oru.sePublications
Change search
Refine search result
1 - 32 of 32
CiteExportLink to result list
Permanent link
Cite
Citation style
  • apa
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Other style
More styles
Language
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Other locale
More languages
Output format
  • html
  • text
  • asciidoc
  • rtf
Rows per page
  • 5
  • 10
  • 20
  • 50
  • 100
  • 250
Sort
  • Standard (Relevance)
  • Author A-Ö
  • Author Ö-A
  • Title A-Ö
  • Title Ö-A
  • Publication type A-Ö
  • Publication type Ö-A
  • Issued (Oldest first)
  • Issued (Newest first)
  • Created (Oldest first)
  • Created (Newest first)
  • Last updated (Oldest first)
  • Last updated (Newest first)
  • Disputation date (earliest first)
  • Disputation date (latest first)
  • Standard (Relevance)
  • Author A-Ö
  • Author Ö-A
  • Title A-Ö
  • Title Ö-A
  • Publication type A-Ö
  • Publication type Ö-A
  • Issued (Oldest first)
  • Issued (Newest first)
  • Created (Oldest first)
  • Created (Newest first)
  • Last updated (Oldest first)
  • Last updated (Newest first)
  • Disputation date (earliest first)
  • Disputation date (latest first)
Select
The maximal number of hits you can export is 250. When you want to export more records please use the Create feeds function.
  • 1.
    Asghar, Naveed
    et al.
    Örebro University, School of Medical Sciences.
    Gunaltay, Sezin
    School of Medical Sciences, Örebro University, Örebro, Sweden.
    Tran, Pham Tue Hung
    Örebro University, School of Medical Sciences.
    Melik, Wessam
    Örebro University, School of Medical Sciences.
    Höglund, Urban
    Adlego Biomedical AB, Uppsala, Sweden.
    Johansson, Christer
    Academy of Quality Pharm Science and BiQ Pharma AB, Södertälje, Sweden.
    Frelin, Lars
    Laboratory Medicine, Karolinska Institutet, Stockholm, Sweden.
    Sällberg, Matti
    Laboratory Medicine, Karolinska Institutet, Stockholm, Sweden.
    Johansson, Magnus
    Örebro University, School of Medical Sciences.
    DNA launched suicidal flaviviruses as therapeutic vaccine candidates2018Conference paper (Refereed)
    Abstract [en]

    Chronic liver disease, resulting from Hepatitis B virus (HBV), Hepatitis D virus (HDV), or Hepatitis C virus (HCV) infections, contributes to a major health burden worldwide. The relativelyhigh cost of the HCV treatment brings concerns about the accessibility, especially in the developing countries. Hence, there exists a need for cost effect interventions with high efficiency. We aim to develop therapeutic vaccine candidates against HBV, HCV and HDV using DNA based subgenomic flavivirus replicons as a delivery system. Tick-borne encephalitis virus (TBEV), Langat virus (LGTV), West-Nile virus (WNV), or Kunjinvirus (KUNV) replicon with firefly luciferase geneas a reporter were expressed and characterized in cell culture studies. WNV and KUNV replicons showed significantly higher replication compared to their respective negative controls with unfunctional viral RNA dependent RNA polymerase. KUNV and WNV replicons were chosen for cloning the HCV or HB/DV vaccine candidate gene by replacing luciferasegene. Owing to the self-replicating trait of the flavivirus subgenomic replicons, Western blotting demonstrated that the antigen expression by KUNV and WNV replicons was several folds higher than the positive control. These results suggest that DNA based KUNV and WNV replicons may function as carriers for the hepatitis vaccine candidate genes, and these replicons are currently used for in vivostudies in animal models.

  • 2.
    Asghar, Naveed
    et al.
    School of Natural Science, Technology & Environmental Studies, Södertörn University, Huddinge, Sweden .
    Lee, Yi-Ping
    Department of Clinical Microbiology, Virology,Umeå University, Umeå, Sweden; The Laboratory for Molecular Medicine Sweden (MIMS), Umeå University, Umeå, Sweden.
    Nilsson, Emma
    Department of Clinical Microbiology, Virology, Umeå University, Umeå, Sweden; The Laboratory for Molecular Medicine Sweden (MIMS), Umeå University, Umeå, Sweden.
    Lindqvist, Richard
    Department of Clinical Microbiology, Virology, Umeå University, Umeå, Sweden; The Laboratory for Molecular Medicine Sweden (MIMS), Umeå University, Umeå, Sweden.
    Melik, Wessam
    Örebro University, School of Medical Sciences.
    Kröger, Andrea
    Innate Immunity and Infection, Helmholtz Centre for Infection Research, Braunschweig, Germany; Institute for Microbiology, University of Magdeburg, Magdeburg, Germany.
    Överby, Anna K.
    Department of Clinical Microbiology, Virology, Umeå University, Umeå, Sweden; The Laboratory for Molecular Medicine Sweden (MIMS), Umeå University, Umeå, Sweden.
    Johansson, Magnus
    Örebro University, School of Medical Sciences.
    The role of the poly(A) tract in the replication and virulence of tick-borne encephalitis virus2016In: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 6, article id 39265Article in journal (Refereed)
    Abstract [en]

