TNF and PGE2 in human monocyte-derived macrophages infected with Chlamydia trachomatis

In this study levels of prostaglandin E2 (PGE2), tumour necrosis factor (TNF) and interleukin-1 (IL-1) alpha in medium from monocyte derived macrophages (MdM) infected with Chlamydia trachomatis (L2/434/Bu or K biovars). TNF and PGE2 were found in both cases while IL-1 alpha was not detected. Both TNF and PGE2 levels were higher in the medium of the MdM infected with K biovars. TNF reached maximum levels 24 h postinfection, and then declined, while PGE2 levels increased continuously during the infection time up to 96 h post-infection. Addition of dexamethasone inhibited production of TNF and PGE2. Inhibition of PGE2 production by indomethacin resulted in increased production of TNF, while addition of PGE2 caused partial inhibition of TNF production from infected MdM

The RNA-binding protein Arrest (Bruno) regulates alternative splicing to enable myofibril maturation in Drosophila flight muscle.

In Drosophila, fibrillar flight muscles (IFMs) enable flight, while tubular muscles mediate other body movements. Here, we use RNA-sequencing and isoform-specific reporters to show that spalt major (salm) determines fibrillar muscle physiology by regulating transcription and alternative splicing of a large set of sarcomeric proteins. We identify the RNA-binding protein Arrest (Aret, Bruno) as downstream of salm. Aret shuttles between the cytoplasm and nuclei and is essential for myofibril maturation and sarcomere growth of IFMs. Molecularly, Aret regulates IFM-specific splicing of various salm-dependent sarcomeric targets, including Stretchin and wupA (TnI), and thus maintains muscle fiber integrity. As Aret and its sarcomeric targets are evolutionarily conserved, similar principles may regulate mammalian muscle morphogenesis

Prevalence of head lice and hygiene practices among women over twelve years of age in Sindh, Balochistan, and North West Frontier Province: National Health Survey of Pakistan, 1990-1994

Background: Head lice infestation is an infection of the scalp and skin which causes blood loss, discomfort, and social and psychological distress with the possibility of secondary bacterial infections occurring at scratch sites. In Pakistan, although some small scale studies have been conducted to investigate prevalence of head lice in school children and the general population, no population based estimates have been reported. The National Health Survey of Pakistan (NHSP 1990 - 94) was a nationally representative health examination survey of the Pakistani population. The NHSP is the first population based household survey to collect data on the prevalence of head lice in adult women in Pakistan. In this paper we use data from the NHSP to present an epidemiological profile of personal hygiene practices and head lice infestation among women aged 12 years or older in three provinces of Pakistan, Balochistan, Sindh and North West Frontier Province (NWFP). Results: Overall about 7% women aged 12 years and older suffered from head lice infestation. Multivariable logistic regression analysis identified factors independently associated with presence of head lice. Age less than 16 years and crowding at home were associated with higher infestation-rates. The impact of household socio-economic status on infestation rates among women was different in urban and rural settings, urban women with low socio-economic status were more vulnerable than similar women in rural settings. Bathing infrequently in summer was associated with higher prevalence rates only in Sindh, possibly due to the fact that among the three provinces Sindh has a hotter and more humid summer. Conclusions: The results of our analysis of NHSP indicate high levels of head lice infestation among girls and women in the three Provinces. The epidemiological profile of hygienic practices of women indicated that NWFP and Balochistan as compared to Sindh, and rural as compared to urban areas were less developed with respect to access to water supply and soap for maintaining personal hygiene. Simple and cost-effective measures such as provision of water and soap, and improving awareness regarding maintaining personal hygiene can contribute significantly towards improving public health status of the women in Pakistan

Genome-Wide Identification of Binding Sites Defines Distinct Functions for Caenorhabditis elegans PHA-4/FOXA in Development and Environmental Response

Transcription factors are key components of regulatory networks that control development, as well as the response to environmental stimuli. We have established an experimental pipeline in Caenorhabditis elegans that permits global identification of the binding sites for transcription factors using chromatin immunoprecipitation and deep sequencing. We describe and validate this strategy, and apply it to the transcription factor PHA-4, which plays critical roles in organ development and other cellular processes. We identified thousands of binding sites for PHA-4 during formation of the embryonic pharynx, and also found a role for this factor during the starvation response. Many binding sites were found to shift dramatically between embryos and starved larvae, from developmentally regulated genes to genes involved in metabolism. These results indicate distinct roles for this regulator in two different biological processes and demonstrate the versatility of transcription factors in mediating diverse biological roles.Molecular and Cellular Biolog

A genome-wide resource for the analysis of protein localisation in Drosophila

The Drosophila genome contains >13000 protein-coding genes, the majority of which remain poorly investigated. Important reasons include the lack of antibodies or reporter constructs to visualise these proteins. Here, we present a genome-wide fosmid library of 10000 GFP-tagged clones, comprising tagged genes and most of their regulatory information. For 880 tagged proteins, we created transgenic lines, and for a total of 207 lines, we assessed protein expression and localisation in ovaries, embryos, pupae or adults by stainings and live imaging approaches. Importantly, we visualised many proteins at endogenous expression levels and found a large fraction of them localising to subcellular compartments. By applying genetic complementation tests, we estimate that about two-thirds of the tagged proteins are functional. Moreover, these tagged proteins enable interaction proteomics from developing pupae and adult flies. Taken together, this resource will boost systematic analysis of protein expression and localisation in various cellular and developmental contexts

