Regulation of neutrophil senescence by microRNAs

Neutrophils are rapidly recruited to sites of tissue injury or infection, where they protect against invading pathogens. Neutrophil functions are limited by a process of neutrophil senescence, which renders the cells unable to respond to chemoattractants, carry out respiratory burst, or degranulate. In parallel, aged neutrophils also undergo spontaneous apoptosis, which can be delayed by factors such as GMCSF. This is then followed by their subsequent removal by phagocytic cells such as macrophages, thereby preventing unwanted inflammation and tissue damage. Neutrophils translate mRNA to make new proteins that are important in maintaining functional longevity. We therefore hypothesised that neutrophil functions and lifespan might be regulated by microRNAs expressed within human neutrophils. Total RNA from highly purified neutrophils was prepared and subjected to microarray analysis using the Agilent human miRNA microarray V3. We found human neutrophils expressed a selected repertoire of 148 microRNAs and that 6 of these were significantly upregulated after a period of 4 hours in culture, at a time when the contribution of apoptosis is negligible. A list of predicted targets for these 6 microRNAs was generated from http://mirecords.biolead.org and compared to mRNA species downregulated over time, revealing 83 genes targeted by at least 2 out of the 6 regulated microRNAs. Pathway analysis of genes containing binding sites for these microRNAs identified the following pathways: chemokine and cytokine signalling, Ras pathway, and regulation of the actin cytoskeleton. Our data suggest that microRNAs may play a role in the regulation of neutrophil senescence and further suggest that manipulation of microRNAs might represent an area of future therapeutic interest for the treatment of inflammatory disease

Uniformly curated signaling pathways reveal tissue-specific cross-talks and support drug target discovery

Motivation: Signaling pathways control a large variety of cellular processes. However, currently, even within the same database signaling pathways are often curated at different levels of detail. This makes comparative and cross-talk analyses difficult. Results: We present SignaLink, a database containing 8 major signaling pathways from Caenorhabditis elegans, Drosophila melanogaster, and humans. Based on 170 review and approx. 800 research articles, we have compiled pathways with semi-automatic searches and uniform, well-documented curation rules. We found that in humans any two of the 8 pathways can cross-talk. We quantified the possible tissue- and cancer-specific activity of cross-talks and found pathway-specific expression profiles. In addition, we identified 327 proteins relevant for drug target discovery. Conclusions: We provide a novel resource for comparative and cross-talk analyses of signaling pathways. The identified multi-pathway and tissue-specific cross-talks contribute to the understanding of the signaling complexity in health and disease and underscore its importance in network-based drug target selection. Availability: http://SignaLink.orgComment: 9 pages, 4 figures, 2 tables and a supplementary info with 5 Figures and 13 Table

Regulation of ethylene biosynthesis in Festuca novae-zelandiae (Hack.) Cockayne and in Festuca aruninaceae (Schreb.) in response to a water deficit : a thesis presented in partial fulfilment of the requirements for the degree of Master of Science in Plant Biology at Massey University, Palmerston North, New Zealand

