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  • 51.
    Bergström, Sven
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Normark, Johan
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Microbiological features distinguishing Lyme disease and relapsing fever spirochetes2018Inngår i: Wiener Klinische Wochenschrift, ISSN 0043-5325, E-ISSN 1613-7671, Vol. 130, nr 15-16, s. 484-490Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The recent proposal of splitting the genus Borrelia into two genera in the newly formed family of Borreliaceae, i.aEuro<overline>e. Borrelia and Borreliella has motivated us to reflect upon how these organisms has been characterized and differentiated. This article therefore aims to take a closer look on the biology and virulence attributes of the two suggested genera, i.aEuro<overline>e. those causing Lyme borreliosis and relapsing fever borreliosis. Both genera have much in common with similar infection biological features. They are both characterized as bacterial zoonoses, transmitted by hematophagous arthropods with almost identical microbiological appearance. Nevertheless, a closer look at the genotypic and phenotypic characteristics clearly reveals several differences that might motivate the suggested split. On the other hand, a change of this well-established classification within the genus Borrelia might impose an economical burden as well as a great confusion in society, including medical and scientific societies as well as the general population.

  • 52.
    Bergström, Sven
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Zückert, Wolfram R
    Structure, function and biogenesis of the Borrelia cell envelope2010Inngår i: Borrelia, molecular biology, host interactions and pathogenesis / [ed] Eds DS Samuels and JD Radolf, Norfolk, UK: Caister Academic Press , 2010, s. 139-166Kapittel i bok, del av antologi (Annet vitenskapelig)
  • 53.
    Bernardo-Garcia, Noelia
    et al.
    Department of Crystallography and Structural Biology, Instituto de Química Física "Rocasolano", CSIC, Madrid, Spain.
    Sanchez-Murcia, Pedro
    Univ Alcala De Henares, Area Farmacol, Dept Ciencias Biomed, Unidad Asociada I D I,CSIC, Madrid, Spain.
    Gago, Federico
    Univ Alcala De Henares, Area Farmacol, Dept Ciencias Biomed, Unidad Asociada I D I,CSIC, Madrid, Spain.
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Hermoso, Juan A.
    CSIC, Inst Quim Fis Rocasolano, Dept Crystallog & Struct Chem, Madrid, Spain.
    Structural Bioinformatics in Broad-Spectrum Racemases: a new path in anti-microbial research2016Inngår i: Current organic chemistry, ISSN 1385-2728, E-ISSN 1875-5348, Vol. 20, nr 11, s. 1222-1231Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    D-amino acids are essential components of the bacterial cell wall and play notable roles in microbiology as regulators, for example in sporulation, biofilm formation or interspecies communication. Racemases are the specific enzymes catalyzing the interconversion of L-amino acids to D-amino acids. While most of racemases are mono-specific, a family of broad-spectrum racemases that can racemize ten of the 19 natural chiral amino acids has been recently reported. These enzymes can interconvert radically different residues such as aliphatic and positively charged residues producing non-canonical D-amino acids. Crystal structures together with bioinformatics allowed identification of the residues defining the molecular footprint in broad-spectrum racemases, the specific features of their active sites and the structural basis of their promiscuity. Here we review the recent knowledge on this family compared with the well established of alanine racemases. This structural information is a prerequisite for the development of novel drugs against the important human pathogens for which broad-spectrum racemases play a key role.

  • 54. Bernardo-Garcia, Noelia
    et al.
    Sánchez-Murcia, Pedro A.
    Espaillat, Akbar
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Martínez-Caballero, Siseth
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Hermoso, Juan A.
    Gago, Federico
    Cold-induced aldimine bond cleavage by Tris in Bacillus subtilis alanine racemase2019Inngår i: Organic and biomolecular chemistry, ISSN 1477-0520, E-ISSN 1477-0539, Vol. 17, nr 17, s. 4350-4358Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Pyridoxal 5'-phosphate (PLP) is a versatile cofactor involved in a large variety of enzymatic processes. Most of PLP-catalysed reactions, such as those of alanine racemases (AlaRs), present a common resting state in which the PLP is covalently bound to an active-site lysine to form an internal aldimine. The crystal structure of BsAlaR grown in the presence of Tris lacks this covalent linkage and the PLP cofactor appears deformylated. However, loss of activity in a Tris buffer only occurred after the solution was frozen prior to carrying out the enzymatic assay. This evidence strongly suggests that Tris can access the active site at subzero temperatures and behave as an alternate racemase substrate leading to mechanism-based enzyme inactivation, a hypothesis that is supported by additional X-ray structures and theoretical results from QM/ MM calculations. Taken together, our findings highlight a possibly underappreciated role for a common buffer component widely used in biochemical and biophysical experiments.

  • 55. Bianchi, Matteo
    et al.
    Niemiec, Maria J
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Siler, Ulrich
    Urban, Constantin F.
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Reichenbach, Janine
    Restoration of anti-Aspergillus defense by neutrophil extracellular traps in human chronic granulomatous disease after gene therapy is calprotectin-dependent2011Inngår i: Journal of Allergy and Clinical Immunology, ISSN 0091-6749, E-ISSN 1097-6825, Vol. 127, nr 5, s. 1243-1252 e.7Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    BACKGROUND: Aspergillus spp infection is a potentially lethal disease in patients with neutropenia or impaired neutrophil function. We showed previously that Aspergillus hyphae, too large for neutrophil phagocytosis, are inhibited by reactive oxygen species-dependent neutrophil extracellular trap (NET) formation. This process is defective in chronic granulomatous disease (CGD) because of impaired phagocyte nicotinamide adenine dinucleotide phosphate (NADPH) oxidase function.

    OBJECTIVE: To determine the antifungal agent and mechanism responsible for reconstitution of Aspergillus growth inhibition within NETs after complementation of NADPH oxidase function by gene therapy (GT) for CGD.

    METHODS: Antifungal activity of free and NET-released calprotectin was assessed by incubation of Aspergillus nidulans with purified calprotectin, induced NETs from human controls, and CGD neutrophils after GT in the presence or absence of Zn(2+) or alpha-S100A9 antibody, and with induced NETs from wild-type or S100A9(-/-) mouse neutrophils.

    RESULTS: We identified the host Zn(2+) chelator calprotectin as a neutrophil-associated antifungal agent expressed within NETs, reversibly preventing A nidulans growth at low concentrations, and leading to irreversible fungal starvation at higher concentrations. Specific antibody-blocking and Zn(2+) addition abolished calprotectin-mediated inhibition of A nidulans proliferation in vitro. The role of calprotectin in anti-Aspergillus defense was confirmed in calprotectin knockout mice.

    CONCLUSION: Reconstituted NET formation by GT for human CGD was associated with rapid cure of pre-existing therapy-refractory invasive pulmonary aspergillosis in vivo, underlining the role of functional NADPH oxidase in NET formation and calprotectin release for antifungal activity. These results demonstrate the critical role of calprotectin in human innate immune defense against Aspergillus infection.

  • 56.
    Billker, Oliver
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet).
    CRISPRing the elephant in the room2018Inngår i: Cell Host and Microbe, ISSN 1931-3128, E-ISSN 1934-6069, Vol. 24, nr 6, s. 754-755Artikkel i tidsskrift (Annet vitenskapelig)
    Abstract [en]

    The importance of guanylyl-cyclases (GCs) in apicomplexa has remained elusive due to the large size of the genes. Two recent studies, including Brown and Sibley, 2018 in this issue of Cell Host & Microbe, make elegant use of genome editing with CRISPR-Cas9 to characterize roles of GCs in Toxoplasma and Plasmodium.

  • 57.
    Binesse, Johan
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Lindgren, Helena
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Lindgren, Lena
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Conlan, Wayne
    Sjöstedt, Anders
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Roles of Reactive Oxygen Species-Degrading Enzymes of Francisella tularensis SCHU S42015Inngår i: Infection and Immunity, ISSN 0019-9567, E-ISSN 1098-5522, Vol. 83, nr 6, s. 2255-2263Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Francisella tularensis is a facultative intracellular bacterium utilizing macrophages as its primary intracellular habitat and is therefore highly capable of resisting the effects of reactive oxygen species (ROS), potent mediators of the bactericidal activity of macrophages. We investigated the roles of enzymes presumed to be important for protection against ROS. Four mutants of the highly virulent SCHU S4 strain with deletions of the genes encoding catalase (katG), glutathione peroxidase (gpx), a DyP-type peroxidase (FTT0086), or double deletion of FTT0086 and katG showed much increased susceptibility to hydrogen peroxide (H2O2) and slightly increased susceptibility to paraquat but not to peroxynitrite (ONOO-) and displayed intact intramacrophage replication. Nevertheless, mice infected with the double deletion mutant showed significantly longer survival than SCHU S4-infected mice. Unlike the aforementioned mutants, deletion of the gene coding for alkyl-hydroperoxide reductase subunit C (ahpC) generated a mutant much more susceptible to paraquat and ONOO- but not to H2O2. It showed intact replication in J774 cells but impaired replication in bone marrow-derived macrophages and in internal organs of mice. The live vaccine strain, LVS, is more susceptible than virulent strains to ROS-mediated killing and possesses a truncated form of FTT0086. Expression of the SCHU S4 FTT0086 gene rendered LVS more resistant to H2O2, which demonstrates that the SCHU S4 strain possesses additional detoxifying mechanisms. Collectively, the results demonstrate that SCHU S4 ROS-detoxifying enzymes have overlapping functions, and therefore, deletion of one or the other does not critically impair the intracellular replication or virulence, although AhpC appears to have a unique function.