    The tick-borne encephalitis virus (TBEV) is a flavivirus transmitted to humans, usually via tick bites. The virus causes tick-borne encephalitis (TBE) in humans, and symptoms range from mild flu-like symptoms to severe and long-lasting sequelae, including permanent brain damage. It has been suggested that within the population of viruses transmitted to the mammalian host, quasispecies with neurotropic properties might become dominant in the host resulting in neurological symptoms. We previously demonstrated the existence of TBEV variants with variable poly(A) tracts within a single blood-fed tick. To characterize the role of the poly(A) tract in TBEV replication and virulence, we generated infectious clones of Torö-2003 with the wild-type (A)3C(A)6 sequence (Torö-6A) or with a modified (A)3C(A)38 sequence (Torö-38A). Torö-38A replicated poorly compared to Torö-6A in cell culture, but Torö-38A was more virulent than Torö-6A in a mouse model of TBE. Next-generation sequencing of TBEV genomes after passaging in cell culture and/or mouse brain revealed mutations in specific genomic regions and the presence of quasispecies that might contribute to the observed differences in virulence. These data suggest a role for quasispecies development within the poly(A) tract as a virulence determinant for TBEV in mice.

  • 3.
    Asghar, Naveed
    et al.
    School of Natural Science, Technology & Environmental Studies, Södertörn University, Huddinge, Sweden.
    Lindblom, Pontus
    Division of Medical Microbiology, Department of Clinical and Experimental Medicine, Linköping University, Linköping, Sweden.
    Melik, Wessam
    School of Natural Science, Technology & Environmental Studies, Södertörn University, Huddinge, Sweden.
    Lindqvist, Richard
    Department of Clinical Microbiology, Virology, Umeå University, Umeå, Sweden.
    Haglund, Mats
    Department of Infectious Diseases, County Hospital, Kalmar, Sweden.
    Forsberg, Pia
    Division of Infectious Diseases, Department of Clinical and Experimental Medicine, Linköping University, Linköping, Sweden; Clinic of Infectious Diseases, Linköping University Hospital, Linköping, Sweden.
    Överby, Anna K.
    Department of Clinical Microbiology, Virology, Umeå University, Umeå, Sweden.
    Andreassen, Åshild
    Division of Infectious Disease Control, Department of Virology, Norwegian Institute of Public Health, Oslo, Norway.
    Lindgren, Per-Eric
    Division of Medical Microbiology, Department of Clinical and Experimental Medicine, Linköping University, Linköping, Sweden; Division of Medical Services, Department of Microbiology, County Hospital Ryhov, Jönköping, Sweden.
    Johansson, Magnus
    Örebro University, School of Medicine, Örebro University, Sweden. School of Natural Science, Technology & Environmental Studies, Södertörn University, Huddinge, Sweden; RiSC - Inflammatory Response and Infection Susceptibility Centre, Faculty of Medicine and Health, Örebro University, Örebro, Sweden.
    Tick-Borne Encephalitis Virus Sequenced Directly from Questing and Blood-Feeding Ticks Reveals Quasispecies Variance2014In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 9, no 7, article id e103264Article in journal (Refereed)
    Abstract [en]

    The increased distribution of the tick-borne encephalitis virus (TBEV) in Scandinavia highlights the importance of characterizing novel sequences within the natural foci. In this study, two TBEV strains: the Norwegian Mandal 2009 (questing nymphs pool) and the Swedish Saringe 2009 (blood-fed nymph) were sequenced and phylogenetically characterized. Interestingly, the sequence of Mandal 2009 revealed the shorter form of the TBEV genome, similar to the highly virulent Hypr strain, within the 3' non-coding region (3'NCR). A different genomic structure was found in the 3'NCR of Saringe 2009, as in-depth analysis demonstrated TBEV variants with different lengths within the poly(A) tract. This shows that TBEV quasispecies exists in nature and indicates a putative shift in the quasispecies pool when the virus switches between invertebrate and vertebrate environments. This prompted us to further sequence and analyze the 3'NCRs of additional Scandinavian TBEV strains and control strains, Hypr and Neudoerfl. Toro 2003 and Habo 2011 contained mainly a short (A) 3C(A)6 poly(A) tract. A similar pattern was observed for the human TBEV isolates 1993/783 and 1991/4944; however, one clone of 1991/4944 contained an (A) 3C(A)11 poly(A) sequence, demonstrating that quasispecies with longer poly(A) could be present in human isolates. Neudoerfl has previously been reported to contain a poly(A) region, but to our surprise the resequenced genome contained two major quasispecies variants, both lacking the poly(A) tract. We speculate that the observed differences are important factors for the understanding of virulence, spread, and control of the TBEV.