Changes in ethylene evolution and the associated biosynthetic enzyme ACC oxidase to a water deficit, were examined in intact leaves of Fostuca novae-zelandiae and F. arundinacea cultivar 'Roa' (syn. Schedonorus phoenix). The aim was to establish a role, or otherwise, for ACC oxidase as a regulator of ethylene biosynthesis in response to a water deficit. While ACC synthase has long been recognised as the major rate-limiting enzyme in ethylene biosynthesis, there is mounting evidence to suggest that ACC oxidase may also regulate the ethylene biosynthesic pathway in higher plants. Leaf tissues from the two species were harvested at regular intervals during the experimental dry-down, and dissected into two leaf zones, regions enclosed by the ligule. comprising the meristematic and elongating leaf zone (the enclosed tissue), and exposed regions composing the mature green leaf zones. Leaf proline content and the rate of leaf elongation (LER) were used as internal and external indicators of physiological changes in response to the water-deficit. Ethylene evolution in response to a water-deficit was found to be tissue-specific in F.arundinacea. In the rapidly expanding leaf zones, i.e. enclosed tissue, ethylene was maintained at levels similar to control tissue. In the mature green regions of leaves, ethylene followed changes in the leaf elongation rate (LER) with observed peaks in ethylene evolution occurnng approximately 48 hours after a rapid decline in the LER. This burst of ethylene was found to precede any accumulation of proline. Increases in the proline content in both leaf zones, only became significant after the ethylene evolution had subsided to below base levels. This stage-specific ethylene evolution in leaves suggests preferential protection of the rapidly expanding leaf cells, an observation that has been documented by other authors. ACO specific enzyme activity was greatest at soil water contents of ca. 9% in the enclosed and 10% in the exposed leaf tissues of F.arundinacea. On further purification of the enzyme, two novel proteins were recognised by polyclonal antibodies in water-stressed leaves of F.arundinacea. A 32 kDa protein was identified in the enclosed leaf tissue and a 37 kDa protein was identified in the exposed leaf tissue, by SDS-PAGE. These proteins eluted from a Mono Q column at different points in the separation process, i.e at salt concentrations of 320-340 and 300-320 mM NaCI respectively, indicating that they may represent two distinct isoforms of the ACO enzyme. Both proteins are active at pH 7.5 with saturating substrate (ACC) and co-substrate (Na ascorbate) concentrations of 1 mM and 30 mM respectively, and co-factor concentrations of 0.02 mM Fe² + and 30 mM NaHCO₃. When compared with results from western analyses, maximum specific enzyme activity correlated well with the water-deficit induced protein from partially purified enclosed leaf tissue, but only loosely with the protein identified in the exposed leaf tissue. The presence of high molecular weight proteins in both the crude and the purrfied (Mono Q) leaf extracts of F.arundinacea together with the novel proteins, suggests that the ACO enzyme in this species may exist as a dimer In F.novae-zelandiae, the presence of high molecular weight molecules m the crude and partially purified (Sephadex G-25) extracts also suggests dimensation of the enzyme in this species. From this study however, it is not possible to establish a clear regulatory role for the ACO enzyme in ethylene biosynthesis in either F.arvndinacea or F.novae-zelandiae While two novel water-deficit-induced proteins were associated with increased ACO activity in purified leaf extracts of F. amndinacea, there was no obvious correlation between ethylene evolution and enzyme activity

Systematic analysis of the Hippo pathway organization and oncogenic alteration in evolution.

The Hippo pathway is a central regulator of organ size and a key tumor suppressor via coordinating cell proliferation and death. Initially discovered in Drosophila, the Hippo pathway has been implicated as an evolutionarily conserved pathway in mammals; however, how this pathway was evolved to be functional from its origin is still largely unknown. In this study, we traced the Hippo pathway in premetazoan species, characterized the intrinsic functions of its ancestor components, and unveiled the evolutionary history of this key signaling pathway from its unicellular origin. In addition, we elucidated the paralogous gene history for the mammalian Hippo pathway components and characterized their cancer-derived somatic mutations from an evolutionary perspective. Taken together, our findings not only traced the conserved function of the Hippo pathway to its unicellular ancestor components, but also provided novel evolutionary insights into the Hippo pathway organization and oncogenic alteration

The role of lipid and lipoprotein metabolism in non-alcoholic fatty liver disease

Due to the epidemic of obesity across the world, nonalcoholic fatty liver disease (NAFLD) has become one of the most prevalent chronic liver disorders in children and adolescents. NAFLD comprises a spectrum of fat-associated liver conditions that can result in end-stage liver disease and the need for liver transplantation. Simple steatosis, or fatty liver, occurs early in NAFLD and may progress to nonalcoholic steatohepatitis, fibrosis and cirrhosis with increased risk of hepatocellular carcinoma. The mechanism of the liver injury in NAFLD is currently thought to be a multiple-hit process where the first hit is an increase in liver fat, followed by multiple additional factors that trigger the inflammatory activity. At the onset of disease, NAFLD is characterized by hepatic triglyceride accumulation and insulin resistance. Liver fat accumulation is associated with increased lipotoxicity from high levels of free fatty acids, free cholesterol and other lipid metabolites. As a consequence, mitochondrial dysfunction with oxidative stress and production of reactive oxygen species and endoplasmic reticulum stress-associated mechanisms, are activated. The present review focuses on the relationship between intra-cellular lipid accumulation and insulin resistance, as well as on lipid and lipoprotein metabolism in NAFLD

