Irf3 polymorphism alters induction of interferon beta in response to Listeria monocytogenes infection

Genetic makeup of the host plays a significant role in the course and outcome of infection. Inbred strains of mice display a wide range of sensitivities to Listeria monocytogenes infection and thus serve as a good model for analysis of the effect of genetic polymorphism. The outcome of L. monocytogenes infection in mice is influenced by the ability of this bacterium to induce expression of interferon beta mRNA, encoded in mouse by the Ifnb1 (interferon beta 1, fibroblast) gene. Mouse strains that lack components of the IFN beta signaling pathway are substantially more resistant to infection. We found that macrophages from the ByJ substrain of the common C57BL/6 inbred strain of mice are impaired in their ability to induce Ifnb1 expression in response to bacterial and viral infections. We mapped the locus that controls differential expression of Ifnb1 to a region on Chromosome 7 that includes interferon regulatory factor 3 (Irf3), which encodes a transcription factor responsible for early induction of Ifnb1 expression. In C57BL/6ByJ mice, Irf3 mRNA was inefficiently spliced, with a significant proportion of the transcripts retaining intron 5. Analysis of the Irf3 locus identified a single base-pair polymorphism and revealed that intron 5 of Irf3 is spliced by the atypical U12-type spliceosome. We found that the polymorphism disrupts a U12-type branchpoint and has a profound effect on the efficiency of splicing of Irf3. We demonstrate that a naturally occurring change in the splicing control element has a dramatic effect on the resistance to L. monocytogenes infection. Thus, the C57BL/6ByJ mouse strain serves as an example of how a mammalian host can counter bacterial virulence strategies by introducing subtle alteration of noncoding sequences

Human metapneumovirus driven IFN-β production antagonizes macrophage transcriptional induction of IL1-β in response to bacterial pathogens

Human metapneumovirus (HMPV) is a pneumovirus that may cause severe respiratory disease in humans. HMPV infection has been found to increase susceptibility to bacterial superinfections leading to increased morbidity and mortality. The molecular mechanisms underlying HMPV-mediated increase in bacterial susceptibility are poorly understood and largely understudied. Type I interferons (IFNs), while critical for antiviral defenses, may often have detrimental effects by skewing the host immune response and cytokine output of immune cells. It is currently unknown if HMPV skews the inflammatory response in human macrophages triggered by bacterial stimuli. Here we report that HMPV pre-infection impacts production of specific cytokines. HMPV strongly suppresses IL-1β transcription in response to LPS or heat-killed Pseudomonas aeruginosa and Streptococcus pneumonia, while enhancing mRNA levels of IL-6, TNF-α and IFN-β. We demonstrate that in human macrophages the HMPV-mediated suppression of IL-1β transcription requires TANK-binding kinase 1 (TBK1) and signaling via the IFN-β-IFNAR axis. Interestingly, our results show that HMPV pre-infection did not impair the LPS-stimulated activation of NF-κB and HIF-1α, transcription factors that stimulate IL-1β mRNA synthesis in human cells. Furthermore, we determined that sequential HMPV-LPS treatment resulted in accumulation of the repressive epigenetic mark H3K27me3 at the IL1B promoter. Thus, for the first time we present data revealing the molecular mechanisms by which HMPV shapes the cytokine output of human macrophages exposed to bacterial pathogens/LPS, which appears to be dependent on epigenetic reprogramming at the IL1B promoter leading to reduced synthesis of IL-1β. These results may improve current understanding of the role of type I IFNs in respiratory disease mediated not only by HMPV, but also by other respiratory viruses that are associated with superinfections.</p