  • 58. Birdsell, D. N.
    et al.
    Özsürekci, Y.
    Rawat, A.
    Aycan, A. E.
    Mitchell, C. L.
    Sahl, J. W.
    Johansson, Anders
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Colman, R. E.
    Schupp, J. M.
    Ceyhan, M.
    Keim, P. S.
    Wagner, D. M.
    Coinfections identified from metagenomic analysis of cervical lymph nodes from tularemia patients2018Inngår i: BMC Infectious Diseases, ISSN 1471-2334, E-ISSN 1471-2334, Vol. 18, artikkel-id 319Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Background: Underlying coinfections may complicate infectious disease states but commonly go unnoticed because an a priori clinical suspicion is usually required so they can be detected via targeted diagnostic tools. Shotgun metagenomics is a broad diagnostic tool that can be useful for identifying multiple microbes simultaneously especially if coupled with lymph node aspirates, a clinical matrix known to house disparate pathogens. The objective of this study was to analyze the utility of this unconventional diagnostic approach (shotgun metagenomics) using clinical samples from human tularemia cases as a test model. Tularemia, caused by the bacterium Francisella tularensis, is an emerging infectious disease in Turkey. This disease commonly manifests as swelling of the lymph nodes nearest to the entry of infection. Because swollen cervical nodes are observed from many different types of human infections we used these clinical sample types to analyze the utility of shotgun metagenomics.

    Methods: We conducted an unbiased molecular survey using shotgun metagenomics sequencing of DNA extracts from fine-needle aspirates of neck lymph nodes from eight tularemia patients who displayed protracted symptoms. The resulting metagenomics data were searched for microbial sequences (bacterial and viral).

    Results: F. tularensis sequences were detected in all samples. In addition, we detected DNA of other known pathogens in three patients. Both Hepatitis B virus (HBV) and Human Parvovirus B-19 were detected in one individual and Human Parvovirus B-19 alone was detected in two other individuals. Subsequent PCR coupled with Sanger sequencing verified the metagenomics results. The HBV status was independently confirmed via serological diagnostics, despite evading notice during the initial assessment.

    Conclusion: Our data highlight that shotgun metagenomics of fine-needle lymph node aspirates is a promising clinical diagnostic strategy to identify coinfections. Given the feasibility of the diagnostic approach demonstrated here, further steps to promote integration of this type of diagnostic capability into mainstream clinical practice are warranted.

  • 59. Birgand, G.
    et al.
    Johansson, Anders
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Szilagyi, E.
    Lucet, J. -C
    Overcoming the obstacles of implementing infection prevention and control guidelines2015Inngår i: Clinical Microbiology and Infection, ISSN 1198-743X, E-ISSN 1469-0691, Vol. 21, nr 12, s. 1067-1071Artikkel, forskningsoversikt (Fagfellevurdert)
    Abstract [en]

    Reasons for a successful or unsuccessful implementation of infection prevention and control (IPC) guidelines are often multiple and interconnected. This article reviews key elements from the national to the individual level that contribute to the success of the implementation of IPC measures and gives perspectives for improvement. Governance approaches, modes of communication and formats of guidelines are discussed with a view to improve collaboration and transparency among actors. The culture of IPC influences practices and varies according to countries, specialties and healthcare providers. We describe important contextual aspects, such as relationships between actors and resources and behavioural features including professional background or experience. Behaviour change techniques providing goal-setting, feedback and action planning have proved effective in mobilizing participants and may be key to trigger social movements of implementation. The leadership of international societies in coordinating actions at international, national and institutional levels using multidisciplinary approaches and fostering collaboration among clinical microbiology, infectious diseases and IPC will be essential for success. Clinical Microbiology and Infection (C) 2015 European Society of Clinical Microbiology and Infectious Diseases. 

  • 60.
    Bitar, Aziz
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Immunologi/immunkemi.
    Aung, Kyaw Min
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Immunologi/immunkemi.
    Wai, Sun Nyunt
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Hammarström, Marie-Louise
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Immunologi/immunkemi.
    Vibrio cholerae derived outer membrane vesicles modulate the inflammatory response of human intestinal epithelial cells by inducing microRNA-146a2019Inngår i: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 9, artikkel-id 7212Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The small intestinal epithelium of Vibrio cholerae infected patients expresses the immunomodulatory microRNAs miR-146a and miR-155 at acute stage of disease. V. cholerae release outer membrane vesicles (OMVs) that serve as vehicles for translocation of virulence factors including V. cholerae cytolysin (VCC). The aim was to investigate whether OMVs, with and/or without VCC-cargo could be responsible for induction of microRNAs in intestinal epithelial cells and thereby contribute to immunomodulation. Polarized tight monolayers of T84 cells were challenged with OMVs of wildtype and a VCC deletion mutant of the non-O1/non-O139 (NOVC) V. cholerae strain V:5/04 and with soluble VCC. OMVs, with and without VCC-cargo, caused significantly increased levels of miR-146a. Increase was seen already after 2 hours challenge with OMVs and persisted after 12 hours. Challenge with soluble VCC caused significant increases in interleukin-8 (IL-8), tumour necrosis factor-α (TNF-α), CCL20, IL-1β, and IRAK2 mRNA levels while challenge with OMVs did not cause increases in expression levels of any of these mRNAs. These results suggest that V. cholerae bacteria release OMVs that induce miR-146a in order to pave the way for colonization by reducing the strength of an epithelial innate immune defence reaction and also preventing inflammation in the mucosa that factors like VCC can evoke.

  • 61.
    Bitar, Aziz
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Immunologi/immunkemi.
    De, Rituparna
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Immunologi/immunkemi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Melgar, Silvia
    Aung, Kyaw Min
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Immunologi/immunkemi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Rahman, Arman
    Qadri, Firdausi
    Wai, Sun Nyunt
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Shirin, Tahmina
    Hammarström, Marie-Louise
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Immunologi/immunkemi.
    Induction of immunomodulatory miR-146a and miR-155 in small intestinal epithelium of Vibrio cholerae infected patients at acute stage of cholera2017Inngår i: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 12, nr 3, artikkel-id 0173817Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The potential immunomodulatory role of microRNAs in small intestine of patients with acute watery diarrhea caused by Vibrio cholerae O1 or enterotoxigenic Escherichia coli (ETEC) infection was investigated. Duodenal biopsies were obtained from study-participants at the acute (day 2) and convalescent (day 21) stages of disease, and from healthy individuals. Levels of miR-146a, miR-155 and miR-375 and target gene (IRAK1, TRAF6, CARD10) and 11 cytokine mRNAs were determined by qRT-PCR. The cellular source of microRNAs in biopsies was analyzed by in situ hybridization. The ability of V. cholerae bacteria and their secreted products to cause changes in microRNA- and mRNA levels in polarized tight monolayers of intestinal epithelial cells was investigated. miR-146a and miR-155 were expressed at significantly elevated levels at acute stage of V. cholerae infection and declined to normal at convalescent stage (P<0.009 versus controls; P = 0.03 versus convalescent stage, pairwise). Both microRNAs were mainly expressed in the epithelium. Only marginal down-regulation of target genes IRAK1 and CARD10 was seen and a weak cytokine-profile was identified in the acute infected mucosa. No elevation of microRNA levels was seen in ETEC infection. Challenge of tight monolayers with the wild type V. cholerae O1 strain C6706 and clinical isolates from two study-participants, caused significant increase in miR-155 and miR-146a by the strain C6706 (P<0.01). One clinical isolate caused reduction in IRAK1 levels (P<0.05) and none of the strains induced inflammatory cytokines. In contrast, secreted factors from these strains caused markedly increased levels of IL-8, IL-1β, and CARD10 (P<0.001), without inducing microRNA expression. Thus, miR-146a and miR-155 are expressed in the duodenal epithelium at the acute stage of cholera. The inducer is probably the V. cholerae bacterium. By inducing microRNAs the bacterium can limit the innate immune response of the host, including inflammation evoked by its own secreted factors, thereby decreasing the risk of being eliminated.

  • 62.
    Bitar, Aziz
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi.
    Kyaw, Min Aung
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Marie-Louise, Hammarström
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Immunologi/immunkemi.
    Wai, Sun Nynt
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Vibrio cholerae derived outer membrane vesicles modulate the inflammatory response ofhuman intestinal epithelial cells by inducing microRNA-146aInngår i: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The small intestinal epithelium of Vibrio cholerae infected patients expresses the immunomodulatory microRNAs miR-146a and miR-155 at acute stage of disease. V. cholerae release outer membrane vesicles (OMVs) that serve as vehicles for translocation of virulencefactors including V. cholerae cytolysin (VCC). The aim was to investigate whether OMVs, with and/or without VCC-cargo could be responsible for induction of microRNAs in intestinal epithelial cells and thereby contribute to immunomodulation. Polarized tight monolayers of T84 cells were challenged with OMVs of wild-type and a VCC deletion mutant of the non-O1/non-O139 (NOVC) V. cholerae strain V:5/04 and with soluble VCC. OMVs, with and without VCC-cargo, caused significantly increased levels of miR-146a. Challenge with soluble VCC caused significant increases in interleukin-8 (IL-8), tumour necrosis factor-α(TNF-α), CCL20, IL-1β, and IRAK2 mRNA levels while challenge with OMVs did not causeany changes. Notably, OMVs from the VCC deficient mutant caused significant decreases in CCL20 and IL-18 mRNA levels. These results suggest that V. cholerae bacteria release OMVs that induce miR-146a in order to pave the way for colonization by reducing thestrength of an epithelial innate immune defence reaction and also preventing inflammation inthe mucosa that factors like VCC can evoke.