  • 4.
    Asghar, Naveed
    et al.
    Örebro University, School of Medical Sciences.
    Maravelia, Panagiota
    aboratory Medicine, Karolinska Institutet, Stockholm, Sweden.
    Caro-Perez, Noelia
    Laboratory Medicine, Karolinska Institutet, Stockholm, Sweden.
    Tarn, Hung
    Örebro University, School of Medical Sciences.
    Melik, Wessam
    Örebro University, School of Medical Sciences.
    Pasetto, Anna
    aboratory Medicine, Karolinska Institutet, Stockholm, Sweden.
    Ahlen, Gustaf
    aboratory Medicine, Karolinska Institutet, Stockholm, Sweden.
    Frelin, Lars
    aboratory Medicine, Karolinska Institutet, Stockholm, Sweden.
    Höglund, Urban
    Johansson, Christer
    Sällberg, Matti
    aboratory Medicine, Karolinska Institutet, Stockholm, Sweden.
    Johansson, Magnus
    Örebro University, School of Medical Sciences.
    Immunogenicity of DNA launched suicidal flavivirus replicons for protective vaccination against hepatitis viruses2019Conference paper (Refereed)
    Abstract [en]

    Chronic liver disease, resulting from Hepatitis B virus (HBV), Hepatitis D virus (HDV), or Hepatitis C virus (HCV) infections, contributes to a major health burden worldwide. Chronic infections with the hepatitis C virus (HCV) can be effectively cured by antivirals. However, as cured patients can be re-infected they lack protective immune responses. In addition, the relativelyhigh cost of the HCV treatment brings concerns about the accessibility, especially in the developing countries. Hence, there exists a need for cost effect vaccines with high efficiency to control and possibly eradicate Hepatitis viruses globally. The vaccine should induce either, or both, neutralizing antibodies and protective T cell responses. We therefore have developed DNA based flavivirus replicons as a potent delivery system that effectively prime HCV-specific T cell responses. We generated suicidal subgenomic DNA replicons of Tick-borne encephalitis virus (TBEV), Langat virus (LGTV), West-Nile virus (WNV), and Kunjinvirus (KUNV) expressing either a fusion protein between the HCV NS3/4A and a stork hepatitis B virus core or a vaccine candidate gene of HB/DV. Transfection experiments showed that the antigen expression by KUNV and WNV replicons was several folds higher than the antigen expression by standard DNA plasmid with CMV promoter. The immunogenicity of three suicidal flaviviral DNA replicons expressing HCV NS3/4A was tested in mice and compared to HCV NS3/4A expression by the standard DNA plasmid. The KUNV-HCV replicon was the best replicon-based immunogen with respect to priming of HCV NS3/4A-specific T cells as determined by ELISpot, dextramer staining, and polyfunctionality. Importantly, a mutant KUNV-HCV immunogen lacking replication failed to induce immune responses. Thus, the newly developed KUNV-based suicidal DNA launched replicon vaccine for HCV is a highly attractive candidate as a prophylactic vaccine against chronic hepatitis C. In addition, we are currently testing the immunogenicity of KUNV-HB/DV replicon in mice.

  • 5.
    Asghar, Naveed
    et al.
    Örebro University, School of Medical Sciences.
    Maravelia, Panagiota
    Laboratory Medicine, Karolinska Institutet, Stockholm, Sweden.
    Caro-Perez, Noelia
    Laboratory Medicine, Karolinska Institutet, Stockholm, Sweden.
    Tran, Pham Tue Hung
    Örebro University, School of Medical Sciences.
    Melik, Wessam
    Örebro University, School of Medical Sciences.
    Pasetto, Anna
    aboratory Medicine, Karolinska Institutet, Stockholm, Sweden.
    Ahlen, Gustaf
    Laboratory Medicine, Karolinska Institutet, Stockholm, Sweden.
    Frelin, Lars
    Laboratory Medicine, Karolinska Institutet, Stockholm, Sweden.
    Höglund, Urban
    Adlego Biomedical AB, Uppsala, Sweden.
    Johansson, Christer
    Academy of Quality Pharm Science and BiQ Pharma AB, Södertälje, Sweden.
    Sällberg, Matti
    Laboratory Medicine, Karolinska Institutet, Stockholm, Sweden.
    Johansson, Magnus
    Örebro University, School of Medical Sciences.
    Immunogenicity of DNA launched suicidal flavivirus replicons for protective vaccination against hepatitis viruses2019Conference paper (Refereed)
    Abstract [en]

    Chronic liver disease, resulting from Hepatitis B virus (HBV), Hepatitis D virus (HDV), or Hepatitis C virus (HCV) infections, contributes to a major health burden worldwide. Chronic infections with the hepatitis C virus (HCV) can be effectively cured by antivirals. However, as cured patients can be re-infected they lack protective immune responses. In addition, the relativelyhigh cost of the HCV treatment brings concerns about the accessibility, especially in the developing countries. Hence, there exists a need for cost effect vaccines with high efficiency to control and possibly eradicate Hepatitis viruses globally. The vaccine should induce either, or both, neutralizing antibodies and protective T cell responses. We therefore have developed DNA based flavivirus replicons as a potent delivery system that effectively prime HCV-specific T cell responses. We generated suicidal subgenomic DNA replicons of Tick-borne encephalitis virus (TBEV), Langat virus (LGTV), West-Nile virus (WNV), and Kunjinvirus (KUNV) expressing either a fusion protein between the HCV NS3/4A and a stork hepatitis B virus core or a vaccine candidate gene of HB/DV. Transfection experiments showed that the antigen expression by KUNV and WNV replicons was several folds higher than the antigen expression by standard DNA plasmid with CMV promoter. The immunogenicity of three suicidal flaviviral DNA replicons expressing HCV NS3/4A was tested in mice and compared to HCV NS3/4A expression by the standard DNA plasmid. The KUNV-HCV replicon was the best replicon-based immunogen with respect to priming of HCV NS3/4A-specific T cells as determined by ELISpot, dextramer staining, and polyfunctionality. Importantly, a mutant KUNV-HCV immunogen lacking replication failed to induce immune responses. Thus, the newly developed KUNV-based suicidal DNA launched replicon vaccine for HCV is a highly attractive candidate as a prophylactic vaccine against chronic hepatitis C. In addition, we are currently testing the immunogenicity of KUNV-HB/DV replicon in mice.