The Classical Pink-Eyed Dilution Mutation Affects Angiogenic Responsiveness

Angiogenesis is the process by which new blood vessels are formed from existing vessels. Mammalian populations, including humans and mice, harbor genetic variations that alter angiogenesis. Angiogenesis-regulating gene variants can result in increased susceptibility to multiple angiogenesis-dependent diseases in humans. Our efforts to dissect the complexity of the genetic diversity that regulates angiogenesis have used laboratory animals due to the availability of genome sequence for many species and the ability to perform high volume controlled breeding. Using the murine corneal micropocket assay, we have observed more than ten-fold difference in angiogenic responsiveness among various mouse strains. This degree of difference is observed with either bFGF or VEGF induced corneal neovascularization. Ongoing mapping studies have identified multiple loci that affect angiogenic responsiveness in several mouse models. In this study, we used F2 intercrosses between C57BL/6J and the 129 substrains 129P1/ReJ and 129P3/J, as well as the SJL/J strain, where we have identified new QTLs that affect angiogenic responsiveness. In the case of AngFq5, on chromosome 7, congenic animals were used to confirm the existence of this locus and subcongenic animals, combined with a haplotype-based mapping approach that identified the pink-eyed dilution mutation as a candidate polymorphism to explain AngFq5. The ability of mutations in the pink-eyed dilution gene to affect angiogenic response was demonstrated using the p-J allele at the same locus. Using this allele, we demonstrate that pink-eyed dilution mutations in Oca2 can affect both bFGF and VEGF-induced corneal angiogenesis

The Histone Demethylase Jhdm1a Regulates Hepatic Gluconeogenesis

Hepatic gluconeogenesis is required for maintaining blood glucose homeostasis; yet, in diabetes mellitus, this process is unrestrained and is a major contributor to fasting hyperglycemia. To date, the impacts of chromatin modifying enzymes and chromatin landscape on gluconeogenesis are poorly understood. Through catalyzing the removal of methyl groups from specific lysine residues in the histone tail, histone demethylases modulate chromatin structure and, hence, gene expression. Here we perform an RNA interference screen against the known histone demethylases and identify a histone H3 lysine 36 (H3K36) demethylase, Jhdm1a, as a key negative regulator of gluconeogenic gene expression. In vivo, silencing of Jhdm1a promotes liver glucose synthesis, while its exogenous expression reduces blood glucose level. Importantly, the regulation of gluconeogenesis by Jhdm1a requires its demethylation activity. Mechanistically, we find that Jhdm1a regulates the expression of a major gluconeogenic regulator, C/EBPα. This is achieved, at least in part, by its USF1-dependent association with the C/EBPα promoter and its subsequent demethylation of dimethylated H3K36 on the C/EBPα locus. Our work provides compelling evidence that links histone demethylation to transcriptional regulation of gluconeogenesis and has important implications for the treatment of diabetes

Human metapneumovirus driven IFN-β production antagonizes macrophage transcriptional induction of IL1-β in response to bacterial pathogens

Human metapneumovirus (HMPV) is a pneumovirus that may cause severe respiratory disease in humans. HMPV infection has been found to increase susceptibility to bacterial superinfections leading to increased morbidity and mortality. The molecular mechanisms underlying HMPV-mediated increase in bacterial susceptibility are poorly understood and largely understudied. Type I interferons (IFNs), while critical for antiviral defenses, may often have detrimental effects by skewing the host immune response and cytokine output of immune cells. It is currently unknown if HMPV skews the inflammatory response in human macrophages triggered by bacterial stimuli. Here we report that HMPV pre-infection impacts production of specific cytokines. HMPV strongly suppresses IL-1β transcription in response to LPS or heat-killed Pseudomonas aeruginosa and Streptococcus pneumonia, while enhancing mRNA levels of IL-6, TNF-α and IFN-β. We demonstrate that in human macrophages the HMPV-mediated suppression of IL-1β transcription requires TANK-binding kinase 1 (TBK1) and signaling via the IFN-β-IFNAR axis. Interestingly, our results show that HMPV pre-infection did not impair the LPS-stimulated activation of NF-κB and HIF-1α, transcription factors that stimulate IL-1β mRNA synthesis in human cells. Furthermore, we determined that sequential HMPV-LPS treatment resulted in accumulation of the repressive epigenetic mark H3K27me3 at the IL1B promoter. Thus, for the first time we present data revealing the molecular mechanisms by which HMPV shapes the cytokine output of human macrophages exposed to bacterial pathogens/LPS, which appears to be dependent on epigenetic reprogramming at the IL1B promoter leading to reduced synthesis of IL-1β. These results may improve current understanding of the role of type I IFNs in respiratory disease mediated not only by HMPV, but also by other respiratory viruses that are associated with superinfections

