MicroRNA profiling of the bovine alveolar macrophage response to Mycobacterium bovis infection suggests pathogen survival is enhanced by microRNA regulation of endocytosis and lysosome trafficking

peer-reviewedMycobacterium bovis, the causative agent of bovine tuberculosis, a major problem for global agriculture, spreads via an airborne route and is taken up by alveolar macrophages (AM) in the lung. Here, we describe the first next-generation sequencing (RNA-seq) approach to temporally profile miRNA expression in primary bovine AMs post-infection with M. bovis. One, six, and forty miRNAs were identified as significantly differentially expressed at 2, 24 and 48 h post-infection, respectively. The differential expression of three miRNAs (bta-miR-142-5p, bta-miR-146a, and bta-miR-423-3p) was confirmed by RT-qPCR. Pathway analysis of the predicted mRNA targets of differentially expressed miRNAs suggests that these miRNAs preferentially target several pathways that are functionally relevant for mycobacterial pathogenesis, including endocytosis and lysosome trafficking, IL-1 signalling and the TGF-β pathway. Over-expression studies using a bovine macrophage cell-line (Bomac) reveal the targeting of two key genes in the innate immune response to M. bovis, IL-1 receptor-associated kinase 1 (IRAK1) and TGF-β receptor 2 (TGFBR2), by miR-146. Taken together, our study suggests that miRNAs play a key role in tuning the complex interplay between M. bovis survival strategies and the host immune response

Standardization of synthetic biology tools and assembly methods for Saccharomyces cerevisiae and emerging yeast species

As redesigning organisms using engineering principles is one of the purposes of synthetic biology (SynBio), the standardization of experimental methods and DNA parts is becoming increasingly a necessity. The synthetic biology community focusing on the engineering of Saccharomyces cerevisiae has been in the foreground in this area, conceiving several well-characterized SynBio toolkits widely adopted by the community. In this review, the molecular methods and toolkits developed for S. cerevisiae are discussed in terms of their contributions to the required standardization efforts. In addition, the toolkits designed for emerging nonconventional yeast species including Yarrowia lipolytica, Komagataella phaffii, and Kluyveromyces marxianus are also reviewed. Without a doubt, the characterized DNA parts combined with the standardized assembly strategies highlighted in these toolkits have greatly contributed to the rapid development of many metabolic engineering and diagnostics applications among others. Despite the growing capacity in deploying synthetic biology for common yeast genome engineering works, the yeast community has a long journey to go to exploit it in more sophisticated and delicate applications like bioautomation.ISSN:2161-506

Development of a high sensitivity label free waveguide interferometry instrument : a project of Creoptix GmbH with the center for Biochemistry ZHAW as main research partner

Creoptix GmbH has developed a novel and innovative technology for label-free detection of molecules based on grating-coupled interferometry (GCI). GCI is a proprietary technology characterized by a very high sensitivity at low technical complexity. One main application will be the measurement of binding affinities in research and development projects such as drug discovery. Goal of an interdisciplinary CTI project with the partners from ZHAW, FHNW and CSEM together with Creoptix is the development of a first GCI instrument including disposables and the optimization of its functionality, followed by field tests to pave the way to market introduction

A20 critically controls microglia activation and inhibits inflammasome-dependent neuroinflammation

Microglia, the mononuclear phagocytes of the central nervous system (CNS), are important for the maintenance of CNS homeostasis, but also critically contribute to CNS pathology. Here we demonstrate that the nuclear factor kappa B (NF-kappa B) regulatory protein A20 is crucial in regulating microglia activation during CNS homeostasis and pathology. In mice, deletion of A20 in microglia increases microglial cell number and affects microglial regulation of neuronal synaptic function. Administration of a sublethal dose of lipopolysaccharide induces massive microglia activation, neuroinflammation, and lethality in mice with microgliaconfined A20 deficiency. Microglia A20 deficiency also exacerbates multiple sclerosis (MS) like disease, due to hyperactivation of the NIrp3 inflammasome leading to enhanced interleukin-113 secretion and CNS inflammation. Finally, we confirm a NIrp3 inflammasome signature and IL-1 beta expression in brain and cerebrospinal fluid from MS patients. Collectively, these data reveal a critical role for A20 in the control of microglia activation and neuroinflammation

Decoding human fetal liver haematopoiesis.