  • 6.
    Asghar, Naveed
    et al.
    Örebro University, School of Medical Sciences. School of Natural Science, Technology & Environmental Studies, Södertörn University, Huddinge, Sweden; iRiSC – Inflammatory Response and Infection Susceptibility Centre, Faculty of Medicine and Health, Örebro University, Örebro, Sweden.
    Pettersson, John H-O
    Department of Infectious Disease Epidemiology and Modelling, Norwegian Institute of Public Health, Oslo, Norway; Department of Microbiology, National Veterinary Institute, Uppsala, Sweden; Department of Medical Biochemistry and Microbiology (IMBIM), Zoonosis Science Center, Uppsala University, Uppsala, Sweden.
    Dinnetz, Patrik
    School of Natural Science, Technology & Environmental Studies, Södertörn University, Huddinge, Sweden.
    Andreassen, Åshild
    Department of Virology, Division of Infectious Disease Control, Norwegian Institute of Public Health, Oslo, Norway.
    Johansson, Magnus
    Örebro University, School of Medical Sciences.
    Deep sequencing analysis of tick-borne encephalitis virus from questing ticks at natural foci reveals similarities between quasispecies pools of the virus2017In: Journal of General Virology, ISSN 0022-1317, E-ISSN 1465-2099, Vol. 98, no 3, p. 413-421Article in journal (Refereed)
    Abstract [en]

    Every year, tick-borne encephalitis virus (TBEV) causes severe central nervous system infection in 10,000 to 15,000 people in Europe and Asia. TBEV is maintained in the environment by an enzootic cycle that requires a tick vector and a vertebrate host, and the adaptation of TBEV to vertebrate and invertebrate environments is essential for TBEV persistence in nature. This adaptation is facilitated by the error-prone nature of the virus' RNA-dependent RNA polymerase that generates genetically distinct virus variants called quasispecies. TBEV shows a focal geographical distribution pattern where each focus represents a TBEV hotspot. Here we sequenced and characterized two TBEV genomes, JP-296 and JP-554, from questing Ixodes ricinus ticks at a TBEV focus in central Sweden. Phylogenetic analysis showed geographical clustering among the newly sequenced strains and three previously sequenced Scandinavian strains, Toro-2003, Saringe-2009, and Mandal-2009, which originated from same ancestor. Among these five Scandinavian TBEV strains, only Mandal-2009 showed a large deletion within the 3´ non-coding region (NCR) similar to the highly virulent TBEV strain Hypr. Deep sequencing of JP-296, JP-554, and Mandal-2009 revealed significantly high quasispecies diversity for JP-296 and JP-554, with intact 3´NCRs, compared to the low diversity in Mandal-2009, with a truncated 3´NCR. SNP analysis showed that 40% of the SNPs were common between quasispecies populations of JP-296 and JP-554, indicating a putative mechanism for how TBEV persists and is maintained within its natural foci.

  • 7.
    Asghar, Naveed
    et al.
    Södertörns Högskola, Huddinge, Sweden.
    Wessam, Melik
    Södertörns Högskola, Huddinge, Sweden.
    Lindblom, Pontus
    Linköpings Universitet, Linköping, Sweden.
    Lindgren, Per-Erik
    Linköpings Universitet, Linköping, Sweden.
    Andreassen, Åshild
    Norska folkhälsoinstitutet, Oslo, Norway.
    Johansson, Magnus
    Örebro University, School of Health and Medical Sciences, Örebro University, Sweden. Örebro University, School of Medicine, Örebro University, Sweden.
    Genomic Sequencing of Tick-borne Encephalitis Virus frin Questing and Blood-Feeding Ixodes ricinus2013Conference paper (Other academic)
  • 8.
    Bertrand, Yann
    et al.
    Dept Plant & Environm Sci, Univ Gothenburg, Gothenburg, Sweden.
    Töpel, Mats
    Dept Plant & Environm Sci, Univ Gothenburg, Gothenburg, Sweden.
    Elväng, Annelie
    Sch Life Sci, Södertörn Univ, Huddinge, Sweden.
    Melik, Wessam
    Sch Life Sci, Södertörn Univ, Huddinge, Sweden; Dept Genet Microbiol & Toxicol, Stockholm Univ, Stockholm, Sweden.
    Johansson, Magnus
    Sch Life Sci, Södertörn Univ, Huddinge, Sweden.
    First dating of a recombination event in mammalian tick-borne flaviviruses2012In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 7, no 2, article id e31981Article in journal (Refereed)
    Abstract [en]