  • 63.
    Björnfot, Ann-Catrin
    et al.
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet).
    Lavander, Moa
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Infektionssjukdomar.
    Forsberg, Åke
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Wolf-Watz, Hans
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Autoproteolysis of YscU of Yersinia pseudotuberculosis is important for regulation of expression and secretion of Yop proteins2009Inngår i: Journal of Bacteriology, ISSN 0021-9193, E-ISSN 1098-5530, Vol. 191, nr 13, s. 4259-4267Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    YscU of Yersinia can be autoproteolysed to generate a 10-kDa C-terminal polypeptide designated YscU(CC). Autoproteolysis occurs at the conserved N downward arrowPTH motif of YscU. The specific in-cis-generated point mutants N263A and P264A were found to be defective in proteolysis. Both mutants expressed and secreted Yop proteins (Yops) in calcium-containing medium (+Ca(2+) conditions) and calcium-depleted medium (-Ca(2+) conditions). The level of Yop and LcrV secretion by the N263A mutant was about 20% that of the wild-type strain, but there was no significant difference in the ratio of the different secreted Yops, including LcrV. The N263A mutant secreted LcrQ regardless of the calcium concentration in the medium, corroborating the observation that Yops were expressed and secreted in Ca(2+)-containing medium by the mutant. YscF, the type III secretion system (T3SS) needle protein, was secreted at elevated levels by the mutant compared to the wild type when bacteria were grown under +Ca(2+) conditions. YscF secretion was induced in the mutant, as well as in the wild type, when the bacteria were incubated under -Ca(2+) conditions, although the mutant secreted smaller amounts of YscF. The N263A mutant was cytotoxic for HeLa cells, demonstrating that the T3SS-mediated delivery of effectors was functional. We suggest that YscU blocks Yop release and that autoproteolysis is required to relieve this block.

  • 64.
    Boal, Frédéric
    et al.
    INSERM U1048, I2MC and Universite´ Paul Sabatier, 31432 Toulouse, France.
    Puhar, Andrea
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). INSERM U1202, Unite´ de Pathogénie Microbienne Moléculaire, Institut Pasteur, 75724 Paris Cedex 15, France.
    Xuereb, Jean-Marie
    INSERM U1048, I2MC and Universite´ Paul Sabatier, 31432 Toulouse, France.
    Kunduzova, Oksana
    INSERM U1048, I2MC and Universite´ Paul Sabatier, 31432 Toulouse, France.
    Sansonetti, Philippe J.
    INSERM U1202, Unite´ de Pathogénie Microbienne Moléculaire, Institut Pasteur, 75724 Paris Cedex 15, France.
    Payrastre, Bernard
    INSERM U1048, I2MC and Universite´ Paul Sabatier, 31432 Toulouse, France; .
    Tronchére, Héléne
    INSERM U1048, I2MC and Universite´ Paul Sabatier, 31432 Toulouse, France.
    PI5P Triggers ICAM-1 Degradation in Shigella Infected Cells, Thus Dampening Immune Cell Recruitment2016Inngår i: Cell reports, ISSN 2211-1247, E-ISSN 2211-1247, Vol. 14, nr 4, s. 750-759Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Shigella flexneri, the pathogen responsible for bacillary dysentery, has evolved multiple strategies to control the inflammatory response. Here, we show that Shigella subverts the subcellular trafficking of the intercellular adhesion molecule-1 (ICAM-1), a key molecule in immune cell recruitment, in a mechanism dependent on the injected bacterial enzyme IpgD and its product, the lipid mediator PI5P. Overexpression of IpgD, but not a phosphatase dead mutant, induced the internalization and the degradation of ICAM-1 in intestinal epithelial cells. Remarkably, addition of permeant PI5P reproduced IpgD effects and led to the inhibition of neutrophil recruitment. Finally, these results were confirmed in an in vivo model of Shigella infection where IpgD-dependent ICAM-1 internalization reduced neutrophil adhesion. In conclusion, we describe here an immune evasion mechanism used by the pathogen Shigella to divert the host cell trafficking machinery in order to reduce immune cell recruitment.

  • 65.
    Bonde, Mari
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Olofsson, Annelie
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Frost, Mikaela
    Jegerschöld, Caroline
    Karolinska Institutet, Sweden.
    Bergström, Sven
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Sandblad, Linda
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Structural analysis of the B. burgdorferi integral outer membrane protein, P13, in lipid bilayer NanodiscsManuskript (preprint) (Annet (populærvitenskap, debatt, mm))
  • 66.
    Bonde, Mari
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Östberg, Yngve
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Bunikis, Ignas
    Uppsala University.
    Nyunt Wai, Sun
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Bergström, Sven
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Effects of osmotic stress in P13 and P66 deficient Borrelia burgdorferi mutantsManuskript (preprint) (Annet (populærvitenskap, debatt, mm))
  • 67.
    Brodiazhenko, Tetiana
    et al.
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Institute of Technology, University of Tartu, Tartu, Estonia.
    Johansson, Marcus J. O.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet).
    Takada, Hiraku
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Nissan, Tracy
    Hauryliuk, Vasili
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Institute of Technology, University of Tartu, Tartu, Estonia.
    Murina, Victoriia
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Elimination of Ribosome Inactivating Factors Improves the Efficiency of Bacillus subtilis and Saccharomyces cerevisiae Cell-Free Translation Systems2018Inngår i: Frontiers in Microbiology, ISSN 1664-302X, E-ISSN 1664-302X, Vol. 9, artikkel-id 3041Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Cell-free translation systems based on cellular lysates optimized for in vitro protein synthesis have multiple applications both in basic and applied science, ranging from studies of translational regulation to cell-free production of proteins and ribosome-nascent chain complexes. In order to achieve both high activity and reproducibility in a translation system, it is essential that the ribosomes in the cellular lysate are enzymatically active. Here we demonstrate that genomic disruption of genes encoding ribosome inactivating factors – HPF in Bacillus subtilis and Stm1 in Saccharomyces cerevisiae – robustly improve the activities of bacterial and yeast translation systems. Importantly, the elimination of B. subtilis HPF results in a complete loss of 100S ribosomes, which otherwise interfere with disome-based approaches for preparation of stalled ribosomal complexes for cryo-electron microscopy studies.

  • 68. Broglia, Laura
    et al.
    Materne, Solange
    Lecrivain, Anne-Laure
    Hahnke, Karin
    Le Rhun, Anaïs
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Max Planck Unit for the Science of Pathogens, Berlin, Germany; Department of Regulation in Infection Biology, Max Planck Institute for Infection Biology, Berlin, Germany; Department of Regulation in Infection Biology, Helmholtz Centre for Infection Research, Braunschweig, Germany.
    Charpentier, Emmanuelle
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Max Planck Unit for the Science of Pathogens, Berlin, Germany; Department of Regulation in Infection Biology, Max Planck Institute for Infection Biology, Berlin, Germany; Institute for Biology, Humboldt University, Berlin, Germany; Department of Regulation in Infection Biology, Helmholtz Centre for Infection Research, Braunschweig, Germany.
    RNase Y-mediated regulation of the streptococcal pyrogenic exotoxin B2018Inngår i: RNA Biology, ISSN 1547-6286, E-ISSN 1555-8584, Vol. 15, nr 10, s. 1336-1347Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Endoribonuclease Y (RNase Y) is a crucial regulator of virulence in Gram-positive bacteria. In the human pathogen Streptococcus pyogenes, RNase Y is required for the expression of the major secreted virulence factor streptococcal pyrogenic exotoxin B (SpeB), but the mechanism involved in this regulation remains elusive. Here, we demonstrate that the 5′ untranslated region of speB mRNA is processed by several RNases including RNase Y. In particular, we identify two RNase Y cleavage sites located downstream of a guanosine (G) residue. To assess whether this nucleotide is required for RNase Y activity in vivo, we mutated it and demonstrate that the presence of this G residue is essential for the processing of the speB mRNA 5′ UTR by RNase Y. Although RNase Y directly targets and processes speB, we show that RNase Y-mediated regulation of speB expression occurs primarily at the transcriptional level and independently of the processing in the speB mRNA 5′ UTR. To conclude, we demonstrate for the first time that RNase Y processing of an mRNA target requires the presence of a G. We also provide new insights on the speB 5′ UTR and on the role of RNase Y in speB regulation.

  • 69.
    Bröms, Jeanette E.
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Ishikawa, Takahiko
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Wai, Sun N.
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Sjöstedt, Anders
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    A functional VipA-VipB interaction is required for the type VI secretion system activity of Vibrio cholerae O1 strain A15522013Inngår i: BMC Microbiology, ISSN 1471-2180, E-ISSN 1471-2180, Vol. 13, s. 96-Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Background: Many Gram-negative bacteria rely on a type VI secretion system (T6SS) to infect eukaryotic cells or to compete against other microbes. Common to these systems is the presence of two conserved proteins, in Vibrio cholerae denoted VipA and VipB, which have been shown to interact in many clinically relevant pathogens. In this study, mutagenesis of a defined region within the VipA protein was used to identify residues important for VipB binding in V. cholerae O1 strain A1552. Results: A dramatically diminished interaction was shown to correlate with a decrease in VipB stability and a loss of hemolysin co-regulated protein (Hcp) secretion and rendered the bacterium unable to compete with Escherichia coli in a competition assay. Conclusions: This confirms the biological relevance of the VipA-VipB interaction, which is essential for the T6SS activity of many important human pathogens.

