Yolk sac, but not hematopoietic stem cell-derived progenitors, sustain erythropoiesis throughout murine embryonic life.

In the embryo, the first hematopoietic cells derive from the yolk sac and are thought to be rapidly replaced by the progeny of hematopoietic stem cells. We used three lineage-tracing mouse models to show that, contrary to what was previously assumed, hematopoietic stem cells do not contribute significantly to erythrocyte production up until birth. Lineage tracing of yolk sac erythromyeloid progenitors, which generate tissue resident macrophages, identified highly proliferative erythroid progenitors that rapidly differentiate after intra-embryonic injection, persisting as the major contributors to the embryonic erythroid compartment.

Core-Modified Coelenterazine Luciferin Analogues: Synthesis and Chemiluminescence Properties.

In this work on the design and studies of luciferins related to the blue-hued coelenterazine, the synthesis of heterocyclic analogues susceptible to produce a photon, possibly at a different wavelength, is undertaken. Here, the synthesis of O-acetylated derivatives of imidazo[1,2-b]pyridazin-3(5 H)-one, imidazo[2,1-f][1,2,4]triazin-7(1 H)-one, imidazo[1,2-a]pyridin-3-ol, imidazo[1,2-a]quinoxalin-1(5 H)-one, benzo[f]imidazo[1,2-a]quinoxalin-3(11 H)-one, imidazo[1',2':1,6]pyrazino[2,3-c]quinolin-3(11 H)-one, and 5,11-dihydro-3 H-chromeno[4,3-e]imidazo[1,2-a]pyrazin-3-one is described thanks to extensive use of the Buchwald-Hartwig N-arylation reaction. The acidic hydrolysis of these derivatives then gave solutions of the corresponding luciferin analogues, which were studied.

PLA2G1B is involved in CD4 anergy and CD4 lymphopenia in HIV-infected patients.

The precise mechanism leading to profound immunodeficiency of HIV-infected patients is still only partially understood. Here, we show that more than 80% of CD4+ T cells from HIV-infected patients have morphological abnormalities. Their membranes exhibited numerous large abnormal membrane microdomains (aMMDs), which trap and inactivate physiological receptors, such as that for IL-7.

Bioluminescence Profiling of NanoKAZ/NanoLuc Luciferase Using a Chemical Library of Coelenterazine Analogues.

We describe here an extensive structure-bioluminescence relationship study of a chemical library of analogues of coelenterazine, using nanoKAZ/NanoLuc, a mutated luciferase originated from the catalytic subunit of the deep-sea shrimp Oplophorus gracilirostris. Out of the 135 O-acetylated precursors that were prepared by using our recently reported synthesis and following their hydrolysis to give solutions of the corresponding luciferins, notable bioluminescence improvements were achieved in comparison with furimazine, which is currently amongst the best substrates of nanoKAZ/NanoLuc. For instance, the rather more lipophilic analogue 8-(2,3-difluorobenzyl)-2-((5-methylfuran-2-yl)methyl)-6-phenylimidazo[1,2-a]pyrazin-3(7H)-one provided a 1.

Real-time analysis of the behaviour of groups of mice via a depth-sensing camera and machine learning.

Preclinical studies of psychiatric disorders use animal models to investigate the impact of environmental factors or genetic mutations on complex traits such as decision-making and social interactions. Here, we introduce a method for the real-time analysis of the behaviour of mice housed in groups of up to four over several days and in enriched environments. The method combines computer vision through a depth-sensing infrared camera, machine learning for animal and posture identification, and radio-frequency identification to monitor the quality of mouse tracking.

Artificial niche microarrays for identifying extrinsic cell-fate determinants.

The complex cellular microenvironment plays an important role in determining cell fate. For example, stem cells located in a microenvironment termed niche integrate a wide variety of extrinsic cues to take distinct fate choices. Capturing this multiple-input/multiple-output system in vitro has proven to be very challenging.

The Use of Combinatorial Hexapeptide Ligand Library (CPLL) in Allergomics.