    The mammalian tick-borne flavivirus group (MTBFG) contains viruses associated with important human and animal diseases such as encephalitis and hemorrhagic fever. In contrast to mosquito-borne flaviviruses where recombination events are frequent, the evolutionary dynamic within the MTBFG was believed to be essentially clonal. This assumption was challenged with the recent report of several homologous recombinations within the Tick-borne encephalitis virus (TBEV). We performed a thorough analysis of publicly available genomes in this group and found no compelling evidence for the previously identified recombinations. However, our results show for the first time that demonstrable recombination (i.e., with large statistical support and strong phylogenetic evidences) has occurred in the MTBFG, more specifically within the Louping ill virus lineage. Putative parents, recombinant strains and breakpoints were further tested for statistical significance using phylogenetic methods. We investigated the time of divergence between the recombinant and parental strains in a Bayesian framework. The recombination was estimated to have occurred during a window of 282 to 76 years before the present. By unravelling the temporal setting of the event, we adduce hypotheses about the ecological conditions that could account for the observed recombination.

  • 9. Brooks, Andrew J
    et al.
    Johansson, Magnus
    John, Anna V
    Xu, Yibin
    Jans, David A
    Vasudevan, Subhash G
    The interdomain region of dengue NS5 protein that binds to the viral helicase NS3 contains independently functional importin beta 1 and importin alpha/beta-recognized nuclear localization signals2002In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 277, no 39, p. 36399-36407Article in journal (Refereed)
    Abstract [en]

    Dengue virus NS5 protein is a multifunctional RNA-dependent RNA polymerase that is essential for virus replication. We have shown previously that the 37- amino acid interdomain spacer sequence (residues (369)X(2)KKX(14)KKKX(11)RKX(3)405) of Dengue2 NS5 contains a functional nuclear localization signal (NLS). In this study, beta-galactosidase fusion proteins carrying point mutations of the positively charged residues or truncations of the interdomain linker region (residues 369-389 or residues 386-405) were analyzed for nuclear import and importin binding activities to show that the N-terminal part of the linker region (residues 369-389, a/bNLS) is critical for nuclear localization and is recognized with high affinity by the conventional NLS-binding importin alpha/beta heterodimeric nuclear import receptor. We also show that the importin beta-binding site (residues 320-368, bNLS) adjacent to the a/bNLS, previously identified by yeast two-hybrid analysis, is functional as an NLS, recognized with high affinity by importin beta, and able to target beta-galactosidase to the nucleus. Intriguingly, the bNLS is highly conserved among Dengue and related flaviviruses, implying a general role for the region and importin beta in the infectious cycle.

  • 10. Cheng, J
    et al.
    Johansson, Magnus
    Nordlund, S
    Expression of P(II) and glutamine synthetase is regulated by P(II), the ntrBC products, and processing of the glnBA mRNA in Rhodospirillum rubrum1999In: Journal of Bacteriology, ISSN 0021-9193, E-ISSN 1098-5530, Vol. 181, no 20, p. 6530-6534Article in journal (Refereed)
    Abstract [en]

    We have studied the transcription of the glnB and glnA genes in Rhodospirillum rubrum with firefly luciferase as a reporter enzyme. Under NH(4)(+) and N(2) conditions, glnBA was cotranscribed from a weak and a strong promoter. In nitrogen-fixing cultures, activity of the latter was highly enhanced by NtrC, but transcription from both promoters occurred under both conditions. There is no promoter controlling transcription of glnA alone, supporting our proposal that the glnA mRNA is produced by processing.

  • 11. Ellencrona, Karin
    et al.
    Syed, Asim
    Johansson, Magnus
    Flavivirus NS5 associates with host-cell proteins zonula occludens-1 (ZO-1) and regulating synaptic membrane exocytosis-2 (RIMS2) via an internal PDZ binding mechanism2009In: Biological chemistry (Print), ISSN 1431-6730, E-ISSN 1437-4315, Vol. 390, no 4, p. 319-323Article in journal (Refereed)
    Abstract [en]

    Dengue virus (DENV) and tick-borne encephalitis virus (TBEV) are flaviviruses, which can cause lethal hemorrhagic fever and encephalitis, respectively. Here, we demonstrate that the TBEV-NS5 and DENV-NS5 proteins use an internal binding mechanism to target human PDZ proteins. TBEV-NS5 has high affinity to regulating synaptic membrane exocytosis-2 (RIMS2) and Scribble, whereas DENV-NS5 binds primarily to the tight junction protein zonula occludens-1 (ZO-1). Targeting of TBEV-NS5 to the plasma membrane is stabilised by ZO-1; however, DENV-NS5 co-localises with ZO-1 in the nucleus. These interactions have potential important roles in the ability of flaviviruses to manipulate cell proliferation, junction permeability and the interferon pathways.