  • 70.
    Bröms, Jeanette E
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Lavander, Moa
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Meyer, Lena
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Sjöstedt, Anders
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    IglG and IglI of the Francisella pathogenicity island are important virulence determinants of Francisella tularensis LVS2011Inngår i: Infection and Immunity, ISSN 0019-9567, E-ISSN 1098-5522, Vol. 79, nr 9, s. 3683-3696Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The Gram-negative bacterium Francisella tularensis is the causative agent of tularemia, a disease intimately associated with the multiplication of the bacterium within host macrophages. This in turn requires the expression of Francisella pathogenicity island (FPI) genes, believed to encode a type VI secretion system. While the exact functions of many of the components have yet to be revealed, some have been found to contribute to the ability of Francisella to cause systemic infection in mice as well as to prevent phagolysosomal fusion and facilitate escape into the host cytosol. Upon reaching this compartment, the bacterium rapidly multiplies, inhibits activation of the inflammasome, and ultimately causes apoptosis of the host cell. In this study, we analyzed the contribution of the FPI-encoded proteins IglG, IglI, and PdpE to the aforementioned processes in F. tularensis LVS. The ΔpdpE mutant behaved similarly to the parental strain in all investigated assays. In contrast, ΔiglG and ΔiglI mutants, although they were efficiently replicating in J774A.1 cells, both exhibited delayed phagosomal escape, conferred a delayed activation of the inflammasome, and exhibited reduced cytopathogenicity as well as marked attenuation in the mouse model. Thus, IglG and IglI play key roles for modulation of the intracellular host response and also for the virulence of F. tularensis.

  • 71.
    Bröms, Jeanette E
    et al.
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi.
    Lavander, Moa
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi.
    Sjöstedt, Anders
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi.
    Dissection of the Functions of the IglC Protein of Francisella tularensis2010Inngår i: The challenge of highly pathogenic microorganisms: mechanisms of virulence and novel medical countermeasures, Springer, 2010, s. 67-75Konferansepaper (Fagfellevurdert)
    Abstract [en]

    Francisella tularensis harbors genes with similarity to genes encoding components of a type VI secretion system (T6SS). These include iglA and iglB, the homologues of which are conserved in T6SSs. They are part of the igl operon, also encompassing the iglC and iglD genes. We have used a yeast two-hybrid system to study the interaction of the Igl proteins of E tularensis LVS. Previously, we identified a region of IglA necessary for efficient binding to IglB as well as for IglAB protein stability and intra-macrophage growth with an essential role for a conserved alpha-helical region. Thus, IglA-IglB complex formation is clearly crucial for Francisella pathogenicity and the same interaction is conserved in other human pathogens. Herein, the interaction of IglC with other members of the operon was investigated. It showed no binding to the other members in the yeast two-hybrid assay and we found also that two cysteine residues, C191 and C192, predicted to be putative prenylation sites, played no role for the important contribution of IglC to the intracellular replication of E tularensis although C191 was important for the stability of the protein.

  • 72.
    Bröms, Jeanette E.
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Meyer, Lena
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Lavander, Moa
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Larsson, Pär
    Sjöstedt, Anders
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    DotU and VgrG, core components of type VI secretion systems, are essential for Francisella LVS pathogenicity2012Inngår i: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 7, nr 4, artikkel-id e34639Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The Gram-negative bacterium Francisella tularensis causes tularemia, a disease which requires bacterial escape from phagosomes of infected macrophages. Once in the cytosol, the bacterium rapidly multiplies, inhibits activation of the inflammasome and ultimately causes death of the host cell. Of importance for these processes is a 33-kb gene cluster, the Francisella pathogenicity island (FPI), which is believed to encode a type VI secretion system (T6SS). In this study, we analyzed the role of the FPI-encoded proteins VgrG and DotU, which are conserved components of type VI secretion (T6S) clusters. We demonstrate that in F. tularensis LVS, VgrG was shown to form multimers, consistent with its suggested role as a trimeric membrane puncturing device in T6SSs, while the inner membrane protein DotU was shown to stabilize PdpB/IcmF, another T6SS core component. Upon infection of J774 cells, both Delta vgrG and Delta dotU mutants did not escape from phagosomes, and subsequently, did not multiply or cause cytopathogenicity. They also showed impaired activation of the inflammasome and marked attenuation in the mouse model. Moreover, all of the DotU-dependent functions investigated here required the presence of three residues that are essentially conserved among all DotU homologues. Thus, in agreement with a core function in T6S clusters, VgrG and DotU play key roles for modulation of the intracellular host response as well as for the virulence of F. tularensis.

  • 73.
    Bröms, Jeanette E.
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Meyer, Lena
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi. Department of Experimental Medical Science, Section for Immunology, Lund University, Lund, Sweden.
    Sjöstedt, Anders
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi.
    A mutagenesis-based approach identifies amino acids in the N-terminal part of Francisella tularensis IglE that critically control type VI system-mediated secretion2017Inngår i: Virulence, ISSN 2150-5594, E-ISSN 2150-5608, Vol. 8, nr 6, s. 821-847Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The Gram-negative bacterium Francisella tularensis is the etiological agent of the zoonotic disease tularemia. Its life cycle is characterized by an ability to survive within phagocytic cells through phagosomal escape and replication in the cytosol, ultimately causing inflammasome activation and host cell death. Required for these processes is the Francisella Pathogenicity Island (FPI), which encodes a Type VI secretion system (T6SS) that is active during intracellular infection. In this study, we analyzed the role of the FPI-component IglE, a lipoprotein which we previously have shown to be secreted in a T6SS-dependent manner. We demonstrate that in F. tularensis LVS, IglE is an outer membrane protein. Upon infection of J774 cells, an Delta iglE mutant failed to escape from phagosomes, and subsequently, to multiply and cause cytopathogenicity. Moreover, Delta iglE was unable to activate the inflammasome, to inhibit LPS-stimulated secretion of TNF-alpha, and showed marked attenuation in the mouse model. In F. novicida, IglE was required for in vitro secretion of IglC and VgrG. A mutagenesis-based approach involving frameshift mutations and alanine substitution mutations within the first similar to 38 residues of IglE revealed that drastic changes in the sequence of the extreme N-terminus (residues 2-6) were well tolerated and, intriguingly, caused hyper-secretion of IglE during intracellular infection, while even subtle mutations further downstream lead to impaired protein function. Taken together, this study highlights the importance of IglE in F. tularensis pathogenicity, and the contribution of the N-terminus for all of the above mentioned processes.

  • 74.
    Bröms, Jeanette E.
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Meyer, Lena
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Sun, Kun
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Lavander, Moa
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Sjöstedt, Anders
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Klinisk bakteriologi. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Unique substrates secreted by the type VI secretion system of Francisella tularensis during intramacrophage infection2012Inngår i: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 7, nr 11, artikkel-id e50473Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Gram-negative bacteria have evolved sophisticated secretion machineries specialized for the secretion of macromolecules important for their life cycles. The Type VI secretion system (T6SS) is the most widely spread bacterial secretion machinery and is encoded by large, variable gene clusters, often found to be essential for virulence. The latter is true for the atypical T6SS encoded by the Francisella pathogenicity island (FPI) of the highly pathogenic, intracellular bacterium Francisella tularensis. We here undertook a comprehensive analysis of the intramacrophage secretion of the 17 FPI proteins of the live vaccine strain, LVS, of F. tularensis. All were expressed as fusions to the TEM beta-lactamase and cleavage of the fluorescent substrate CCF2-AM, a direct consequence of the delivery of the proteins into the macrophage cytosol, was followed over time. The FPI proteins IglE, IglC, VgrG, IglI, PdpE, PdpA, IglJ and IglF were all secreted, which was dependent on the core components DotU, VgrG, and IglC, as well as IglG. In contrast, the method was not directly applicable on F. novicida U112, since it showed very intense native beta-lactamase secretion due to FTN_1072. Its role was proven by ectopic expression in trans in LVS. We did not observe secretion of any of the LVS substrates VgrG, IglJ, IglF or IglI, when tested in a FTN_1072 deficient strain of F. novicida, whereas IglE, IglC, PdpA and even more so PdpE were all secreted. This suggests that there may be fundamental differences in the T6S mechanism among the Francisella subspecies. The findings further corroborate the unusual nature of the T6SS of F. tularensis since almost all of the identified substrates are unique to the species.