Whole Cell Cryo-Electron Tomography Reveals Distinct Disassembly Intermediates of Vaccinia Virus

At each round of infection, viruses fall apart to release their genome for replication, and then reassemble into stable particles within the same host cell. For most viruses, the structural details that underlie these disassembly and assembly reactions are poorly understood. Cryo-electron tomography (cryo-ET), a unique method to investigate large and asymmetric structures at the near molecular resolution, was previously used to study the complex structure of vaccinia virus (VV). Here we study the disassembly of VV by cryo-ET on intact, rapidly frozen, mammalian cells, infected for up to 60 minutes. Binding to the cell surface induced distinct structural rearrangements of the core, such as a shape change, the rearrangement of its surface spikes and de-condensation of the viral DNA. We propose that the cell surface induced changes, in particular the decondensation of the viral genome, are a prerequisite for the subsequent release of the vaccinia DNA into the cytoplasm, which is followed by its cytoplasmic replication. Generally, this is the first study that employs whole cell cryo-ET to address structural details of pathogen-host cell interaction

From RNAi Screens to Molecular Function in Embryonic Stem Cells

The ability of embryonic stem (ES) cells to generate any of the around 220 cell types of the adult body has fascinated scientists ever since their discovery. The capacity to re-program fully differentiated cells into induced pluripotent stem (iPS) cells has further stimulated the interest in ES cell research. Fueled by this interest, intense research has provided new insights into the biology of ES cells in the recent past. The development of large-scale and high throughput RNAi technologies has made it possible to sample the role of every gene in maintaining ES cell identity. Here, we review the RNAi screens performed in ES cells to date and discuss the challenges associated with these large-scale experiments. Furthermore, we provide a perspective on how to streamline the molecular characterization following the initial phenotypic description utilizing bacterial artificial chromosome (BAC) transgenesis

Chasing Migration Genes: A Brain Expressed Sequence Tag Resource for Summer and Migratory Monarch Butterflies (Danaus plexippus)

North American monarch butterflies (Danaus plexippus) undergo a spectacular fall migration. In contrast to summer butterflies, migrants are juvenile hormone (JH) deficient, which leads to reproductive diapause and increased longevity. Migrants also utilize time-compensated sun compass orientation to help them navigate to their overwintering grounds. Here, we describe a brain expressed sequence tag (EST) resource to identify genes involved in migratory behaviors. A brain EST library was constructed from summer and migrating butterflies. Of 9,484 unique sequences, 6068 had positive hits with the non-redundant protein database; the EST database likely represents ∼52% of the gene-encoding potential of the monarch genome. The brain transcriptome was cataloged using Gene Ontology and compared to Drosophila. Monarch genes were well represented, including those implicated in behavior. Three genes involved in increased JH activity (allatotropin, juvenile hormone acid methyltransfersase, and takeout) were upregulated in summer butterflies, compared to migrants. The locomotion-relevant turtle gene was marginally upregulated in migrants, while the foraging and single-minded genes were not differentially regulated. Many of the genes important for the monarch circadian clock mechanism (involved in sun compass orientation) were in the EST resource, including the newly identified cryptochrome 2. The EST database also revealed a novel Na+/K+ ATPase allele predicted to be more resistant to the toxic effects of milkweed than that reported previously. Potential genetic markers were identified from 3,486 EST contigs and included 1599 double-hit single nucleotide polymorphisms (SNPs) and 98 microsatellite polymorphisms. These data provide a template of the brain transcriptome for the monarch butterfly. Our “snap-shot” analysis of the differential regulation of candidate genes between summer and migratory butterflies suggests that unbiased, comprehensive transcriptional profiling will inform the molecular basis of migration. The identified SNPs and microsatellite polymorphisms can be used as genetic markers to address questions of population and subspecies structure

Transcriptome profiling of ontogeny in the acridid grasshopper Chorthippus biguttulus

Acridid grasshoppers (Orthoptera:Acrididae) are widely used model organisms for developmental, evolutionary, and neurobiological research. Although there has been recent influx of orthopteran transcriptomic resources, many use pooled ontogenetic stages obscuring information about changes in gene expression during development. Here we developed a de novo transcriptome spanning 7 stages in the life cycle of the acridid grasshopper Chorthippus biguttulus. Samples from different stages encompassing embryonic development through adults were used for transcriptomic profiling, revealing patterns of differential gene expression that highlight processes in the different life stages. These patterns were validated with semi-quantitative RT-PCR. Embryonic development showed a strongly differentiated expression pattern compared to all of the other stages and genes upregulated in this stage were involved in signaling, cellular differentiation, and organ development. Our study is one of the first to examine gene expression during post-embryonic development in a hemimetabolous insect and we found that only the fourth and fifth instars had clusters of genes upregulated during these stages. These genes are involved in various processes ranging from synthesis of biogenic amines to chitin binding. These observations indicate that post-embryonic ontogeny is not a continuous process and that some instars are differentiated. Finally, genes upregulated in the imago were generally involved in aging and immunity. Our study highlights the importance of looking at ontogeny as a whole and indicates promising directions for future research in orthopteran development