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El-Mowafy M.,Helmholtz Center for Infection Research | El-Mowafy M.,Mansoura University | Bahgat M.M.,National Research Center of Egypt | Bahgat M.M.,Institute for Experimental Infection Research | Bilitewski U.,Helmholtz Center for Infection Research
BMC Microbiology

Background: Microorganisms use two-component signal transduction (TCST) systems to regulate the response of the organism to changes of environmental conditions. Such systems are absent from mammalian cells and are thus of interest as drug targets. Fungal TCST systems are usually composed of a hybrid histidine kinase, comprising the histidine kinase (HisKA) domain and a receiver domain, a histidine phosphotransfer protein and a response regulator. Among the 11 groups of fungal histidine kinases, group III histidine kinases are of particular relevance as they are essential for the activity of different groups of fungicides. A characteristic feature is the N-terminal amino acid repeat domain comprising multiple HAMP domains, of which the function is still largely unknown. In Candida albicans, a fungal human pathogen, three histidine kinases were identified, of which CaNik1p is a group III histidine kinase. Heterologous expression of this protein in Sacchromyces cerevisiae conferred susceptibility to different fungicides. Fungicide activity was associated with phosphorylation of the mitogen activated protein kinase Hog1p. Results: We have constructed mutated versions of CaNik1p, from which either all HAMP domains were deleted (CaNik1pΔHAMP) or in which the histidine kinase or the receiver domains were not-functional. Expression of CaNIK1ΔHAMP in S. cerevisiae led to severe growth inhibition. Normal growth could be restored by either replacing the phosphate-accepting histidine residue in CaNik1pΔHAMP or by expressing CaNIK1ΔHAMP in S. cerevisiae mutants, in which single genes encoding several components of the HOG pathway were deleted. Expression of proteins with non-functional histidine kinase or receiver domains resulted in complete loss of susceptibility to antifungals, such as fludioxonil. Conditions leading to growth inhibition of transformants also led to phosphorylation of the MAP kinase Hog1p. Conclusion: Our results show that functional histidine kinase and receiver domains of CaNik1p were essential for antifungal susceptibility and for activation of the Hog1p. Moreover, for the first time we show that deletion of all HAMP domains from CaNik1p led to activation of Hog1p without an external stimulus. This phenotype was similar to the effects obtained upon treatment with fungicides, as in both cases growth inhibition correlated with Hog1p activation and was dependent on the functionality of the conserved phosphate-accepting histidine residue. © 2013 El-Mowafy et al.; licensee BioMed Central Ltd. Source

Nair S.,Helmholtz Center for Infection Research | Michaelsen-Preusse K.,TU Braunschweig | Finsterbusch K.,Helmholtz Center for Infection Research | Stegemann-Koniszewski S.,Helmholtz Center for Infection Research | And 9 more authors.
PLoS Pathogens

The innate immune system protects cells against invading viral pathogens by the auto- and paracrine action of type I interferon (IFN). In addition, the interferon regulatory factor (IRF)-1 can induce alternative intrinsic antiviral responses. Although both, type I IFN and IRF-1 mediate their antiviral action by inducing overlapping subsets of IFN stimulated genes, the functional role of this alternative antiviral action of IRF-1 in context of viral infections in vivo remains unknown. Here, we report that IRF-1 is essential to counteract the neuropathology of vesicular stomatitis virus (VSV). IFN- and IRF-1-dependent antiviral responses act sequentially to create a layered antiviral protection program against VSV infections. Upon intranasal infection, VSV is cleared in the presence or absence of IRF-1 in peripheral organs, but IRF-1-/- mice continue to propagate the virus in the brain and succumb. Although rapid IFN induction leads to a decline in VSV titers early on, viral replication is re-enforced in the brains of IRF-1-/- mice. While IFN provides short-term protection, IRF-1 is induced with delayed kinetics and controls viral replication at later stages of infection. IRF-1 has no influence on viral entry but inhibits viral replication in neurons and viral spread through the CNS, which leads to fatal inflammatory responses in the CNS. These data support a temporal, non-redundant antiviral function of type I IFN and IRF-1, the latter playing a crucial role in late time points of VSV infection in the brain. © 2014 Nair et al. Source

Volz A.,Ludwig Maximilians University of Munich | Langenmayer M.,Ludwig Maximilians University of Munich | Jany S.,Ludwig Maximilians University of Munich | Jany S.,German Center for Infection Research | And 3 more authors.
Journal of Virology

Immunization with modified vaccinia virus Ankara (MVA) can rapidly protect mice against lethal ectromelia virus (ECTV) infection, serving as an experimental model for severe systemic infections. Importantly, this early protective capacity of MVA vaccination completely depends on virus-specific cytotoxic CD8+ T cell responses. We used MVA vaccination in the mousepox challenge model using ECTV infection to investigate the previously unknown factors required to elicit rapid protective T cell immunity in normal C57BL/6 mice and in mice lacking the interferon alpha/beta receptor (IFNAR-/-). We found a minimal dose of 105 PFU of MVA vaccine fully sufficient to allow robust protection against lethal mousepox, as assessed by the absence of disease symptoms and failure to detect ECTV in organs from vaccinated animals. Moreover, MVA immunization at low dosage also protected IFNAR-/- mice, indicating efficient activation of cellular immunity even in the absence of type I interferon signaling. When monitoring for virus-specific CD8+ T cell responses in mice vaccinated with the minimal protective dose of MVA, we found significantly enhanced levels of antigen-specific T cells in animals that were MVA vaccinated and ECTV challenged compared to mice that were only vaccinated. The initial priming of naive CD8+ T cells by MVA immunization appears to be highly efficient and, even at low doses, mediates a rapid in vivo burst of pathogen-specific T cells upon challenge. Our findings define striking requirements for protective emergency immunization against severe systemic infections with orthopoxviruses. © 2014, American Society for Microbiology. Source

Kalinke U.,Institute for Experimental Infection Research

In mammals, odorants are inhaled through the nose and inside the nasal cavity they trigger olfactory sensory neurons (OSN)  that are located within the olfactory epithelium. OSN project their axons into glomerular structures of the olfactory bulb. There they synapse with dendrites of second-order neurons that project their axons to the olfactory cortex. Thus, olfaction is based on direct interaction of environmental matters with OSN. This poses the question of how neurotropic viruses are prevented from infecting OSN and entering the central nervous system. Recent evidence indicates that upon instillation of neurotropic virus OSN are readily infected. By axonal transport virus reaches the glomerular  layer of the olfactory bulb where it is efficiently curbed by a type I IFN dependent mechanism. In this review local mechanisms limiting virus entry via the olfactory system and virus spread within the CNS are recapitulated in the context of anatomical properties of the olfactory system. Source

Prinz M.,Albert Ludwigs University of Freiburg | Kalinke U.,Institute for Experimental Infection Research
Trends in Molecular Medicine

Type I interferons (IFN-α and IFN-β) were discovered more than five decades ago and are widely used for the treatment of human autoimmune diseases such as multiple sclerosis (MS). Despite their highly beneficial features, the precise mechanism of action remains speculative. Given the frequent side effects of IFN-α/β therapy, understanding its action in an in vivo setting is vital to further improve this therapeutic approach. Major advances in our understanding of the IFN biology have recently been made and are particularly based on the combination of powerful genome-wide expression analysis in humans with gene-targeting techniques available for basic research. The recent discovery of a novel T-cell subset, Th17 cells, sheds new light on type I IFNs in MS. © 2010 Elsevier Ltd. Source

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