Genome-wide gene expression profiling of stress response in a spinal cord clip compression injury model.

BackgroundThe aneurysm clip impact-compression model of spinal cord injury (SCI) is a standard injury model in animals that closely mimics the primary mechanism of most human injuries: acute impact and persisting compression. Its histo-pathological and behavioural outcomes are extensively similar to human SCI. To understand the distinct molecular events underlying this injury model we analyzed global mRNA abundance changes during the acute, subacute and chronic stages of a moderate to severe injury to the rat spinal cord.ResultsTime-series expression analyses resulted in clustering of the majority of deregulated transcripts into eight statistically significant expression profiles. Systematic application of Gene Ontology (GO) enrichment pathway analysis allowed inference of biological processes participating in SCI pathology. Temporal analysis identified events specific to and common between acute, subacute and chronic time-points. Processes common to all phases of injury include blood coagulation, cellular extravasation, leukocyte cell-cell adhesion, the integrin-mediated signaling pathway, cytokine production and secretion, neutrophil chemotaxis, phagocytosis, response to hypoxia and reactive oxygen species, angiogenesis, apoptosis, inflammatory processes and ossification. Importantly, various elements of adaptive and induced innate immune responses span, not only the acute and subacute phases, but also persist throughout the chronic phase of SCI. Induced innate responses, such as Toll-like receptor signaling, are more active during the acute phase but persist throughout the chronic phase. However, adaptive immune response processes such as B and T cell activation, proliferation, and migration, T cell differentiation, B and T cell receptor-mediated signaling, and B cell- and immunoglobulin-mediated immune response become more significant during the chronic phase.ConclusionsThis analysis showed that, surprisingly, the diverse series of molecular events that occur in the acute and subacute stages persist into the chronic stage of SCI. The strong agreement between our results and previous findings suggest that our analytical approach will be useful in revealing other biological processes and genes contributing to SCI pathology

Multiscale metabolic modeling of C4 plants: connecting nonlinear genome-scale models to leaf-scale metabolism in developing maize leaves

C4 plants, such as maize, concentrate carbon dioxide in a specialized compartment surrounding the veins of their leaves to improve the efficiency of carbon dioxide assimilation. Nonlinear relationships between carbon dioxide and oxygen levels and reaction rates are key to their physiology but cannot be handled with standard techniques of constraint-based metabolic modeling. We demonstrate that incorporating these relationships as constraints on reaction rates and solving the resulting nonlinear optimization problem yields realistic predictions of the response of C4 systems to environmental and biochemical perturbations. Using a new genome-scale reconstruction of maize metabolism, we build an 18000-reaction, nonlinearly constrained model describing mesophyll and bundle sheath cells in 15 segments of the developing maize leaf, interacting via metabolite exchange, and use RNA-seq and enzyme activity measurements to predict spatial variation in metabolic state by a novel method that optimizes correlation between fluxes and expression data. Though such correlations are known to be weak in general, here the predicted fluxes achieve high correlation with the data, successfully capture the experimentally observed base-to-tip transition between carbon-importing tissue and carbon-exporting tissue, and include a nonzero growth rate, in contrast to prior results from similar methods in other systems. We suggest that developmental gradients may be particularly suited to the inference of metabolic fluxes from expression data.Comment: 57 pages, 14 figures; submitted to PLoS Computational Biology; source code available at http://github.com/ebogart/fluxtools and http://github.com/ebogart/multiscale_c4_sourc