  • 12.
    Elväng, Annelie
    et al.
    School of Life Sciences, Södertörn University, Huddinge, Sweden.
    Melik, Wessam
    School of Life Sciences, Södertörn University, Huddinge, Sweden; Department of Genetics, Microbiology and Toxicology, Stockholm University, Stockholm, Sweden.
    Bertrand, Yann
    School of Life Sciences, Södertörn University, Huddinge, Sweden.
    Lönn, Mikael
    School of Life Sciences, Södertörn University, Huddinge, Sweden.
    Johansson, Magnus
    School of Life Sciences, Södertörn University, Huddinge, Sweden.
    Sequencing of a tick-borne encephalitis virus from Ixodes ricinus reveals a thermosensitive RNA switch significant for virus propagation in ectothermic arthropods2011In: Vector Borne and Zoonotic Diseases, ISSN 1530-3667, E-ISSN 1557-7759, Vol. 11, no 6, p. 649-658Article in journal (Refereed)
    Abstract [en]

    Tick-borne encephalitis virus (TBEV) is a flavivirus with major impact on global health. The geographical TBEV distribution is expanding, thus making it pivotal to further characterize the natural virus populations. In this study, we completed the earlier partial sequencing of a TBEV pulled out of a pool of RNA extracted from 115 ticks collected on Torö in the Stockholm archipelago. The total RNA was sufficient for all sequencing of a TBEV genome (Torö-2003), without conventional enrichment procedures such as cell culturing or suckling mice amplification. To our knowledge, this is the first time that the genome of TBEV has been sequenced directly from an arthropod reservoir. The Torö-2003 sequence has been characterized and compared with other TBE viruses. In silico analyses of secondary RNA structures formed by the two untranslated regions revealed a temperature-sensitive structural shift between a closed replicative form and an open AUG accessible form, analogous to a recently described bacterial thermoswitch. Additionally, novel phylogenetic conserved structures were identified in the variable part of the 3'-untranslated region, and their sequence and structure similarity when compared with earlier identified structures suggests an enhancing function on virus replication and translation. We propose that the thermo-switch mechanism may explain the low TBEV prevalence often observed in environmentally sampled ticks. Finally, we were able to detect variations that help in the understanding of virus adaptations to varied environmental temperatures and mammalian hosts through a comparative approach that compares RNA folding dynamics between strains with different mammalian cell passage histories.

  • 13.
    Hamsten, C.
    et al.
    Department of Medicine Solna, Clinical Immunology and Allergy Unit, Karolinska Institutet and University Hospital Solna, Stockholm, Sweden; Center for Inflammatory Diseases, Karolinska Institutet, Stockholm, Sweden.
    Starkhammar, M.
    Department of Internal Medicine, Södersjukhuset, Stockholm, Sweden.
    Tran, T. A.
    Department of Medicine Solna, Clinical Immunology and Allergy Unit, Karolinska Institutet and University Hospital Solna, Stockholm, Sweden.
    Johansson, Magnus
    Örebro University, School of Medicine, Örebro University, Sweden. School of Life Sciences, Södertörns University, Stockholm, Sweden.
    Bengtsson, U.
    Department of Internal Medicine, Södersjukhuset, Stockholm, Sweden.
    Ahlén, G.
    Division of Clinical Microbiology, Department of Laboratory Medicine, Karolinska Institutet and University Hospital Huddinge, Stockholm, Sweden.
    Sällberg, M.
    Division of Clinical Microbiology, Department of Laboratory Medicine, Karolinska Institutet and University Hospital Huddinge, Stockholm, Sweden.
    Grönlund, H.
    Department of Medicine Solna, Clinical Immunology and Allergy Unit, Karolinska Institutet and University Hospital Solna, Stockholm, Sweden.
    van Hage, M.
    Department of Medicine Solna, Clinical Immunology and Allergy Unit, Karolinska Institutet and University Hospital Solna, Stockholm, Sweden.
    Identification of galactose-α-1,3-galactose in the gastrointestinal tract of the tick Ixodes ricinus; possible relationship with red meat allergy2013In: Allergy. European Journal of Allergy and Clinical Immunology, ISSN 0105-4538, E-ISSN 1398-9995, Vol. 68, no 4, p. 549-552Article in journal (Refereed)
    Abstract [en]

    Patients with IgE antibodies against the carbohydrate epitope galactose-α-1,3-galactose (α-Gal) have reported severe allergic reactions after consumption of red meat. Investigations have revealed associations between IgE to α-Gal and tick bites. We provide the first direct evidence that α-Gal is present within ticks thus potentially explaining the relationship between tick exposure and sensitization to α-Gal, with development of red meat allergy as a secondary phenomena. Serum from Swedish patients with delayed severe reactions to red meat was included in the study. A dose-dependent inhibition of IgE responses to α-Gal by the tick Ixodes ricinus is demonstrated. Furthermore, using cryostat-cut sections of I. ricinus, we show that both a monoclonal and a polyclonal antibody against α-Gal stains the gastrointestinal tract of the tick. The same pattern is seen when staining with patient sera IgE positive to α-Gal. These results confirm that the α-Gal epitope is present in I. ricinus and imply host exposure to α-Gal during a tick bite. This provides further evidence that tick bites are associated with IgE responses to α-Gal and red meat allergy.

