The SARS Coronavirus E Protein Interacts with PALS1 and Alters Tight Junction Formation and Epithelial Morphogenesis

Intercellular tight junctions define epithelial apicobasal polarity and form a physical fence which protects underlying tissues from pathogen invasions. PALS1, a tight junction-associated protein, is a member of the CRUMBS3-PALS1-PATJ polarity complex, which is crucial for the establishment and maintenance of epithelial polarity in mammals. Here we report that the carboxy-terminal domain of the SARS-CoV E small envelope protein (E) binds to human PALS1. Using coimmunoprecipitation and pull-down assays, we show that E interacts with PALS1 in mammalian cells and further demonstrate that the last four carboxy-terminal amino acids of E form a novel PDZ-binding motif that binds to PALS1 PDZ domain. PALS1 redistributes to the ERGIC/Golgi region, where E accumulates, in SARS-CoV–infected Vero E6 cells. Ectopic expression of E in MDCKII epithelial cells significantly alters cyst morphogenesis and, furthermore, delays formation of tight junctions, affects polarity, and modifies the subcellular distribution of PALS1, in a PDZ-binding motif-dependent manner. We speculate that hijacking of PALS1 by SARS-CoV E plays a determinant role in the disruption of the lung epithelium in SARS patients.     

Novel invadopodia components revealed by differential proteomic analysis

When highly invasive cancer cells are cultured on an extracellular matrix substrate, they extend proteolytically active membrane protrusions, termed invadopodia, from their ventral surface into the underlying matrix. Our understanding of the molecular composition of invadopodia has rapidly advanced in the last few years, but is far from complete.To accelerate component discovery, we resorted to a proteomics approach by applying DIfference Gel Electrophoresis (DIGE) to compare invadopodia-enriched sub-cellular fractions with cytosol and cell body membrane fractions and the whole cell lysate. The fractionation procedure was validated through step-by-step monitoring of the enrichment in typical actin-related invadopodia-associated proteins. After statistical analysis, 129 protein spots were selected for peptide mass fingerprinting analysis; of these 76 were successfully identified and found to correspond to 58 proteins belonging to different functional classes including aerobic glycolysis and other metabolic pathways, protein synthesis, degradation and folding, cytoskeletal components and membrane-associated proteins.Finally, validation of a number of identified proteins was carried out by a combination of immuno-blotting on cell fractions and immunofluorescence localization at invadopodia. These results reveal newly identified components of invadopodia and open further avenues to the molecular study of invasive growth behavior of cancer cells.     

Oviposition pattern of the predator Podisus nigrispinus (Heteroptera: Pentatomidae) under different temperatures

The daily reproductive rate of Podisus nigrispinus (Dallas) (Heteroptera: Pentatomidae) fed with Alabama argillacea (Hübner) (Lepidoptera: Noctuidae) larvae was studied at constant temperatures of 20, 23, 25, 28, 30 and 33±0.2°C, relative humidity of 60±10% and photoperiod of L:D 14:10. Daily reproductive rate of P. nigripinus was affected by age of this predator. Each P. nigrispinus female laid 5.3 (20°C) to 19.9 eggs/day (28°C) which developed into 4.3–16.5 nymphs, respectively. Highest daily reproductive rate of P. nigrispinus was recorded at 28 and 30°C for 5–30-day-old females. This predator showed higher daily reproductive rate than its prey A. argillacea at 25°C. It was also able to reproduce at temperatures from 20 to 33°C with maximum daily reproductive rate between 25 and 30°C. These results are important for optimizing mass rearing of P. nigrispinus in the laboratory.     

Social Environment Affects Acquisition and Color of Structural Nuptial Plumage in a Sexually Dimorphic Tropical Passerine

Structural colors result from the physical interaction of light with organic materials of differing refractive indexes organized at nanoscale dimensions to produce significant interference effects. Because color properties emerge from these finely organized nanostructures, the production of structural coloration could respond to environmental factors and be developmentally more plastic than expected, functioning as an indicator of individual quality. However, there are many unknown factors concerning the function and mechanisms regulating structural coloration, especially relative to social environment. We hypothesized that social environment, in the form of competitive settings, can influence the developmental pathways involving production of feather structural coloration. We experimentally assessed the impact of social environment upon body condition, molt and spectral properties of two types of structural color that compose the nuptial plumage in blue-black grassquits: black iridescent plumage and white underwing patches. We manipulated male social environment during nine months by keeping individuals in three treatments: (1) pairs; (2) all-male groups; and (3) male-female mixed groups. All morphological characters and spectral plumage measures varied significantly through time, but only acquisition of nuptial plumage coverage and nuptial plumage color were influenced by social environment. Compared with males in the paired treatment, those in treatments with multiple males molted into nuptial plumage faster and earlier, and their plumage was more UV-purple-shifted. Our results provide experimental evidence that social context strongly influences development and expression of structural plumage. These results emphasize the importance of long-term experimental studies to identify the phenotypic consequences of social dynamics relative to ornament expression.     