Definitive haematopoiesis in the fetal liver supports self-renewal and differentiation of haematopoietic stem cells and multipotent progenitors (HSC/MPPs) but remains poorly defined in humans. Here, using single-cell transcriptome profiling of approximately 140,000 liver and 74,000 skin, kidney and yolk sac cells, we identify the repertoire of human blood and immune cells during development. We infer differentiation trajectories from HSC/MPPs and evaluate the influence of the tissue microenvironment on blood and immune cell development. We reveal physiological erythropoiesis in fetal skin and the presence of mast cells, natural killer and innate lymphoid cell precursors in the yolk sac. We demonstrate a shift in the haemopoietic composition of fetal liver during gestation away from being predominantly erythroid, accompanied by a parallel change in differentiation potential of HSC/MPPs, which we functionally validate. Our integrated map of fetal liver haematopoiesis provides a blueprint for the study of paediatric blood and immune disorders, and a reference for harnessing the therapeutic potential of HSC/MPPs.We acknowledge funding from the Wellcome Human Cell Atlas Strategic Science Support (WT211276/Z/18/Z); M.H. is funded by Wellcome (WT107931/Z/15/Z), The Lister Institute for Preventive Medicine and NIHR and Newcastle-Biomedical Research Centre; S.A.T. is funded by Wellcome (WT206194), ERC Consolidator and EU MRG-Grammar awards and; S.B. is funded by Wellcome (WT110104/Z/15/Z) and St. Baldrick’s Foundation; E.L. is funded by a Wellcome Sir Henry Dale and Royal Society Fellowships, European Haematology Association, Wellcome and MRC to the Wellcome-MRC Cambridge Stem Cell Institute and BBSRC

Gene expression variability across cells and species shapes innate immunity.

As the first line of defence against pathogens, cells mount an innate immune response, which varies widely from cell to cell. The response must be potent but carefully controlled to avoid self-damage. How these constraints have shaped the evolution of innate immunity remains poorly understood. Here we characterize the innate immune response's transcriptional divergence between species and variability in expression among cells. Using bulk and single-cell transcriptomics in fibroblasts and mononuclear phagocytes from different species, challenged with immune stimuli, we map the architecture of the innate immune response. Transcriptionally diverging genes, including those that encode cytokines and chemokines, vary across cells and have distinct promoter structures. Conversely, genes that are involved in the regulation of this response, such as those that encode transcription factors and kinases, are conserved between species and display low cell-to-cell variability in expression. We suggest that this expression pattern, which is observed across species and conditions, has evolved as a mechanism for fine-tuned regulation to achieve an effective but balanced response

MicroRNA regulation of the bovine alveolar macrophage response to Mycobacterium bovis

THESIS 10878.1Bovine tuberculosis is one of the most economically significant infectious diseases of cattle in Ireland and globally, resulting in economic losses of approximately ?2 billion annually. The causative agent is Mycobacterium bovis, a rod-shaped, Gram positive, acid-fast bacterium that is a member of the Mycobacterium tuberculosis complex (MTBC), a group of highly related pathogens that are spread via an airborne route

Selective effects of benzodiazepines on the acquisition of conditioned taste aversion compared to attenuation of neophobia in C57BL/6 mice

The effects of pre-conditioning administration of anxiolytic benzodiazepines on the acquisition of a conditioned taste aversion (CTA) and on the acquisition of attenuation of neophobia (AN) were investigated in C57BL/6 mice.status: publishe

Probing entanglement in Compton interactions

This theoretical research aims to examine areas of the Compton cross section of entangled annihilation photons for the purpose of testing for possible break down of theory, which could have consequences for predicted optimal capabilities of Compton PET systems. We provide maps of the cross section for entangled annihilation photons for experimental verification. We introduce a strategy to derive cross sections in a relatively straight forward manner for the Compton scattering of a hypothetical separable, mixed and entangled states. To understand the effect that entanglement has on the cross section for annihilation photons, we derive the cross section so that it is expressed in terms of the cross section of a hypothetical separable state and of a hypothetical forbidden maximally entangled state. We find lobe-like structures in the cross section which are regions where entanglement has the greatest effect. We also find that mixed states do not reproduce the cross section for annihilation photons, contrary to a recent investigation which reported otherwise. We review the motivation and method of the most precise Compton scattering experiment for annihilation photons, in order to resolve conflicting reports regarding the extent to which the cross section itself has been experimentally verified

Regulation of cerebral endothelial cell morphology by extracellular calcium

Cerebral endothelial cells interconnected by tight and adherens junctions constitute the structural basis of the blood-brain barrier. Extracellular calcium ions have been reported to play an important role in the formation and maintenance of the junctional complex. However, little is known about the action of calcium depletion on the structural characteristics of cerebral endothelial cells. Using atomic force microscopy we analyzed the effect of calcium depletion and readdition on the shape and size of living brain endothelial cells. It was found that the removal of extracellular calcium from confluent cell cultures induced the dissociation of the cells from each other accompanied by an increase in their height. After readdition of calcium a gradual recovery was observed until total confluency was regained. We have also demonstrated that Rho-kinase plays an important role in the calcium-depletion-induced disassembly of endothelial tight and adherens junctions. The Rho-kinase inhibitor Y27632 could prevent the morphological changes induced by a lack of calcium as well. Our results suggest that calcium depletion induces Rho-kinase-dependent cytoskeletal changes that may be partly responsible for the disassembly of the junctional complex