  • 14.
    Johansson, Magnus
    Örebro University, School of Medical Sciences.
    Development of an edible Tick-borne encephalitis vaccine2017Conference paper (Other academic)
  • 15. Johansson, Magnus
    et al.
    Brooks, A J
    Jans, D A
    Vasudevan, S G
    A small region of the dengue virus-encoded RNA-dependent RNA polymerase, NS5, confers interaction with both the nuclear transport receptor importin-beta and the viral helicase, NS32001In: Journal of General Virology, ISSN 0022-1317, E-ISSN 1465-2099, Vol. 82, no Pt 4, p. 735-745Article in journal (Refereed)
    Abstract [en]

    The dengue virus RNA-dependent RNA polymerase, NS5, and the protease/helicase, NS3, are multidomain proteins that have been shown to interact both in vivo and in vitro. A hyperphosphorylated form of NS5 that does not interact with NS3 has been detected in the nuclei of virus-infected cells, presumably as the result of the action of a functional nuclear localization sequence within the interdomain region of NS5 (residues 369-405). In this study, it is shown by using the yeast two-hybrid system that the C-terminal region of NS3 (residues 303-618) interacts with the N-terminal region of NS5 (residues 320-368). Further, it is shown that this same region of NS5 is also recognized by the cellular nuclear import receptor importin-beta. The interaction between NS5 and importin-beta and competition by NS3 with the latter for the same binding site on NS5 were confirmed by pull-down assays. The direct interaction of importin-beta with NS5 has implications for the mechanism by which this normally cytoplasmic protein may be targetted to the nucleus.

  • 16.
    Johansson, Magnus
    et al.
    Örebro University, School of Medical Sciences.
    Frelin, Lars
    Laboratory Medicine, Karolinska Institutet, Stockholm, Sweden.
    Maravelia, Panagiota
    Laboratory Medicine, Karolinska Institutet, Stockholm, Sweden.
    Asghar, Naveed
    Melik, Wessam
    Örebro University, School of Medical Sciences.
    Caro-Perez, Noelia
    Laboratory Medicine, Karolinska Institutet, Stockholm, Sweden.
    Pasetto, Anna
    Laboratory Medicine, Karolinska Institutet, Stockholm, Sweden.
    Ahlen, Gustaf
    Laboratory Medicine, Karolinska Institutet, Stockholm, Sweden.
    Sallberg, Matti
    Laboratory Medicine, Karolinska Institutet, Stockholm, Sweden.
    Immunogenicity of a New Flaviviral-Based DNA Launched Suicidal Replicon for Protective Vaccination Against Hepatitis C2019In: Molecular Therapy, ISSN 1525-0016, E-ISSN 1525-0024, Vol. 27, no 4, p. 139-139Article in journal (Other academic)
  • 17. Johansson, Magnus
    et al.
    Nordlund, S
    Purification of P(II) and P(II)-UMP and in vitro studies of regulation of glutamine synthetase in Rhodospirillum rubrum1999In: Journal of Bacteriology, ISSN 0021-9193, E-ISSN 1098-5530, Vol. 181, no 20, p. 6524-6529Article in journal (Refereed)
    Abstract [en]

    The P(II) protein from Rhodospirillum rubrum was fused with a histidine tag, overexpressed in Escherichia coli, and purified by Ni(2+)-chelating chromatography. The uridylylated form of the P(II) protein could be generated in E. coli. The effects on the regulation of glutamine synthetase by P(II), P(II)-UMP, glutamine, and alpha-ketoglutarate were studied in extracts from R. rubrum grown under different conditions. P(II) and glutamine were shown to stimulate the ATP-dependent inactivation (adenylylation) of glutamine synthetase, which could be totally inhibited by alpha-ketoglutarate. Deadenylylation (activation) of glutamine synthetase required phosphate, but none of the effectors studied had any major effect, which is different from their role in the E. coli system. In addition, deadenylylation was found to be much slower than adenylylation under the conditions investigated.

  • 18. Johansson, Magnus
    et al.
    Nordlund, S
    Transcription of the glnB and glnA genes in the photosynthetic bacterium Rhodospirillum rubrum1996In: Microbiology, ISSN 1350-0872, E-ISSN 1465-2080, Vol. 142 ( Pt 5), p. 1265-1272Article in journal (Refereed)
    Abstract [en]

    The PII protein, encoded by glnB, has a central role in the control of nitrogen metabolism in nitrogen-fixing prokaryotes. The glnB gene of Rhodospirillum rubrum was isolated and sequenced. The deduced amino acid sequence had very high sequence identity to other PII proteins. The glnA gene, encoding glutamine synthetase, was located 135 bp downstream of glnB and was partially sequenced. glnB is cotranscribed with glnA from a promoter with high similarity to the sigma 54-dependent promoter consensus sequence. A putative sigma 70 promoter was also identified further upstream of glnB. Northern blotting analyses showed that in addition glnA is either transcribed from an unidentified promoter or, more likely, that the glnBA transcript is processed to give the glnA mRNA. The total level of the two transcripts was much higher in nitrogen-fixing cells than in ammonia-grown cells.