  • 75.
    Buckland, Robert J
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Watt, Danielle L
    Chittoor, Balasubramanyam
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Nilsson, Anna Karin
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Kunkel, Thomas A
    Chabes, Andrei
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Increased and Imbalanced dNTP Pools Symmetrically Promote Both Leading and Lagging Strand Replication Infidelity2014Inngår i: PLOS Genetics, ISSN 1553-7390, E-ISSN 1553-7404, Vol. 10, nr 12, artikkel-id e1004846Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The fidelity of DNA replication requires an appropriate balance of dNTPs, yet the nascent leading and lagging strands of the nuclear genome are primarily synthesized by replicases that differ in subunit composition, protein partnerships and biochemical properties, including fidelity. These facts pose the question of whether imbalanced dNTP pools differentially influence leading and lagging strand replication fidelity. Here we test this possibility by examining strand-specific replication infidelity driven by a mutation in yeast ribonucleotide reductase, rnr1-Y285A, that leads to elevated dTTP and dCTP concentrations. The results for the CAN1 mutational reporter gene present in opposite orientations in the genome reveal that the rates, and surprisingly even the sequence contexts, of replication errors are remarkably similar for leading and lagging strand synthesis. Moreover, while many mismatches driven by the dNTP pool imbalance are efficiently corrected by mismatch repair, others are repaired less efficiently, especially those in sequence contexts suggesting reduced proofreading due to increased mismatch extension driven by the high dTTP and dCTP concentrations. Thus the two DNA strands of the nuclear genome are at similar risk of mutations resulting from this dNTP pool imbalance, and this risk is not completely suppressed even when both major replication error correction mechanisms are genetically intact.

  • 76.
    Bueno, Emilio
    et al.
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). CSIC, Estac Expt Zaidin, Dept Soil Microbiol & Symbiot Syst, C Profesor Albareda 1, E-18008 Granada, Spain.
    Robles, Eloy F.
    Torres, Maria J.
    Krell, Tino
    Bedmar, Eulogio J.
    Delgado, Maria J.
    Mesa, Socorro
    Disparate response to microoxia and nitrogen oxides of the Bradyrhizobium japonicum napEDABC, nirK and norCBQD denitrification genes2017Inngår i: Nitric oxide, ISSN 1089-8603, E-ISSN 1089-8611, Vol. 68, s. 137-149Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Expression of the Bradyrhizobium japonicum napEDABC, nirK and norCBQD denitrification genes requires low oxygen (O-2) tension and nitrate (NO3), through a regulatory network comprised of two coordinated cascades, FixLJ-FixK(2)-NnrR and RegSR-NifA. To precisely understand how these signals are integrated in the FixLJ-FixK(2)-NnrR circuit, we analyzed beta-Galactosidase activities from napE-lacZ, nirK-lacZ and norC-lacZ fusions, and performed analyses of NapC and NorC levels as well as periplasmic nitrate reductase (Nap) activity, in B. japonicum wildtype and fixK(2) and nnrR mutant backgrounds. While microoxic conditions (2% O-2 at headspace) were sufficient to induce expression of napEDABC and nirK genes and this control depends on FixK(2), norCBQD expression requires, in addition to microoxia, nitric oxide gas (NO) and both FixK(2) and NnrR transcription factors. Purified FixK(2) protein directly interacted and activated transcription in collaboration with B. japonicum RNA polymerase (RNAP) from the napEDABC and nirK promoters, but not from the norCBQD promoter. Further, recombinant NnrR protein bound exclusively to the norCBQD promoter in an O-2-sensitive manner. Our work suggest a disparate regulation of B. japonicum denitrifying genes expression with regard to their dependency to microoxia, nitrogen oxides (NOx), and the regulatory proteins FixK(2) and NnrR. In this control, expression of napEDABC and nirK genes requires microoxic conditions and directly depends on FixK2, while expression of norCBQD genes relies on NO, being NnrR the candidate which directly interacts with the norCBQD promoter. 

  • 77.
    Bueno, Emilio
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Sit, Brandon
    Waldor, Matthew K.
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Anaerobic nitrate reduction divergently governs population expansion of the enteropathogen Vibrio cholerae2018Inngår i: Nature Microbiology, E-ISSN 2058-5276, Vol. 3, nr 12, s. 1346-1353Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    To survive and proliferate in the absence of oxygen, many enteric pathogens can undergo anaerobic respiration within the host by using nitrate (NO3-) as an electron acceptor(1,2). In these bacteria, NO3- is typically reduced by a nitrate reductase to nitrite (NO2-), a toxic intermediate that is further reduced by a nitrite reductase(3). However, Vibrio cholerae, the intestinal pathogen that causes cholera, lacks a nitrite reductase, leading to NO2- accumulation during nitrate reduction 4(.) Thus, V. cholerae is thought to be unable to undergo NO3-(-)dependent anaerobic respiration(4). Here, we show that during hypoxic growth, NO3- reduction in V. cholerae divergently affects bacterial fitness in a manner dependent on environmental pH. Remarkably, in alkaline conditions, V. cholerae can reduce NO3- to support population growth. Conversely, in acidic conditions, accumulation of NO2- from NO3- reduction simultaneously limits population expansion and preserves cell viability by lowering fermentative acid production. Interestingly, other bacterial species such as Salmonella typhimurium, enterohaemorrhagic Escherichia coli (EHEC) and Citrobacter rodentium also reproduced this pH-dependent response, suggesting that this mechanism might be conserved within enteric pathogens. Our findings explain how a bacterial pathogen can use a single redox reaction to divergently regulate population expansion depending on the fluctuating environmental pH.

  • 78. Butina, Karen
    et al.
    Loffler, Susanne
    Rhen, Mikael
    Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Richter-Dahlfors, Agneta
    Electrochemical sensing of bacteria via secreted redox active compounds using conducting polymers2019Inngår i: Sensors and actuators. B, Chemical, ISSN 0925-4005, E-ISSN 1873-3077, Vol. 297, artikkel-id UNSP 126703Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Bacterial infections and antibiotic resistance represent major global threats to public health. Current diagnostics use culture based assays that are reliable but slow, hence appealing for new rapid methods. Here we describe redox sensing as a novel concept for rapid, label-free detection of bacteria. We utilize a two-electrode poly(3,4-ethylenedioxythiophene):polystyrenesulfonate (PEDOT:PSS) based sensor for detection of bacterially secreted redox-active compounds. Using purified redox-active compounds, we show the ability of the sensor to detect and quantify compounds in micromolar concentrations within minutes. When applied for detection and quantification of Salmonella, we show that secreted, low molecular weight redox compounds cause reduction of the PEDOT:PSS electrode. A potential role of redox sensing in infection diagnostics was demonstrated as uropathogenic strains of E. coli., Staphylococcus, Enterococcus, Pseudomonas, Proteus, and Klebsiella spp., major causes of complicated urinary tract infections, were successfully detected in complex media or processed urine. Since numerous bacterial species are capable of extracellular electron transfer, redox sensing may find use as a generic method for bacterial detection with applications in research laboatories, the clinic and industry alike.

  • 79. Byström, Jonas
    et al.
    Thomson, Scott J.
    Johansson, Jörgen
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Edin, Matthew L.
    Zeldin, Darryl C.
    Gilroy, Derek W.
    Smith, Andrew M.
    Bishop-Bailey, David
    Inducible CYP2J2 and its product 11,12-EET promotes bacterial phagocytosis: a role for CYP2J2 deficiency in the pathogenesis of Crohn's disease?2013Inngår i: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 8, nr 9, artikkel-id e75107Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The epoxygenase CYP2J2 has an emerging role in inflammation and vascular biology. The role of CYP2J2 in phagocytosis is not known and its regulation in human inflammatory diseases is poorly understood. Here we investigated the role of CYP2J2 in bacterial phagocytosis and its expression in monocytes from healthy controls and Crohns disease patients. CYP2J2 is anti-inflammatory in human peripheral blood monocytes. Bacterial LPS induced CYP2J2 mRNA and protein. The CYP2J2 arachidonic acid products 11,12-EET and 14,15-EET inhibited LPS induced TNFα release. THP-1 monocytes were transformed into macrophages by 48h incubation with phorbol 12-myristate 13-acetate. Epoxygenase inhibition using a non-selective inhibitor SKF525A or a selective CYP2J2 inhibitor Compound 4, inhibited E. coli particle phagocytosis, which could be specifically reversed by 11,12-EET. Moreover, epoxygenase inhibition reduced the expression of phagocytosis receptors CD11b and CD68. CD11b also mediates L. monocytogenes phagocytosis. Similar, to E. coli bioparticle phagocytosis, epoxygenase inhibition also reduced intracellular levels of L. monocytogenes, which could be reversed by co-incubation with 11,12-EET. Disrupted bacterial clearance is a hallmark of Crohn's disease. Unlike macrophages from control donors, macrophages from Crohn's disease patients showed no induction of CYP2J2 in response to E. coli. These results demonstrate that CYP2J2 mediates bacterial phagocytosis in macrophages, and implicates a defect in the CYP2J2 pathway may regulate bacterial clearance in Crohn's disease.

  • 80.
    Caraballo, Rémi
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen. Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Saleeb, Michael
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen. Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Bauer, Johannes
    Interfaculty Institute of Biochemistry, University of Tübingen, Germany.
    Liaci, Antonio-Manuel
    Interfaculty Institute of Biochemistry, University of Tübingen, Germany.
    Chandra, Naresh
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Virologi. Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Storm, Rickard J
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Virologi. Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Frängsmyr, Lars
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Virologi. Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Qian, Weixing
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Stehle, Thilo
    Interfaculty Institute of Biochemistry, University of Tübingen, Germany ; Department of Pediatrics, Vanderbilt University School of Medicine, USA.
    Arnberg, Niklas
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Virologi. Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Elofsson, Mikael
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen. Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Triazole linker-based trivalent sialic acid inhibitors of adenovirus type 37 infection of human corneal epithelial cells2015Inngår i: Organic and biomolecular chemistry, ISSN 1477-0520, E-ISSN 1477-0539, Vol. 13, nr 35, s. 9194-9205Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Adenovirus type 37 (Ad37) is one of the principal agents responsible for epidemic keratoconjunctivitis (EKC), a severe ocular infection that remains without any available treatment. Recently, a trivalent sialic acid derivative (ME0322, Angew. Chem. Int. Ed., 2011, 50, 6519) was shown to function as a highly potent inhibitor of Ad37, efficiently preventing the attachment of the virion to the host cells and subsequent infection. Here, new trivalent sialic acid derivatives were designed, synthesized and their inhibitory properties against Ad37 infection of the human corneal epithelial cells were investigated. In comparison to ME0322, the best compound (17a) was found to be over three orders of magnitude more potent in a cell-attachment assay (IC50 = 1.4 nM) and about 140 times more potent in a cell-infection assay (IC50 = 2.9nM). X-ray crystallographic analysis demonstrated a trivalent binding mode of all compounds to the Ad37 fiber knob. For the most potent compound ophthalmic toxicity in rabbits was investigated and it was concluded that repeated eye administration did not cause any adverse effects.

