidth 532–587?nm), resulting in punctuated foci accumulation in the cytoplasm. It also showed low toxicity in human osteosarcoma cells. These combined properties provide an interesting prospective approach for opto-tagging single or a sub-population of cells and seeking their location without the need for continuous monitoring.     

Resource density regulates the foraging investment in higher termite species

1. Resource density can regulate the area that animals use. At low resource density, there is a conflict in terms of balance between costs of foraging and benefits acquired. The foraging of the higher termite Nasutitermes aff. coxipoensis consists of searching throughout trails and a building galleries phase. 2. In this study, a manipulative field experiment was used to test the hypothesis that colonies of N. aff. coxipoensis forage towards a more profitable balance between the establishment of trails and gallery construction at low resource density. 3. The experiment was conducted in north‐eastern Brazil. Seven experimental plots were established with a continuous increase in resource density (sugarcane baits). Entire colonies of N. aff. coxipoensis were transplanted from their original sites to the experimental plot, totalling 35 nests. The number, branches and total length of trails and galleries were quantified. 4. The results show that N. aff. coxipoensis optimises its foraging output, intensifying the establishment of trails at the cost of gallery construction when resource density is low. The number of trails, the number of trail branches and the total length of trails decreased with increasing resource density. Interestingly, at low resource density, the search effort was concentrated on forming longer and a greater number of trails, a small proportion of which were converted into galleries. The opposite relationship was observed at high resource density. 5. These results suggest an optimisation of search efforts during foraging depending on resource density, a mechanism that may help researchers to understand the use of space by higher termite species.     

Detection of PrPsc in Blood from Sheep Infected with the Scrapie and Bovine Spongiform Encephalopathy Agents

The role of blood in the iatrogenic transmission of transmissible spongiform encephalopathy (TSE) or prion disease has become an increasing concern since the reports of variant Creutzfeldt-Jakob disease (vCJD) transmission through blood transfusion from humans with subclinical infection. The development of highly sensitive rapid assays to screen for prion infection in blood is of high priority in order to facilitate the prevention of transmission via blood and blood products. In the present study we show that PrP^sc, a surrogate marker for TSE infection, can be detected in cells isolated from the blood from naturally and experimentally infected sheep by using a rapid ligand-based immunoassay. In sheep with clinical disease, PrP^sc was detected in the blood of 55% of scrapie agent-infected animals (n = 80) and 71% of animals with bovine spongiform encephalopathy (n = 7). PrP^sc was also detected several months before the onset of clinical signs in a subset of scrapie agent-infected sheep, followed from 3 months of age to clinical disease. This study confirms that PrP^sc is associated with the cellular component of blood and can be detected in preclinical sheep by an immunoassay in the absence of in vitro or in vivo amplification.Transmission of variant Creutzfeldt-Jakob disease (vCJD) has been linked with blood transfusion in four reported cases in Great Britain (19, 24, 26, 32), indicating that this is likely to be an efficient route of transmission. Such findings highlight a significant risk to recipients of vCJD-contaminated blood components, and blood services in the United Kingdom have responded by putting in place precautionary measures, including leucodepletion. However, it remains uncertain whether such a procedure is able to remove all prion infectivity. For example, in two studies by Gregori et al. (13, 14) only 42 and 72% of infectivity was removed by leucodepletion from blood from hamsters with scrapie. Therefore, a rapid blood test for vCJD that is able to screen for likely infected blood is critical given that the presymptomatic stages of vCJD are long and that the prevalence of infection in the human population is unknown (6, 9). This knowledge has given rise to concerns that a large-scale vCJD epidemic could occur by human-to-human transmission (16, 21).Infectivity in human blood is consistent with the demonstration of transmission of disease by blood transfusion in sheep incubating both scrapie and experimental BSE infection (17, 18, 20). Transmission was demonstrated from both whole blood and buffy coat fractions from sheep blood, indicating a cellular source of prions although, from studies done in rodent models, it is likely that the plasma fraction also contains infectivity (4, 13, 14). Furthermore, transmission was possible from sheep showing clinical signs and from sheep that were infected but still in the preclinical phase. However, identification of the abnormal prion protein (PrP^sc) in blood as a surrogate marker for infection has proved more elusive (3). Recently, PrP^sc has been amplified from the blood of experimentally infected rodents (5, 25, 28) and from sheep naturally infected with scrapie agent (29) using protein misfolded cyclic amplification (PMCA), but often these studies take days or weeks to complete. Here, we demonstrate, using a ligand-based immunoassay, that PrP^sc is associated with blood leukocytes from sheep with terminal scrapie or bovine spongiform encephalopathy (BSE) and in sheep incubating scrapie prior to the onset of clinical signs. This assay is a modification of a test that has been validated for use as a postmortem test for BSE, scrapie, and chronic wasting disease (CWD) in Europe and the United States (7).