Molecular basis for substrate recognition and septum cleavage by AtlA,the major N-acetylglucosaminidase of Enterococcus faecalis

The cleavage of septal peptidoglycan at the end of cell division facilitates the separation of the two daughter cells. The hydrolases involved in this process (called autolysins) are potentially lethal enzymes that can cause cell death; their activity, therefore, must be tightly controlled during cell growth. In Enterococcus faecalis, the N-acetylglucosaminidase AtlA plays a predominant role in cell separation. atlA mutants form long cell chains and are significantly less virulent in the zebrafish model of infection. The attenuated virulence of atlA mutants is underpinned by a limited dissemination of bacterial chains in the host organism and a more efficient uptake by phagocytes that clear the infection. AtlA has structural homologs in other important pathogens, such as Listeria monocytogenes and Salmonella typhimurium, and therefore represents an attractive model to design new inhibitors of bacterial pathogenesis. Here, we provide a 1.45 Å crystal structure of the E. faecalis AtlA catalytic domain that reveals a closed conformation of a conserved β-hairpin and a complex network of hydrogen bonds that bring two catalytic residues to the ideal distance for an inverting mechanism. Based on the model of the AtlA–substrate complex, we identify key residues critical for substrate recognition and septum cleavage during bacterial growth. We propose that this work will provide useful information for the rational design of specific inhibitors targeting this enterococcal virulence factor and its orthologs in other pathogens.     

The Roles of Sex,Mass and Individual Specialisation in Partitioning Foraging-Depth Niches of a Pursuit-Diving Predator

Intra-specific foraging niche partitioning can arise due to gender differences or individual specialisation in behaviour or prey selection. These may in turn be related to sexual size dimorphism or individual variation in body size through allometry. These variables are often inter-related and challenging to separate statistically. We present a case study in which the effects of sex, body mass and individual specialisation on the dive depths of the South Georgia shag on Bird Island, South Georgia are investigated simultaneously using a linear mixed model. The nested random effects of trip within individual explained a highly significant amount of the variance. The effects of sex and body mass were both significant independently but could not be separated statistically owing to them being strongly interrelated. Variance components analysis revealed that 45.5% of the variation occurred among individuals, 22.6% among trips and 31.8% among Dives, while R² approximations showed gender explained 31.4% and body mass 55.9% of the variation among individuals. Male dive depths were more variable than those of females at the levels of individual, trip and dive. The effect of body mass on individual dive depths was only marginally significant within sexes. The percentage of individual variation in dive depths explained by mass was trivial in males (0.8%) but substantial in females (24.1%), suggesting that differences in dive depths among males was largely due to them adopting different behavioural strategies whereas in females allometry played an additional role. Niche partitioning in the study population therefore appears to be achieved through the interactive effects of individual specialisation and gender upon vertical foraging patch selection, and has the potential to interact in complex ways with other axes of the niche hypervolume such as foraging locations, timing of foraging and diet.     

Silver enhances stamen development in female white campion (Silene latifolia [Caryophyllaceae])

Sex expression in the dioecious plant white campion (Silene latifolia Poiret subsp. alba) appears to be insensitive to exogenous applications of auxins, cytokinins, gibberellic acid, and ethylene; however, silver thiosulfate (Ag(2)S(2)O(3)), an ethylene inhibitor, enhanced stamen development in female white campion. In wild-type females, stamen development is arrested before the microspore mother cells are formed. In contrast, stamens of Ag(2)S(2)O(3)-treated females completed meiosis and produced microspores. Stamen development for these females was incomplete, however, and pollen did not mature. Ag(2)S(2)O(3) stimulated stamen development to the same extent in asexual white campion mutants that retained a Y chromosome but had lost Y-linked genes needed for early stages of stamen development. Although Ag(2)S(2)O(3) can inhibit ethylene signaling, the enhancement of stamen development in female white campion cannot be explained as a loss of ethylene response because no other ethylene inhibitor tested (1-methylcyclopropene, trans-cyclooctene, aminoethoxyvinylglycine, and cobalt chloride) caused stamens to develop in female plants. In addition, application of other metal ions could not enhance stamen development. Therefore, the effect we observed on female white campion was specifically caused by silver ions but not by their action on ethylene signaling.     