  • 81. Castanheira, Sonia
    et al.
    Cestero, Juan J.
    Rico-Perez, Gadea
    Garcia, Pablo
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Ayala, Juan A.
    Graciela Pucciarelli, M.
    Garcia-del Portillo, Francisco
    A Specialized Peptidoglycan Synthase Promotes Salmonella Cell Division inside Host Cells2017Inngår i: mBio, ISSN 2161-2129, E-ISSN 2150-7511, Vol. 8, nr 6, artikkel-id e01685-17Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Bacterial cell division has been studied extensively under laboratory conditions. Despite being a key event in the bacterial cell cycle, cell division has not been explored in vivo in bacterial pathogens interacting with their hosts. We discovered in Salmonella enterica serovar Typhimurium a gene absent in nonpathogenic bacteria and encoding a peptidoglycan synthase with 63% identity to penicillin-binding protein 3 (PBP3). PBP3 is an essential cell division-specific peptidoglycan synthase that builds the septum required to separate daughter cells. Since S. Typhimurium carries genes that encode a PBP3 paralog-which we named PBP3(SAL)-and PBP3, we hypothesized that there are different cell division events in host and nonhost environments. To test this, we generated S. Typhimurium isogenic mutants lacking PBP3(SAL) or the hitherto considered essential PBP3. While PBP3 alone promotes cell division under all conditions tested, the mutant producing only PBP3(SAL) proliferates under acidic conditions (pH <= 5.8) but does not divide at neutral pH. PBP3(SAL) production is tightly regulated with increased levels as bacteria grow in media acidified up to pH 4.0 and in intracellular bacteria infecting eukaryotic cells. PBP3(SAL) activity is also strictly dependent on acidic pH, as shown by beta-lactam antibiotic binding assays. Live-cell imaging microscopy revealed that PBP3(SAL) alone is sufficient for S. Typhimurium to divide within phagosomes of the eukaryotic cell. Additionally, we detected much larger amounts of PBP3(SAL) than those of PBP3 in vivo in bacteria colonizing mouse target organs. Therefore, PBP3(SAL) evolved in S. Typhimurium as a specialized peptidoglycan synthase promoting cell division in the acidic intraphagosomal environment. IMPORTANCE During bacterial cell division, daughter cells separate by a transversal structure known as the division septum. The septum is a continuum of the cell wall and therefore is composed of membrane(s) and a peptidoglycan layer. To date, actively growing bacteria were reported to have only a "cell division-specific" peptidoglycan synthase required for the last steps of septum formation and consequently, essential for bacterial life. Here, we discovered that Salmonella enterica has two peptidoglycan synthases capable of synthesizing the division septum. One of these enzymes, PBP3(SAL), is present only in bacterial pathogens and evolved in Salmonella to function exclusively in acidic environments. PBP3(SAL) is used preferentially by Salmonella to promote cell division in vivo in mouse target organs and inside acidified phagosomes. Our data challenge the concept of only one essential cell division-specific peptidoglycan synthase and demonstrate that pathogens can divide in defined host locations using alternative mechanisms.

  • 82.
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Biology of Vibrio cholerae: editorial overview2017Inngår i: International Microbiology, ISSN 1139-6709, E-ISSN 1618-1905, Vol. 20, nr 3Artikkel i tidsskrift (Annet vitenskapelig)
  • 83.
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Divergent functional roles of D-amino acids secreted by Vibrio cholerae2017Inngår i: International Microbiology, ISSN 1139-6709, E-ISSN 1618-1905, Vol. 20, nr 3, s. 149-150Artikkel, forskningsoversikt (Fagfellevurdert)
    Abstract [en]

    The L-forms of amino acids are used in all kingdoms of life to synthesize proteins. However, the bacterium Vibrio cholerae, the causative agent of cholera, produces D-amino acids which are released to the environment at millimolar concentrations. We baptized these D-amino acids as non-canonical D-amino acids (NCDAAs) since they are different from those (i.e. D-alanine and D-glutamate) normally present in the bacterial cell wall. In V. cholerae, production of NCDAAs relies on the BsrV enzyme, a periplasmic broad spectrum racemase. BsrV multispecific activity, produces of a wide range of distinct D-amino acids. Using a combination of genetics and molecular physiology approaches we have demonstrated that NCDAAs target different cellular processes which may function as part of a cooperative strategy in vibrio communities to protect non-producing members from competing bacteria. Because NCDAA production is widespread in bacteria, we anticipate that NCDAAs are relevant modulators of microbial subpopulations in diverse ecosystems.

  • 84.
    Cava, Felipe
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    de Pedro, Miguel A.
    Peptidoglycan plasticity in bacteria: emerging variability of the murein sacculus and their associated biological functions2014Inngår i: Current Opinion in Microbiology, ISSN 1369-5274, E-ISSN 1879-0364, Vol. 18, s. 46-53Artikkel, forskningsoversikt (Fagfellevurdert)
    Abstract [en]

    The peptidoglycan (PG) sacculus once thought to be just a reinforcing, static and uniform structure, is fast becoming recognized as a dynamic cell constituent involved in every aspect of bacterial physiology. Recent advances showed that in addition to 'classical' tasks - as an essential element to define bacterial shape, size, division and resistance to osmotic stress the sacculus plays very important roles in many other fields. The very few chemical and structural changes that were once considered as bizarre, or maybe exotic exceptions, are now universally accepted as fundamental pieces in bacterial cell wall adaptation to different kinds of environmental stresses; immune response; intra-specific and inter-specific signalling and antibiotics, just to mention a few. Most, if not all, of these implications are a consequence of the enormous adaptability of PG metabolism to cope with changing conditions, a characteristic for which the term plasticity is proposed. Here we overview and comment on a number of recent contributions on the cell wall adaptive responses to environmental challenges that has greatly impacted the already high complexity of the PG biology field. These new evidences have revived the interest in PG plasticity as an exciting and trendy topic in current microbiology which considers this variability as the trustworthy picture of bacterial PG in nature.

  • 85.
    Cava, Felipe
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Kuru, Erkin
    Brun, Yves V.
    de Pedro, Miguel A.
    Modes of cell wall growth differentiation in rod-shaped bacteria2013Inngår i: Current Opinion in Microbiology, ISSN 1369-5274, E-ISSN 1879-0364, Vol. 16, nr 6, s. 731-737Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    A bacterial cell takes on the challenge to preserve and reproduce its shape at every generation against a substantial internal pressure by surrounding itself with a mechanical support, a peptidoglycan cell wall. The enlargement of the cell wall via net incorporation of precursors into the pre-existing wall conditions bacterial growth and morphology. However, generation, reproduction and/or modification of a specific shape requires that the incorporation takes place at precise locations for a defined time period. Much has been learnt in the past few years about the biochemistry of the peptidoglycan synthesis process, but topological approaches to the understanding of shape generation have been hindered by a lack of appropriate techniques. Recent technological advances are paving the way for substantial progress in understanding the mechanisms of bacterial morphogenesis. Here we review the latest developments, focusing on the impact of new techniques on the precise mapping of cell wall growth sites.

  • 86. Cerveny, Lukas
    et al.
    Straskova, Adela
    Dankova, Vera
    Hartlova, Anetta
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Ceckova, Martina
    Staud, Frantisek
    Stulik, Jiri
    Tetratricopeptide Repeat Motifs in the World of Bacterial Pathogens: Role in Virulence Mechanisms2013Inngår i: Infection and Immunity, ISSN 0019-9567, E-ISSN 1098-5522, Vol. 81, nr 3, s. 629-635Artikkel, forskningsoversikt (Fagfellevurdert)
    Abstract [en]

    The tetratricopeptide repeat (TPR) structural motif is known to occur in a wide variety of proteins present in prokaryotic and eukaryotic organisms. The TPR motif represents an elegant module for the assembly of various multiprotein complexes, and thus, TPR-containing proteins often play roles in vital cell processes. As the TPR profile is well defined, the complete TPR protein repertoire of a bacterium with a known genomic sequence can be predicted. This provides a tremendous opportunity for investigators to identify new TPR-containing proteins and study them in detail. In the past decade, TPR-containing proteins of bacterial pathogens have been reported to be directly related to virulence-associated functions. In this minireview, we summarize the current knowledge of the TPR-containing proteins involved in virulence mechanisms of bacterial pathogens while high-lighting the importance of TPR motifs for the proper functioning of class II chaperones of a type III secretion system in the pathogenesis of Yersinia, Pseudomonas, and Shigella.