Exploitation of conserved eukaryotic host cell farnesylation machinery by an F-box effector of Legionella pneumophila

Farnesylation involves covalent linkage of eukaryotic proteins to a lipid moiety to anchor them into membranes, which is essential for the biological function of Ras and other proteins. A large cadre of bacterial effectors is injected into host cells by intravacuolar pathogens through elaborate type III–VII translocation machineries, and many of these effectors are incorporated into the pathogen-containing vacuolar membrane by unknown mechanisms. The Dot/Icm type IV secretion system of Legionella pneumophila injects into host cells the F-box effector Ankyrin B (AnkB), which functions as platforms for the docking of polyubiquitinated proteins to the Legionella-containing vacuole (LCV) to enable intravacuolar proliferation in macrophages and amoeba. We show that farnesylation of AnkB is indispensable for its anchoring to the cytosolic face of the LCV membrane, for its biological function within macrophages and Dictyostelium discoideum, and for intrapulmonary proliferation in mice. Remarkably, the protein farnesyltransferase, RCE-1 (Ras-converting enzyme-1), and isoprenyl cysteine carboxyl methyltransferase host farnesylation enzymes are recruited to the LCV in a Dot/Icm-dependent manner and are essential for the biological function of AnkB. In conclusion, this study shows novel localized recruitment of the host farnesylation machinery and its anchoring of an F-box effector to the LCV membrane, and this is essential for biological function in vitro and in vivo

A literature-based similarity metric for biological processes

BACKGROUND: Recent analyses in systems biology pursue the discovery of functional modules within the cell. Recognition of such modules requires the integrative analysis of genome-wide experimental data together with available functional schemes. In this line, methods to bridge the gap between the abstract definitions of cellular processes in current schemes and the interlinked nature of biological networks are required. RESULTS: This work explores the use of the scientific literature to establish potential relationships among cellular processes. To this end we haveused a document based similarity method to compute pair-wise similarities of the biological processes described in the Gene Ontology (GO). The method has been applied to the biological processes annotated for the Saccharomyces cerevisiae genome. We compared our results with similarities obtained with two ontology-based metrics, as well as with gene product annotation relationships. We show that the literature-based metric conserves most direct ontological relationships, while reveals biologically sounded similarities that are not obtained using ontology-based metrics and/or genome annotation. CONCLUSION: The scientific literature is a valuable source of information from which to compute similarities among biological processes. The associations discovered by literature analysis are a valuable complement to those encoded in existing functional schemes, and those that arise by genome annotation. These similarities can be used to conveniently map the interlinked structure of cellular processes in a particular organism

FTS and 2-DG induce pancreatic cancer cell death and tumor shrinkage in mice

The Ras inhibitor S-trans-trans farnesylthiosalicylic acid (FTS) inhibits active Ras, which controls cell proliferation, differentiation, survival, and metabolism. FTS also inhibits HIF1α expression in cancer cells, leading to an energy crisis. The synthetic glucose analog 2-deoxy-D-glucose (2-DG), which inhibits glycolysis, is selectively directed to tumor cells that exhibit increased glucose consumption. The 2-DG enters tumor cells, where it competes with glucose for glycolytic enzymes. In cancer models, as well as in human phase 1 trials, 2-DG inhibits tumor growth without toxicity. We postulated that under normoxic conditions, tumor cells treated with FTS would be more sensitive than normal cells to 2-DG. We show here that combined treatment with FTS and 2-DG inhibited cancer cell proliferation additively, yet induced apoptotic cell death synergistically both in vitro and in vivo. The induced apoptosis was inferred from QVD-OPH inhibition, an increase in cleaved caspase 3, and loss of survivin. FTS and 2-DG when combined, but not separately, also induced an increase in fibrosis of the tumor tissue, chronic inflammation, and tumor shrinkage. Overall, these results suggest a possible new treatment of pancreatic tumors by the combined administration of FTS and 2-DG, which together induce pancreatic tumor cell death and tumor shrinkage under non-toxic conditions