Design and effectiveness of a novel trap for capturing nesting songbirds

ABSTRACT Capturing songbirds at their nests can be challenging and time consuming. Although traps designed for capturing songbirds at their nests have been described in the literature, few are effective for capturing species with open‐cup nests. We describe a cylindrical trap designed to capture songbirds at nests up to 2 m above ground in grasses, forbs, shrubs, and small saplings. The nest trap is constructed using a rigid hoop, two pieces of mist net, three stakes, and twist ties. We used this trap to capture female Dickcissels (Spiza americana) and female Indigo Buntings (Passerina cyanea) at their nests, with success rates of 85% (N= 196) and 60–73% (N= 16), respectively. Trapping success was comparable to that using other passerine nest trap designs. Nest abandonment after trapping attempts was rare and similar to that reported in previous studies. Our nest trap is lightweight, easy to make, versatile enough to use in a variety of grassland and shrub habitats, and easily carried and deployed in the field.     

Host plant quality,selection history and trade-offs shape the immune responses of Manduca sexta

Immune defences are an important component of fitness. Yet susceptibility to pathogens is common, suggesting the presence of ecological and evolutionary limitations on immune defences. Here, we use structural equation modelling to quantify the direct effects of resource quality and selection history, and their indirect effects mediated via body condition prior to an immune challenge on encapsulation and melanization immune defences in the tobacco hornworm, Manduca sexta. We also investigate allocation trade-offs among immune defences and growth rate following an immune challenge. We found considerable variation in the magnitude and direction of the direct effects of resource quality and selection history on immune defences and their indirect effects mediated via body condition and allocation trade-offs. Greater resource quality and evolutionary exposure to pathogens had positive direct effects on encapsulation and melanization. The indirect effect of resource quality on encapsulation mediated via body condition was substantial, whereas indirect effects on melanization were negligible. Individuals in better condition prior to the immune challenge had greater encapsulation; however, following the immune challenge, greater encapsulation traded off with slower growth rate. Our study demonstrates the importance of experimentally and analytically disentangling the relative contributions of direct and indirect effects to understand variation in immune defences.     

Cholera toxin promotes the induction of regulatory T cells specific for bystander antigens by modulating dendritic cell activation

It has previously been reported that cholera toxin (CT) is a potent mucosal adjuvant that enhances Th2 or mixed Th1/Th2 type responses to coadministered foreign Ag. Here we demonstrate that CT also promotes the generation of regulatory T (Tr) cells against bystander Ag. Parenteral immunization of mice with Ag in the presence of CT induced T cells that secreted high levels of IL-4 and IL-10 and lower levels of IL-5 and IFN-gamma. Ag-specific CD4(+) T cell lines and clones generated from these mice had cytokine profiles characteristic of Th2 or type 1 Tr cells, and these T cells suppressed IFN-gamma production by Th1 cells. Furthermore, adoptive transfer of bone marrow-derived dendritic cells (DC) incubated with Ag and CT induced T cells that secreted IL-4 and IL-10 and low concentrations of IL-5. It has previously been shown that IL-10 promotes the differentiation or expansion of type 1 Tr cells. Here we found that CT synergized with low doses of LPS to induce IL-10 production by immature DC. CT also enhanced the expression of CD80, CD86, and OX40 (CD134) on DC and induced the secretion of the chemokine, macrophage inflammatory protein-2 (MIP-2), but inhibited LPS-driven induction of CD40 and ICAM-I expression and production of the inflammatory cytokines/chemokines IL-12, TNF-alpha, MIP-1alpha, MIP-1beta, and monocyte chemoattractant protein-1. Our findings suggest that CT induces maturation of DC, but, by inducing IL-10, inhibiting IL-12, and selectively affecting surface marker expression, suppresses the generation of Th1 cells and promotes the induction of T cells with regulatory activity.     

RNA sequencing reveals two major classes of gene expression levels in metazoan cells

The expression level of a gene is often used as a proxy for determining whether the protein or RNA product is functional in a cell or tissue. Therefore, it is of fundamental importance to understand the global distribution of gene expression levels, and to be able to interpret it mechanistically and functionally. Here we use RNA sequencing (RNA‐seq) of mouse Th2 cells, coupled with a range of other techniques, to show that all genes can be separated, based on their expression abundance, into two distinct groups: one group comprised of lowly expressed and putatively non‐functional mRNAs, and the other of highly expressed mRNAs with active chromatin marks at their promoters. These observations are confirmed in many other microarray and RNA‐seq data sets of metazoan cell types.