  • 87.
    Chabes, Andrei
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    dNTPs and maintenance of genome stability2019Inngår i: FEBS Open Bio, E-ISSN 2211-5463, Vol. 9, s. 21-21Artikkel i tidsskrift (Annet vitenskapelig)
  • 88.
    Chabes, Andrei
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik. Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Stillman, Bruce
    Constitutively high dNTP concentration inhibits cell cycle progression and the DNA damage checkpoint in yeast Saccharomyces cerevisiae.2007Inngår i: Proceedings of the National Academy of Sciences of the United States of America, ISSN 0027-8424, Vol. 104, nr 4, s. 1183-8Artikkel i tidsskrift (Fagfellevurdert)
  • 89.
    Chandra, Naresh
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Avdelningen för virologi. Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå University.
    The glycobiology of human adenovirus infections: implications for tropism and treatment2019Doktoravhandling, med artikler (Annet vitenskapelig)
    Abstract [en]

    Human adenoviruses (HAdVs) are common human pathogens, causing gastrointestinal, ocular, and respiratory infections on a regular basis. Epidemic keratoconjunctivitis (EKC) is a severe ocular infection for which no approved antivirals are available. HAdV-D37 is one of the causative agents of EKC and uses sialic acid (SA)-containing glycans as cellular receptors. HAdV-D37 interacts with SA via the knob domain of the trimeric virus fiber protein, containing three SA-binding sites. HAdV-D37 also bind to glycosaminoglycans (GAGs), but the outcome of this interaction remains unknown. Here, using biochemical and cell-based assays, the impact of GAGs on HAdV-D37 infection (paper I) was investigated. We found that HAdV-D37 interacts with both soluble and cell-surface sulfated GAGs via the knob domain of the viral fiber protein. Remarkably, removal of heparan sulfate (HS; a type of GAG) from human corneal epithelial (HCE) cells by heparinase III enhanced HAdV-D37 infection. We propose that sulfated GAGs in bodily secretions and on plasma membranes function as decoy receptors that prevent the virus from binding to SA-containing receptors and inhibit subsequent virus infection. We also found abundant HS in the basement membrane of the human corneal epithelium. We suggest that this layer of HS functions as a barrier to sub-epithelial infection of HAdV-D37. Based on this finding, we hypothesized that GAG-mimetics may act as artificial decoy receptors and inhibit HAdV-D37 infection. Here, the antiviral effect of suramin (a known GAG-mimetic) and its analogs against HAdV-D37 (paper II) was evaluated. Interestingly, all compounds displayed antiviral effects by inhibiting the binding of HAdV-D37 to HCE cells. The antiviral effect of suramin was HAdV species-specific. We report for the first time that virus binding to cell-surface decoy receptor constitutes a potential target for antiviral drug development.

    HAdVs are the major cause of infectious conjunctivitis, constituting up to 75% of all conjunctivitis cases worldwide. Species B HAdV type 3 (HAdV-B3) causes pharyngoconjunctival fever (PCF), whereas HAdV-D8, -D37, and -D64 cause EKC. Recently, HAdV-D53, -D54, and -D56 have emerged as new EKC-causing agents. HAdV-E4 causes both PCF and EKC. SA-containing glycans have been established as cellular receptors for HAdV-D37. By means of cell-based assays, we investigated if ocular HAdVs other than HAdV-D37 also use SA-containing glycans as receptors on HCE cells (paper III). It was found that SA-containing glycans function as cellular receptors for five (HAdV-D8, -D37, -D53, -D54, and -D64) out of six EKC-causing species D HAdVs. We showed that these viruses interact with SAs via the knob domain of the viral fiber protein. HAdV-E4 and -D56 infection of cells was independent of SAs. Surprisingly, HCE cells were completely refractory to HAdV-B3 infection. A trivalent sialic acid (TSA) derivative ME0462 (compound 17a in paper II), designed to bind to SA-binding sites on HAdV-D37 fiber knob, also showed potent antiviral activity against several EKC-causing HAdVs. This suggests that ME0462 can be used as a broad-spectrum antiviral against known and emerging EKC-causing HAdVs. Surface plasmon resonance (SPR) analysis confirmed a direct interaction between ME0462 and fiber knobs of EKC-causing HAdVs.

    Recently, a TSA derivative (ME0322; designed to bind to SA-binding sites on HAdV-D37 fiber knob) was shown potent antiviral against HAdV-D37 in vitro. To improve the antiviral potency of this compound, six new TSA derivatives were synthesized and their inhibitory effects were evaluated against HAdV-D37 (paper IV). Interestingly, the best compound 17a was found approximately three orders of magnitude more potent (IC50 (binding) = 1.4 nM, IC50 (infection) = 2.9 nM) than ME0322 (IC50 in µM range). SPR data showed that HAdV-D37 fiber knob binds to TSA compounds with high affinities. Structural data revealed the trivalent binding mode of all newly synthesized TSA compounds to HAdV-D37 fiber knob. Ophthalmic toxicity of compound 17a (best compound) was also investigated in rabbits without any sign of toxicity.

    HAdV-D36 is a member of species D HAdV and has the ability to infect a broad range of animals, which is unusual for HAdVs. Another remarkable feature of HAdV-D36 is that this virus induces obesity in experimental animals. Several epidemiological studies highlighted a link between HAdV-D36 and human obesity. There is no information about the cellular receptor usage by HAdV-D36. Using structural biology and cell-based approaches, we investigated the cellular receptor(s) for HAdV-D36 (paper V).  We show that HAdV-D36 attaches to host cells (via the fiber knob) using the coxsackie and adenovirus receptor (CAR), SA-containing glycans, and one or more unknown proteins or glycoproteins. Using glycan microarray, we found that HAdV-D36 displays binding preference to a rare SA-variant: 4-O,5-N-diacetylneuraminic acid (Neu4,5Ac2), over the more common SA (in humans) i.e. 5-N-acetylneuraminic acid (Neu5Ac). Structural analysis of HAdV-D36 fiber knob:Neu4,5Ac2 complex explained this preference. To date, Neu4,5Achas not been detected in humans, although it is synthesized by many domestic and livestock animals. Our results indicate that HAdV-D36 has evolved to utilize a specialized set of cellular receptors that coincide with a unique host range and pathogenicity profile.

    These studies provide insights into multiple roles of glycans in HAdV infection cycle and highlight the therapeutic potential of glycans/glycan-mimetics in HAdV-D37 infection.

  • 90.
    Charpentier, Emmanuelle
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Department of Regulation in Infection Biology, Helmholtz Centre for Infection Research, Braunschweig, Germany; Hannover Medical School, Hannover, Germany.
    CRISPR-Cas9: how research on a bacterial RNA-guided mechanism opened new perspectives in biotechnology and biomedicine2015Inngår i: EMBO Molecular Medicine, ISSN 1757-4676, E-ISSN 1757-4684, Vol. 7, nr 4, s. 363-365Artikkel i tidsskrift (Fagfellevurdert)
  • 91.
    Charpentier, Emmanuelle
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Programmable RNA-guided Cas9 endonuclease: a novel tool for genome engineering2014Inngår i: Transgenic research, ISSN 0962-8819, E-ISSN 1573-9368, Vol. 23, nr 1, s. 188-189Artikkel i tidsskrift (Annet vitenskapelig)
  • 92.
    Charpentier, Emmanuelle
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Department of Regulation in Infection Biology, Max Planck Institute for Infection Biology, Berlin, Germany; Institute for Biology, Humboldt University, Berlin, Germany.
    Spotlight on... Emmanuelle Charpentier2018Inngår i: FEMS Microbiology Letters, ISSN 0378-1097, E-ISSN 1574-6968, Vol. 365, nr 4, artikkel-id fnx271Artikkel i tidsskrift (Annet vitenskapelig)
  • 93.
    Charpentier, Emmanuelle
    et al.
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Doudna, Jennifer A.
    Biotechnology: rewriting a genome2013Inngår i: Nature, ISSN 0028-0836, E-ISSN 1476-4687, Vol. 495, nr 7439, s. 50-51Artikkel i tidsskrift (Annet vitenskapelig)
  • 94.
    Charpentier, Emmanuelle
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Richter, Hagen
    van der Oost, John
    White, Malcolm F.
    Biogenesis pathways of RNA guides in archaeal and bacterial CRISPR-Cas adaptive immunity2015Inngår i: FEMS Microbiology Reviews, ISSN 0168-6445, E-ISSN 1574-6976, Vol. 39, nr 3, s. 428-441Artikkel, forskningsoversikt (Fagfellevurdert)
    Abstract [en]

    CRISPR-Cas is an RNA-mediated adaptive immune system that defends bacteria and archaea against mobile genetic elements. Short mature CRISPR RNAs (crRNAs) are key elements in the interference step of the immune pathway. A CRISPR array composed of a series of repeats interspaced by spacer sequences acquired from invading mobile genomes is transcribed as a precursor crRNA (pre-crRNA) molecule. This pre-crRNA undergoes one or two maturation steps to generate the mature crRNAs that guide CRISPR-associated (Cas) protein(s) to cognate invading genomes for their destruction. Different types of CRISPR-Cas systems have evolved distinct crRNA biogenesis pathways that implicate highly sophisticated processing mechanisms. In Types I and III CRISPR-Cas systems, a specific endoribonuclease of the Cas6 family, either standalone or in a complex with other Cas proteins, cleaves the pre-crRNA within the repeat regions. In Type II systems, the trans-acting small RNA (tracrRNA) base pairs with each repeat of the pre-crRNA to form a dual-RNA that is cleaved by the housekeeping RNase III in the presence of the protein Cas9. In this review, we present a detailed comparative analysis of pre-crRNA recognition and cleavage mechanisms involved in the biogenesis of guide crRNAs in the three CRISPR-Cas types.

