A Genome-Wide Collection of Mos1 Transposon Insertion Mutants for the C. elegans Research Community

Methods that use homologous recombination to engineer the genome of C. elegans commonly use strains carrying specific insertions of the heterologous transposon Mos1. A large collection of known Mos1 insertion alleles would therefore be of general interest to the C. elegans research community. We describe here the optimization of a semi-automated methodology for the construction of a substantial collection of Mos1 insertion mutant strains. At peak production, more than 5,000 strains were generated per month. These strains were then subject to molecular analysis, and more than 13,300 Mos1 insertions characterized. In addition to targeting directly more than 4,700 genes, these alleles represent the potential starting point for the engineered deletion of essentially all C. elegans genes and the modification of more than 40% of them. This collection of mutants, generated under the auspices of the European NEMAGENETAG consortium, is publicly available and represents an important research resource

On tail trend detection: modeling relative risk

The climate change dispute is about changes over time of environmental characteristics (such as rainfall). Some people say that a possible change is not so much in the mean but rather in the extreme phenomena (that is, the average rainfall may not change much but heavy storms may become more or less frequent). The paper studies changes over time in the probability that some high threshold is exceeded. The model is such that the threshold does not need to be specified, the results hold for any high threshold. For simplicity a certain linear trend is studied depending on one real parameter. Estimation and testing procedures (is there a trend?) are developed. Simulation results are presented. The method is applied to trends in heavy rainfall at 18 gauging stations across Germany and The Netherlands. A tentative conclusion is that the trend seems to depend on whether or not a station is close to the sea.Comment: 38 page

C2-O-sLeX Glycoproteins Are E-Selectin Ligands that Regulate Invasion of Human Colon and Hepatic Carcinoma Cells

Similar to mechanisms of recruitment of activated leukocytes to inflamed tissues, selectins mediate adhesion and extravasation of circulating cancer cells. Our objective was to determine whether sialyl Lewis X modified core 2 O-glycans (C2-O-sLeX) present on colon and hepatic carcinoma cells promote their adhesion and invasion. We examined membrane expression of C2-O-sLeX, selectin binding, invasion of human colon and hepatic carcinoma cell lines, and mRNA levels of alpha-2,3 fucosyltransferase (FucT-III) and core 2 beta-1,6 N-acetylglucosaminyltransferase (C2GnT1) genes, necessary for C2-O-sLeX synthesis, by quantitative reverse-transcriptase (RT) PCR. Synthesis of core 2 branched O-glycans decorated by sLeX is dependent on C2GnT1 function and thus we determined enzyme activity of C2GnT1. The cell lines that expressed C2GnT1 and FucT-III mRNA by quantitative RT-PCR were highly positive for C2-O-sLeX by flow cytometry, and colon carcinoma cells possessed highly active C2GnT1 enzyme. Cells bound avidly to E-selection but not to P- and L-selectin. Gene knock-down of C2GnT1 in colon and hepatic carcinoma cells using short hairpin RNAs (shRNA) resulted in a 40–90% decrease in C2-O-sLeX and a 30–50% decrease in E-selectin binding compared to control cells. Invasion of hepatic and colon carcinoma cells containing C2GnT1 shRNA was significantly reduced compared to control cells in Matrigel assays and C2GnT1 activity was down-regulated in the latter cells. The sLeX epitope was predominantly distributed on core 2 O-glycans on colon and hepatic carcinoma cells. Our findings indicate that C2GnT1 gene expression and the resulting C2-O-sLeX carbohydrates produced mediate the adhesive and invasive behaviors of human carcinomas which may influence their metastatic potential

Hemotin, a regulator of phagocytosis encoded by a small ORF and xonserved across metazoans

Translation of hundreds of small ORFs (smORFs) of less than 100 amino acids has recently been revealed in vertebrates and Drosophila. Some of these peptides have essential and conserved cellular functions. In Drosophila, we have predicted a particular smORF class encoding ~80 aa hydrophobic peptides, which may function in membranes and cell organelles. Here, we characterise hemotin, a gene encoding an 88aa transmembrane smORF peptide localised to early endosomes in Drosophila macrophages. hemotin regulates endosomal maturation during phagocytosis by repressing the cooperation of 14-3-3ζ with specific phosphatidylinositol (PI) enzymes. hemotin mutants accumulate undigested phagocytic material inside enlarged endo-lysosomes and as a result, hemotin mutants have reduced ability to fight bacteria, and hence, have severely reduced life span and resistance to infections. We identify Stannin, a peptide involved in organometallic toxicity, as the Hemotin functional homologue in vertebrates, showing that this novel regulator of phagocytic processing is widely conserved, emphasizing the significance of smORF peptides in cell biology and disease

An overview of NMR-based metabolomics to identify secondary plant compounds involved in host plant resistance

Secondary metabolites provide a potential source for the generation of host plant resistance and development of biopesticides. This is especially important in view of the rapid and vast spread of agricultural and horticultural pests worldwide. Multiple pests control tactics in the framework of an integrated pest management (IPM) programme are necessary. One important strategy of IPM is the use of chemical host plant resistance. Up to now the study of chemical host plant resistance has, for technical reasons, been restricted to the identification of single compounds applying specific chemical analyses adapted to the compound in question. In biological processes however, usually more than one compound is involved. Metabolomics allows the simultaneous detection of a wide range of compounds, providing an immediate image of the metabolome of a plant. One of the most universally used metabolomic approaches comprises nuclear magnetic resonance spectroscopy (NMR). It has been NMR which has been applied as a proof of principle to show that metabolomics can constitute a major advancement in the study of host plant resistance. Here we give an overview on the application of NMR to identify candidate compounds for host plant resistance. We focus on host plant resistance to western flower thrips (Frankliniella occidentalis) which has been used as a model for different plant species

ABCB4 missense mutations D243A, K435T, G535D, I490T, R545C, and S978P significantly impair the lipid floppase and likely predispose to secondary pathologies in the human population

EJA was funded by Barts and the London Charity (award 458/1495). MN was funded by a Medical Research Council Centenary Award