Oesophagostomum dentatum - potential as a model for genomic studies of strongylid nematodes, with biotechnological prospects

There are substantial gaps in the knowledge of the molecular processes of development and reproduction in parasitic nematodes, despite the fact that understanding such processes could lead to novel ways of treating and controlling parasitic diseases, through blocking or disrupting key biological pathways. Biotechnological advances through large-scale sequencing projects, approaches for the analysis of differential gene and protein expression and functional genomics (e.g., double-stranded RNA interference) now provide opportunities to investigate the molecular basis of developmental processes in some parasitic nematodes. The porcine nodule worm, Oesophagostomum dentatum (order Strongylida), may provide a platform for testing the function of genes from this and related nematodes, given that this species can be grown and maintained in culture in vitro for periods longer than other nematodes of the same order. In this article, we review relevant biological, biochemical and molecular biological and genomic information about O. dentatum and propose that the O. dentatum — pig system provides an attractive model for exploring molecular developmental and reproductive processes in strongylid nematodes, leading toward new intervention methods and biotechnological outcomes.13 page(s

A microRNA catalog of the developing chicken embryo identified by a deep sequencing approach

MicroRNA (miRNA) and other types of small regulatory RNAs play a crucial role in the regulation of gene expression in eukaryotes. Several distinct classes of small regulatory RNAs have been discovered in recent years. To extend the repertoire of small regulatory RNAs characterized in chickens we used a deep sequencing approach developed by Solexa (now Illumina Inc.). We sequenced three small RNA libraries prepared from different developmental stages of the chicken embryo (days five, seven, and nine) to produce over 9.5 million short sequence reads. We developed a bioinformatics pipeline to distinguish authentic mature miRNA sequences from other classes of small RNAs and short RNA fragments represented in the sequencing data. Using this approach we detected almost all of the previously known chicken miRNAs and their respective miRNA* sequences. In addition we discovered 449 new chicken miRNAs including 88 miRNA candidates. Of these, 430 miRNAs appear to be specific to the avian lineage. Another six new miRNAs had evidence of evolutionary conservation in at least one vertebrate species outside of the bird lineage. The remaining 13 putative miRNAs appear to represent chicken orthologs of known vertebrate miRNAs. We discovered 39 additional putative miRNA candidates originating from miRNA generating intronic sequences known as mirtrons

VAPB/ALS8 MSP Ligands Regulate Striated Muscle Energy Metabolism Critical for Adult Survival in <i>Caenorhabditis elegans</i>

<div><p>Mutations in VAPB/ALS8 are associated with amyotrophic lateral sclerosis (ALS) and spinal muscular atrophy (SMA), two motor neuron diseases that often include alterations in energy metabolism. We have shown that <i>C. elegans</i> and Drosophila neurons secrete a cleavage product of VAPB, the N-terminal major sperm protein domain (vMSP). Secreted vMSPs signal through Roundabout and Lar-like receptors expressed on striated muscle. The muscle signaling pathway localizes mitochondria to myofilaments, alters their fission/fusion balance, and promotes energy production. Here, we show that neuronal loss of the <i>C. elegans</i> VAPB homolog triggers metabolic alterations that appear to compensate for muscle mitochondrial dysfunction. When vMSP levels drop, cytoskeletal or mitochondrial abnormalities in muscle induce elevated DAF-16, the Forkhead Box O (FoxO) homolog, transcription factor activity. DAF-16 promotes muscle triacylglycerol accumulation, increases ATP levels in adults, and extends lifespan, despite reduced muscle mitochondria electron transport chain activity. Finally, <i>Vapb</i> knock-out mice exhibit abnormal muscular triacylglycerol levels and FoxO target gene transcriptional responses to fasting and refeeding. Our data indicate that impaired vMSP signaling to striated muscle alters FoxO activity, which affects energy metabolism. Abnormalities in energy metabolism of ALS patients may thus constitute a compensatory mechanism counterbalancing skeletal muscle mitochondrial dysfunction.</p></div

Characterisation of novel microRNAs in the Black flying fox (Pteropus alecto) by deep sequencing

Background: Bats are a major source of new and emerging viral diseases. Despite the fact that bats carry and shed highly pathogenic viruses including Ebola, Nipah and SARS, they rarely display clinical symptoms of infection. Host factors influencing viral replication are poorly understood in bats and are likely to include both pre- and post-transcriptional regulatory mechanisms. MicroRNAs are a major mechanism of post-transcriptional gene regulation, however very little is known about them in bats. Results: This study describes 399 microRNAs identified by deep sequencing of small RNA isolated from tissues of the Black flying fox, Pteropus alecto, a confirmed natural reservoir of the human pathogens Hendra virus and Australian bat lyssavirus. Of the microRNAs identified, more than 100 are unique amongst vertebrates, including a subset containing mutations in critical seed regions. Clusters of rapidly-evolving microRNAs were identified, as well as microRNAs predicted to target genes involved in antiviral immunity, the DNA damage response, apoptosis and autophagy. Closer inspection of the predicted targets for several highly supported novel miRNA candidates suggests putative roles in host-virus interaction. Conclusions: MicroRNAs are likely to play major roles in regulating virus-host interaction in bats, via dampening of inflammatory responses (limiting the effects of immunopathology), and directly limiting the extent of viral replication, either through restricting the availability of essential factors or by controlling apoptosis. Characterisation of the bat microRNA repertoire is an essential step towards understanding transcriptional regulation during viral infection, and will assist in the identification of mechanisms that enable bats to act as natural virus reservoirs. This in turn will facilitate the development of antiviral strategies for use in humans and other species.M.R.F. is supported by EMBO Long-Term fellowship ALTF 225–201

DAF-16 localization and activity in wild-type and mutant worms.

<p>(A) Transgenic strains expressing DAF-16::GFP under its endogenous promoter. Transgenic controls raised at 20°C are similar to those raised at 20°C then shifted to 35°C for 30 minutes (see panel B for quantification). Close up images of boxed areas are shown. Anterior is to the left in all panels. Low magnification bar, 50 µm; high magnification bar, 25 µm. (B) Quantification of DAF-16::GFP localization in control (n = 157) and <i>vpr-1(tm1411)</i> mutants (n = 49). (−), incubation under normal growth condition; (+), incubation at 35°C for 30 minutes. (C) Magnified images showing transgenic lines expressing GFP under the <i>sod-3</i> promoter. <i>arx-2</i> encodes Arp2. Arrows indicate vulva muscle region. Anterior is to the left in all panels. Bar, 50 µm.</p

Effect of Arp2/3 inactivation on muscle fat levels.

<p>DIC and fluorescent images of muscle in live 3-day-old hermaphrodite worms fed Bodipy-FAs. <i>arx-2</i> encodes the Arp2 component of the Arp2/3 complex. Arrowheads indicate Bodipy-FA-stained fat droplets. Bar, 5 µm.</p

Effect of <i>Vapb</i> ablation on fasting/refeeding energy metabolism in mice.

<p>(A) TAG concentration in GA muscle and liver of wild-type (+/+) and <i>Vapb</i> knock-out (−/−) mice after 24-hour fasting (red) or 24 hours fasting followed by 6 hours of refeeding (blue). (B and C) Quantitative RT-PCR of indicated genes in liver (B) and TA muscle (C) of wild-type (+/+) and <i>Vapb</i> knock-out (−/−) mice after 24-hour fasting (red) or 24 hours fasting followed by 6 hours of refeeding (blue). Relative mRNA levels are shown on the Y-axis. #, <i>P</i><0.05 compared to fed mice of the same genotype. <i>*</i>, <i>P</i><0.05 compared to +/+ under the same condition.</p