A keystone predator controls bacterial diversity in the pitcher-plant (Sarracenia purpurea) microecosystem

The community of organisms inhabiting the water-filled leaves of the carnivorous pitcher-plant Sarracenia purpurea includes arthropods, protozoa and bacteria, and serves as a model system for studies of food web dynamics. Despite the wealth of data collected by ecologists and zoologists on this food web, very little is known about the bacterial assemblage in this microecosystem. We used terminal restriction fragment length polymorphism (T-RFLP) analysis to quantify bacterial diversity within the pitchers as a function of pitcher size, pH of the pitcher fluid and the presence of the keystone predator in this food web, larvae of the pitcher-plant mosquito Wyeomyia smithii. Results were analysed at two spatial scales: within a single bog and across three isolated bogs. Pitchers were sterile before they opened and composition of the bacterial assemblage was more variable between different bogs than within bogs. Measures of bacterial richness and diversity were greater in the presence of W. smithii and increased with increasing pitcher size. Our results suggest that fundamental ecological concepts derived from macroscopic food webs can also be used to predict the bacterial assemblages in pitcher plants.     

Enhanced Slow-Wave EEG Activity and Thermoregulatory Impairment following the Inhibition of the Lateral Hypothalamus in the Rat

Neurons within the lateral hypothalamus (LH) are thought to be able to evoke behavioural responses that are coordinated with an adequate level of autonomic activity. Recently, the acute pharmacological inhibition of LH has been shown to depress wakefulness and promote NREM sleep, while suppressing REM sleep. These effects have been suggested to be the consequence of the inhibition of specific neuronal populations within the LH, i.e. the orexin and the MCH neurons, respectively. However, the interpretation of these results is limited by the lack of quantitative analysis of the electroencephalographic (EEG) activity that is critical for the assessment of NREM sleep quality and the presence of aborted NREM-to-REM sleep transitions. Furthermore, the lack of evaluation of the autonomic and thermoregulatory effects of the treatment does not exclude the possibility that the wake-sleep changes are merely the consequence of the autonomic, in particular thermoregulatory, changes that may follow the inhibition of LH neurons. In the present study, the EEG and autonomic/thermoregulatory effects of a prolonged LH inhibition provoked by the repeated local delivery of the GABA_A agonist muscimol were studied in rats kept at thermoneutral (24°C) and at a low (10°C) ambient temperature (Ta), a condition which is known to depress sleep occurrence. Here we show that: 1) at both Tas, LH inhibition promoted a peculiar and sustained bout of NREM sleep characterized by an enhancement of slow-wave activity with no NREM-to-REM sleep transitions; 2) LH inhibition caused a marked transitory decrease in brain temperature at Ta 10°C, but not at Ta 24°C, suggesting that sleep changes induced by LH inhibition at thermoneutrality are not caused by a thermoregulatory impairment. These changes are far different from those observed after the short-term selective inhibition of either orexin or MCH neurons, suggesting that other LH neurons are involved in sleep-wake modulation.     

Genetic mapping and QTL analysis in European hazelnut (<Emphasis Type="Italic">Corylus avellana</Emphasis> L.)

The European hazelnut (Corylus avellana L.) is the most economically important nut species in the Betulaceae family. Despite the need for new improved hazelnut cultivars, few breeding programs are carried out because of the large plant size, the long lifecycle of the plant, and the expense and time required. To date, there are no reports of maps with quantitative trait loci (QTL) in hazelnut. Our objective in the present study was to identify QTL associated with vegetative traits to allow marker-assisted selection (MAS). An F1 progeny (275 plants) of Tonda Gentile delle Langhe × Merveille de Bollwiller obtained in 2009 was used to develop a QTL linkage map for vigour, sucker habit, and time of bud burst, after three years of observations. A set of 163 plants were analysed with 152 microsatellite markers. A map of 11 linkage groups was obtained, covering 663.1 cM, and 15 QTLs were identified and mapped for the traits examined. Of them, 10 were ‘major’ QTL, including a stably expressed region on LG_02 for leaf bud burst. At least one major QTL for each year underlies the variation in each trait and a clustering of QTL for trunk circumference and suckers/trunk circumference ratio with high inter-trait correlations was observed on LG_05, suggesting a single pleiotropic locus. This research represents an initial step in the future identification of chromosomal regions carrying genes of interest, important for breeding programs and MAS.     

Elastic fiber assembly in the adult mouse pubic symphysis during pregnancy and postpartum

Impairment of pelvic organ support has been described in mice with genetic modifications of the proteins involved in elastogenesis, such as lysyl oxidase-like 1 (LOXL1) and fibulin 5. During pregnancy, elastic fiber-enriched pelvic tissues are modified to allow safe delivery. In addition, the mouse pubic symphysis is remodeled in a hormone-controlled process that entails the modification of the fibrocartilage into an interpubic ligament (IpL) and the relaxation of this ligament. After first parturition, recovery occurs to ensure pelvic tissue homeostasis. Because ligaments are the main supports of the pelvic organs, this study aimed to evaluate elastogenesis in the IpL during mouse pregnancy and postpartum. Accordingly, virgin, pregnant, and postpartum C57BL/6 mice were studied using light, confocal, and transmission electron microscopy as well as Western blots and real-time PCR. Female mice exhibited the separation of the pubic bones and the formation, relaxation, and postpartum recovery of the IpL. By the time the IpL was formed, the elastic fibers had increased in profile length and diameter, and they consisted of small conglomerates of amorphous material distributed among the bundles of microfibrils. Our analyses also indicated that elastin/tropoelastin, fibrillin 1, LOXL1/Loxl1, and fibulin 5 were spatially and temporally regulated, suggesting that these molecules may contribute to the synthesis of new elastic fibers during IpL development. Overall, this work revealed that adult elastogenesis may be important to assure the elasticity of the pelvic girdle during preparation for parturition and postpartum recovery. This finding may contribute to our understanding of pathological processes involving elastogenesis in the reproductive tract.