The recent progress of proteomic protocols led to more efficient protein extraction and concentration procedures to remove nonprotein interfering compounds present in the starting material and to increase the concentration of underrepresented proteins. Combinatorial hexapeptide ligand libraries (CPLL) were recently applied to both plant- and animal-derived tissues for capturing the low- and very low-abundance allergens. Several IgE-binding proteins which were previously absent or poorly represented by using conventional proteomics tools have been detected and characterized through a CPLL-based approach.

Double One-Dimensional Electrophoresis (D1-DE) Adapted for Immunoproteomics.

The classical proteomics approach for the identification of allergen candidates consists on the separation of proteins by high-resolution two-dimensional electrophoresis (2-DE) with subsequent IgE immunoblotting and further analysis of IgE-reactive protein spots with mass spectrometry. In this approach at least two gels most be run. One gel is used for staining and the other is for immunoblotting by antibodies labeled with specific immunostains.

Single-Cell Acoustic Force Spectroscopy: Resolving Kinetics and Strength of T Cell Adhesion to Fibronectin.

Assessing the strength and kinetics of molecular interactions of cells with the extracellular matrix is fundamental to understand cell adhesion processes. Given the relevance of these processes, there is a strong need for physical methods to quantitatively assess the mechanism of cell adhesion at the single-cell level, allowing discrimination of cells with different behaviors. Here we introduce single-cell acoustic force spectroscopy (scAFS), an approach that makes use of acoustic waves to exert controlled forces, up to 1 nN, to hundreds of individual cells in parallel.

Migration of Type III Secretion System Transcriptional Regulators Links Gene Expression to Secretion.

Many plant-pathogenic bacteria of considerable economic importance rely on type III secretion systems (T3SSs) of the Hrc-Hrp 1 family to subvert their plant hosts. T3SS gene expression is regulated through the HrpG and HrpV proteins, while secretion is controlled by the gatekeeper HrpJ. A link between the two mechanisms was so far unknown.

Expansion of the SOS regulon of Vibrio cholerae through extensive transcriptome analysis and experimental validation.

Background: The SOS response is an almost ubiquitous response of cells to genotoxic stresses. The full complement of genes in the SOS regulon for Vibrio species has only been addressed through bioinformatic analyses predicting LexA binding box consensus and in vitro validation. Here, we perform whole transcriptome sequencing from Vibrio cholerae treated with mitomycin C as an SOS inducer to characterize the SOS regulon and other pathways affected by this treatment.

Entry and Disassembly of Large DNA Viruses: Electron Microscopy Leads the Way.

Nucleocytoplasmic large DNA viruses are a steadily growing group of viruses that infect a wide range of hosts and are characterized by large particle dimensions and genome sizes. Understanding how they enter into the host cell and deliver their genome in the cytoplasm is therefore particularly intriguing. Here, we review the current knowledge on the entry of two of the best-characterized nucleocytoplasmic large DNA viruses: the poxvirus Vaccinia virus (VACV) and the giant virus Mimivirus.

SpdC, a novel virulence factor, controls histidine kinase activity in Staphylococcus aureus.

The success of Staphylococcus aureus, as both a human and animal pathogen, stems from its ability to rapidly adapt to a wide spectrum of environmental conditions. Two-component systems (TCSs) play a crucial role in this process. Here, we describe a novel staphylococcal virulence factor, SpdC, an Abi-domain protein, involved in signal sensing and/or transduction.

Rhinoscleroma pathogenesis: The type K3 capsule of Klebsiella rhinoscleromatis is a virulence factor not involved in Mikulicz cells formation.

Rhinoscleroma is a human specific chronic granulomatous infection of the nose and upper airways caused by the Gram-negative bacterium Klebsiella pneumoniae subsp. rhinoscleromatis. Although considered a rare disease, it is endemic in low-income countries where hygienic conditions are poor.

Influenza virus genome reaches the plasma membrane via a modified endoplasmic reticulum and Rab11-dependent vesicles.

