A simple smiFISH pipeline to quantify mRNA at the single-cell level in 3D.

Techniques allowing the precise quantification of mRNA at the cellular level are essential for understanding biological processes. Here, we present a semi-automated smiFISH (single-molecule inexpensive FISH) pipeline enabling quantification of mRNA in a small number of cells (∼40) in fixed whole mount tissue. We describe steps for sample preparation, hybridization, image acquisition, cell segmentation, and mRNA quantification.

Comparison of lipidome profiles of Caenorhabditis elegans-results from an inter-laboratory ring trial.

Introduction: Lipidomic profiling allows 100s if not 1000s of lipids in a sample to be detected and quantified. Modern lipidomics techniques are ultra-sensitive assays that enable the discovery of novel biomarkers in a variety of fields and provide new insight in mechanistic investigations. Despite much progress in lipidomics, there remains, as for all high throughput "omics" strategies, the need to develop strategies to standardize and integrate quality control into studies in order to enhance robustness, reproducibility, and usability of studies within specific fields and beyond.

A TIR effector mediates immune evasion by targeting UBAP1 and TLR adaptors.

Bacterial pathogens often subvert the innate immune system to establish a successful infection. The direct inhibition of downstream components of innate immune pathways is particularly well documented but how bacteria interfere with receptor proximal events is far less well understood. Here, we describe a Toll/interleukin 1 receptor (TIR) domain-containing protein (PumA) of the multi-drug resistant PA7 strain.

Genome-wide Screen of Pseudomonas aeruginosa in Saccharomyces cerevisiae Identifies New Virulence Factors.

Pseudomonas aeruginosa is a human opportunistic pathogen that causes mortality in cystic fibrosis and immunocompromised patients. While many virulence factors of this pathogen have already been identified, several remain to be discovered. In this respect we set an unprecedented genome-wide screen of a P.

Repression of somatic cell fate in the germline.

Germ cells must transmit genetic information across generations, and produce gametes while also maintaining the potential to form all cell types after fertilization. Preventing the activation of somatic programs is, therefore, crucial to the maintenance of germ cell identity. Studies in Caenorhabditis elegans, Drosophila melanogaster, and mouse have revealed both similarities and differences in how somatic gene expression is repressed in germ cells, thereby preventing their conversion into somatic tissues.

DI-ICR-FT-MS-based high-throughput deep metabotyping: a case study of the Caenorhabditis elegans-Pseudomonas aeruginosa infection model.

In metabolomics there is an ever-growing need for faster and more comprehensive analysis methods to cope with the increasing size of biological studies. Direct-infusion ion-cyclotron-resonance Fourier-transform spectrometry (DI-ICR-FT-MS) is used in non-targeted metabolomics to obtain high-resolution snapshots of the metabolic state of a system. We applied this technology to a Caenorhabditis elegans-Pseudomonas aeruginosa infection model and optimized times needed for cultivation and mass-spectrometric analysis.

The SET-2/SET1 histone H3K4 methyltransferase maintains pluripotency in the Caenorhabditis elegans germline.

Histone H3 Lys 4 methylation (H3K4me) is deposited by the conserved SET1/MLL methyltransferases acting in multiprotein complexes, including Ash2 and Wdr5. Although individual subunits contribute to complex activity, how they influence gene expression in specific tissues remains largely unknown. In Caenorhabditis elegans, SET-2/SET1, WDR-5.

Optimizing a ultrahigh pressure liquid chromatography-time of flight-mass spectrometry approach using a novel sub-2μm core-shell particle for in depth lipidomic profiling of Caenorhabditis elegans.

Lipid profiling or lipidomics is currently applied in many different research fields. It refers to the global analysis of a samples lipid content using different analytical chemistry methods, with mass spectrometry as the mostly employed technology. We developed a comprehensive in-depth analysis method for the lipidome of the soil-dwelling nematode Caenorhabitis elegans, a widely used model organism.

Characterization of a novel two-partner secretion system implicated in the virulence of Pseudomonas aeruginosa.

Pseudomonas aeruginosa is an opportunistic human pathogen implicated in nosocomial infection and infecting people with compromised immune systems such as cystic fibrosis patients. Although multiple genes involved in P. aeruginosa pathogenesis have been characterized, the overall mechanism of virulence is not fully understood.

The Caenorhabditis elegans HP1 family protein HPL-2 maintains ER homeostasis through the UPR and hormesis.

Cellular adaptation to environmental changes and stress relies on a wide range of regulatory mechanisms that are tightly controlled at several levels, including transcription. Chromatin structure and chromatin binding proteins are important factors contributing to the transcriptional response to stress. However, it remains largely unknown to what extent specific chromatin factors influence the response to distinct forms of stress in a developmental context.

Divergent control of two type VI secretion systems by RpoN in Pseudomonas aeruginosa.

Three Type VI Secretion System (T6SS) loci called H1- to H3-T6SS coexist in Pseudomonas aeruginosa. H1-T6SS targets prokaryotic cells whereas H2-T6SS mediates interactions with both eukaryotic and prokaryotic host cells. Little is known about the third system, except that it may be connected to H2-T6SS during the host infection.

The second type VI secretion system of Pseudomonas aeruginosa strain PAO1 is regulated by quorum sensing and Fur and modulates internalization in epithelial cells.

The genome of Pseudomonas aeruginosa PAO1 contains three type VI secretion systems (T6SSs) called H1-, H2-, and H3-T6SS. The H1-T6SS secretes three identified toxins that target other bacteria, providing a fitness advantage for P. aeruginosa, and likely contributes to bacterial pathogenesis in chronic infections.

Identification of a fibronectin-binding domain within the Campylobacter jejuni CadF protein.

The binding of Campylobacter jejuni to fibronectin (Fn), a component of the extracellular matrix, is mediated by a 37 kDa outer membrane protein termed CadF for Campylobacter adhesion to Fn. Previous studies have indicated that C. jejuni binds to Fn on the basolateral surface of T84 human colonic cells.

Staphylococcus aureus svrA: a gene required for virulence and expression of the agr locus.

A Staphylococcus aureus gene originally identified by signature-tagged mutagenesis as being required for virulence was cloned, sequenced and named svrA. Hydropathy profiles revealed that SvrA is likely to be membrane associated, having two regions with six membrane-spanning domains, the regions separated by an extended hydrophilic loop. When compared with the wild-type strain, an svrA mutant expressed greatly reduced amounts of alpha-, beta- and delta-toxins and an increased amount of protein A.