  • 95.
    Charpentier, Emmanuelle
    et al.
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Schroeder, Renée
    RNA techniques for bacteria2007Inngår i: Current Opinion in Microbiology, ISSN 1369-5274, E-ISSN 1879-0364, Vol. 10, nr 3, s. 254-256Artikkel i tidsskrift (Annet vitenskapelig)
  • 96. Chauhan, Deepika
    et al.
    Srivastava, Pulkit Anupam
    Ritzl, Barbara
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Yennamalli, Ragothaman M.
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Priyadarshini, Richa
    Amino Acid-Dependent Alterations in Cell Wall and Cell Morphology of Deinococcus indicus DR12019Inngår i: Frontiers in Microbiology, ISSN 1664-302X, E-ISSN 1664-302X, Vol. 10, artikkel-id 1449Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Deinococcus radiodurans exhibits growth medium-dependent morphological variation in cell shape, but there is no evidence whether this phenomenon is observed in other members of the Deinococcaceae family. In this study, we isolated a red-pigmented, aerobic, Deinococcus indicus strain DR1 from Dadri wetland, India. This D. indicus strain exhibited cell-morphology transition from rod-shaped cells to multi-cell chains in a growth-medium-dependent fashion. In response to addition of 1% casamino acids in the minimal growth medium, rod-shaped cells formed multi-cell chains. Addition of all 20 amino acids to the minimal medium was able to recapitulate the phenotype. Specifically, a combination of L-methionine, L-lysine, L-aspartate, and L-threonine caused morphological alterations. The transition from rod shape to multi-cell chains is due to delay in daughter cell separation after cell division. Minimal medium supplemented with L-ornithine alone was able to cause cell morphology changes. Furthermore, a comparative UPLC analysis of PG fragments isolated from D. indicus cells propagated in different growth media revealed alterations in the PG composition. An increase in the overall cross-linkage of PG was observed in muropeptides from nutrient-rich TSB and NB media versus PYE medium. Overall our study highlights that environmental conditions influence PG composition and cell morphology in D. indicus.

  • 97.
    Chylinski, Krzysztof
    et al.
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Le Rhun, Anaïs
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Charpentier, Emmanuelle
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    The tracrRNA and Cas9 families of type II CRISPR-Cas immunity systems2013Inngår i: RNA Biology, ISSN 1547-6286, E-ISSN 1555-8584, Vol. 10, nr 5, s. 726-737Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    CRISPR-Cas is a rapidly evolving RNA-mediated adaptive immune system that protects bacteria and archaea against mobile genetic elements. The system relies on the activity of short mature CRISPR RNAs (crRNAs) that guide Cas protein(s) to silence invading nucleic acids. A set of CRISPR-Cas, type II, requires a trans-activating small RNA, tracrRNA, for maturation of precursor crRNA (pre-crRNA) and interference with invading sequences. Following co-processing of tracrRNA and pre-crRNA by RNase III, dual-tracrRNA:crRNA guides the CRISPR-associated endonuclease Cas9 (Csn1) to cleave site-specifically cognate target DNA. Here, we screened available genomes for type II CRISPR-Cas loci by searching for Cas9 orthologs. We analyzed 75 representative loci, and for 56 of them we predicted novel tracrRNA orthologs. Our analysis demonstrates a high diversity in cas operon architecture and position of the tracrRNA gene within CRISPR-Cas loci. We observed a correlation between locus heterogeneity and Cas9 sequence diversity, resulting in the identification of various type II CRISPR-Cas subgroups. We validated the expression and co-processing of predicted tracrRNAs and pre-crRNAs by RNA sequencing in five bacterial species. This study reveals tracrRNA family as an atypical, small RNA family with no obvious conservation of structure, sequence or localization within type II CRISPR-Cas loci. The tracrRNA family is however characterized by the conserved feature to base-pair to cognate pre-crRNA repeats, an essential function for crRNA maturation and DNA silencing by dual-RNA:Cas9. The large panel of tracrRNA and Cas9 ortholog sequences should constitute a useful database to improve the design of RNA-programmable Cas9 as genome editing tool.

  • 98.
    Chylinski, Krzysztof
    et al.
    Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Max F. Perutz Laboratories, University of Vienna, Austria .
    Makarova, Kira S.
    Charpentier, Emmanuelle
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Helmholtz Centre for Infection Research, Department of Regulation in Infection Biology, Braunschweig, Germany ; Hannover Medical School, Hannover, Germany .
    Koonin, Eugene V.
    Classification and evolution of type II CRISPR-Cas systems2014Inngår i: Nucleic Acids Research, ISSN 0305-1048, E-ISSN 1362-4962, Vol. 42, nr 10, s. 6091-6105Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The CRISPR-Cas systems of archaeal and bacterial adaptive immunity are classified into three types that differ by the repertoires of CRISPR-associated (cas) genes, the organization of cas operons and the structure of repeats in the CRISPR arrays. The simplest among the CRISPR-Cas systems is type II in which the endonuclease activities required for the interference with foreign deoxyribonucleic acid (DNA) are concentrated in a single multidomain protein, Cas9, and are guided by a co-processed dual-tracrRNA: crRNA molecule. This compact enzymatic machinery and readily programmable site-specific DNA targeting make type II systems top candidates for a new generation of powerful tools for genomic engineering. Here we report an updated census of CRISPR-Cas systems in bacterial and archaeal genomes. Type II systems are the rarest, missing in archaea, and represented in similar to 5% of bacterial genomes, with an over-representation among pathogens and commensals. Phylogenomic analysis suggests that at least three cas genes, cas1, cas2 and cas4, and the CRISPR repeats of the type II-B system were acquired via recombination with a type I CRISPR-Cas locus. Distant homologs of Cas9 were identified among proteins encoded by diverse transposons, suggesting that type II CRISPR-Cas evolved via recombination of mobile nuclease genes with type I loci.

  • 99. Coquel, Flavie
    et al.
    Silva, Maria-Joao
    Técher, Hervé
    Zadorozhny, Karina
    Sharma, Sushma
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Nieminuszczy, Jadwiga
    Mettling, Clément
    Dardillac, Elodie
    Barthe, Antoine
    Schmitz, Anne-Lyne
    Promonet, Alexy
    Cribier, Alexandra
    Sarrazin, Amélie
    Niedzwiedz, Wojciech
    Lopez, Bernard
    Costanzo, Vincenzo
    Krejci, Lumir
    Chabes, Andrei
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik. Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Benkirane, Monsef
    Lin, Yea-Lih
    Pasero, Philippe
    SAMHD1 acts at stalled replication forks to prevent interferon induction2018Inngår i: Nature, ISSN 0028-0836, E-ISSN 1476-4687, Vol. 557, nr 7703, s. 57-61Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    SAMHD1 was previously characterized as a dNTPase that protects cells from viral infections. Mutations in SAMHD1 are implicated in cancer development and in a severe congenital inflammatory disease known as Aicardi-Goutières syndrome. The mechanism by which SAMHD1 protects against cancer and chronic inflammation is unknown. Here we show that SAMHD1 promotes degradation of nascent DNA at stalled replication forks in human cell lines by stimulating the exonuclease activity of MRE11. This function activates the ATR-CHK1 checkpoint and allows the forks to restart replication. In SAMHD1-depleted cells, single-stranded DNA fragments are released from stalled forks and accumulate in the cytosol, where they activate the cGAS-STING pathway to induce expression of pro-inflammatory type I interferons. SAMHD1 is thus an important player in the replication stress response, which prevents chronic inflammation by limiting the release of single-stranded DNA from stalled replication forks.

  • 100.
    Costa, Tiago
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Amer, Ayad
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Farag, Salah
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Wolf-Watz, Hans
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Fällman, Maria
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Fahlgren, Anna
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Edgren, Tomas
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Francis, Matthew
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Type III secretion translocon assemblies that attenuate Yersinia virulence2013Inngår i: Cellular Microbiology, ISSN 1462-5814, E-ISSN 1462-5822, Vol. 15, nr 7, s. 1088-1110Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Type III secretion enables bacteria to intoxicate eukaryotic cells with anti-host effectors. A class of secreted cargo are the two hydrophobic translocators that form a translocon pore in the host cell plasma membrane through which the translocated effectors may gain cellular entry. In pathogenic Yersinia, YopB and YopD shape this translocon pore. Here, four in cis yopD mutations were constructed to disrupt a predicted α-helix motif at the C-terminus. Mutants YopD(I262P) and YopD(K267P) poorly localized Yop effectors into target eukaryotic cells and failed to resist uptake and killing by immune cells. These defects were due to deficiencies in host-membrane insertion of the YopD-YopB translocon. Mutants YopD(A263P) and YopD(A270P) had no measurable in vitro translocation defect, even though they formed smaller translocon pores in erythrocyte membranes. Despite this, all four mutants were attenuated in a mouse infection model. Hence, YopD variants have been generated that can spawn translocons capable of targeting effectors in vitro, yet were bereft of any lethal effect in vivo. Therefore, Yop translocators may possess other in vivo functions that extend beyond being a portal for effector delivery into host cells.

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