Transport of neo-synthesized influenza A virus (IAV) viral ribonucleoproteins (vRNPs) from the nucleus to the plasma membrane involves Rab 11 but the precise mechanism remains poorly understood. We used metal-tagging and immunolabeling to visualize viral proteins and cellular endomembrane markers by electron microscopy of IAV-infected cells. Unexpectedly, we provide evidence that the vRNP components and the Rab11 protein are present at the membrane of a modified, tubulated endoplasmic reticulum (ER) that extends all throughout the cell, and on irregularly coated vesicles (ICVs).

Genome packaging of reovirus is mediated by the scaffolding property of the microtubule network.

Reovirus replication occurs in the cytoplasm of the host cell, in virally induced mini-organelles called virus factories. On the basis of the serotype of the virus, the virus factories can manifest as filamentous (type 1 Lang strain) or globular structures (type 3 Dearing strain). The filamentous factories morphology is dependent on the microtubule cytoskeleton; however, the exact function of the microtubule network in virus replication remains unknown.

Vaccinia virus A11 is required for membrane rupture and viral membrane assembly.

Although most enveloped viruses acquire their membrane from the host by budding or by a wrapping process, collective data argue that nucleocytoplasmic large DNA viruses (NCLDVs) may be an exception. The prototype member of NCLDVs, vaccinia virus (VACV) may induce rupture of endoplasmic-reticulum-derived membranes to build an open-membrane sphere that closes after DNA uptake. This unconventional membrane assembly pathway is also used by at least 3 other members of the NCLDVs.

Identification of a polygalacturonase (Cup s 2) as the major CCD-bearing allergen in Cupressus sempervirens pollen.

As IgE glyco-epitopes, also referred to as cross-reactive carbohydrate determinants (CCDs), can share significant structural homologies between different plants, they are prone to extensive cross-reactivity among allergen pollen extracts. Here, cypress pollen allergens, especially a polygalacturonase (PG), were further characterized using double one-dimensional electrophoresis (D1-DE). The presence of specific IgE directed against CCDs was investigated by bromelain IgE inhibition and concanavalin A binding assays using sera of cypress pollen-sensitized patients.

An Optimized Method for Extracting Bacterial RNA from Mouse Skin Tissue Colonized by .

Bacterial transcriptome analyses during host colonization are essential to decipher the complexity of the relationship between the bacterium and its host. RNA sequencing (RNA-seq) is a promising approach providing valuable information about bacterial adaptation, the host response and, in some cases, mutual tolerance underlying crosstalk, as recently observed in the context of infection. Buruli ulcer is caused by .

Rae1/YacP, a new endoribonuclease involved in ribosome-dependent mRNA decay in .

The PIN domain plays a central role in cellular RNA biology and is involved in processes as diverse as rRNA maturation, mRNA decay and telomerase function. Here, we solve the crystal structure of the Rae1 (YacP) protein of , a founding member of the NYN (Nedd4-BP1/YacP nuclease) subfamily of PIN domain proteins, and identify potential substrates Unexpectedly, degradation of a characterised target mRNA was completely dependent on both its translation and reading frame. We provide evidence that Rae1 associates with the ribosome and cleaves between specific codons of this mRNA Critically, we also demonstrate translation-dependent Rae1 cleavage of this substrate in a purified translation assay Multiple lines of evidence converge to suggest that Rae1 is an A-site endoribonuclease.

The Legionella pneumophila genome evolved to accommodate multiple regulatory mechanisms controlled by the CsrA-system.

The carbon storage regulator protein CsrA regulates cellular processes post-transcriptionally by binding to target-RNAs altering translation efficiency and/or their stability. Here we identified and analyzed the direct targets of CsrA in the human pathogen Legionella pneumophila. Genome wide transcriptome, proteome and RNA co-immunoprecipitation followed by deep sequencing of a wild type and a csrA mutant strain identified 479 RNAs with potential CsrA interaction sites located in the untranslated and/or coding regions of mRNAs or of known non-coding sRNAs.