  • 19. Johansson, Magnus
    et al.
    Nordlund, S
    Uridylylation of the P(II) protein in the photosynthetic bacterium Rhodospirillum rubrum1997In: Journal of Bacteriology, ISSN 0021-9193, E-ISSN 1098-5530, Vol. 179, no 13, p. 4190-4194Article in journal (Refereed)
    Abstract [en]

    The regulatory protein P(II) has been studied in great detail in enteric bacteria; however, its function in photosynthetic bacteria has not been clearly established. As a number of these bacteria have been shown to regulate nitrogenase activity by a metabolic control system, it is of special interest to establish the role of P(II) in these diazotrophs. In this study, we show that P(II) in Rhodospirillum rubrum is modified in response to the N status in the cell and that addition of ammonium or glutamine leads to demodification. We also provide evidence that P(II) is uridylylated. In addition, we show that not only these compounds but also NAD+ promotes demodification of P(II), which is of particular interest as this pyridine nucleotide has been shown to act as a switch-off effector of nitrogenase. Demodification of P(II) by ammonium or NAD+ did not occur in cultures treated with an inhibitor of glutamine synthetase (methionine sulfoximine), whereas treatment with the glutamate synthase inhibitor 6-diazo-5-oxo-norleucine led to total demodification of P(II) without any other addition. The results indicate that P(II) probably is not directly involved in darkness switch-off of nitrogenase but that a role in ammonium switch-off cannot be excluded.

  • 20. Lindblad, A
    et al.
    Jansson, J
    Brostedt, E
    Johansson, Magnus
    Hellman, U
    Nordlund, S
    Identification and sequence of a nifJ-like gene in Rhodospirillum rubrum: partial characterization of a mutant unaffected in nitrogen fixation1996In: Molecular Microbiology, ISSN 0950-382X, E-ISSN 1365-2958, Vol. 20, no 3, p. 559-568Article in journal (Refereed)
    Abstract [en]

    A nifJ-like gene was identified in the photosynthetic purple non-sulphur bacterium Rhodospirillum rubrum. A DNA segment hybridizing to Klebsiella pneumoniae nifJ was isolated, the gene was inactivated, and a mutant strain, SNJ-1, was constructed by allele replacement. The mutation was confirmed by DNA sequencing. Northern blotting and by the lack of pyruvate oxidoreductase activity. This is the first report of a nifJ-like gene in photosynthetic bacteria. Unexpectedly, SNJ-1 was capable of nitrogen fixation, and growth was similar to the wild-type strain under all conditions investigated. Therefore, this is also the first demonstration that a nifJ homologue, when present, is not essential for nitrogen fixation in a diazotroph. The nifJ-like gene was sequenced and found to have considerable similarity to published nifJ gene sequences from other organisms. By primer extension, the initiation site for transcription was located, and a typical sigma 54 promoter sequence was identified.

  • 21.
    Maravelia, Panagiota
    et al.
    Division of Clinical Microbiology, Department of Laboratory Medicine, Karolinska Institutet, Stockholm, Sweden.
    Frelin, Lars
    Division of Clinical Microbiology, Department of Laboratory Medicine, Karolinska Institutet, Stockholm, Sweden.
    Ni, Yi
    Department of Molecular Virology, University of Heidelberg, Heidelberg, Germany.
    Pérez, Noelia Caro
    Division of Clinical Microbiology, Department of Laboratory Medicine, Karolinska Institutet, Stockholm, Sweden.
    Ahlén, Gustaf
    Division of Clinical Microbiology, Department of Laboratory Medicine, Karolinska Institutet, Stockholm, Sweden.
    Jagya, Neetu
    Division of Clinical Microbiology, Department of Laboratory Medicine, Karolinska Institutet, Stockholm, Sweden.
    Verch, Georg
    Department of Molecular Virology, University of Heidelberg, Heidelberg, Germany.
    Verhoye, Lieven
    Laboratory of Liver Infectious Diseases, Ghent University, Gent, Belgium.
    Pater, Lena
    Division of Clinical Microbiology, Department of Laboratory Medicine, Karolinska Institutet, Stockholm, Sweden.
    Johansson, Magnus
    Örebro University, School of Medical Sciences.
    Pasetto, Anna
    Division of Clinical Microbiology, Department of Laboratory Medicine, Karolinska Institutet, Stockholm, Sweden.
    Meuleman, Philip
    Laboratory of Liver Infectious Diseases, Ghent University, Gent, Belgium.
    Urban, Stephan
    Department of Molecular Virology, University of Heidelberg, Heidelberg, Germany.
    Sällberg, Matti
    Division of Clinical Microbiology, Department of Laboratory Medicine, Karolinska Institutet, Stockholm, Sweden.
    Blocking entry of hepatitis B and D viruses to hepatocytes as a novel immunotherapy for treating chronic infections2020In: Journal of Infectious Diseases, ISSN 0022-1899, E-ISSN 1537-6613, article id jiaa036Article in journal (Refereed)
    Abstract [en]

    BACKGROUND: Chronic hepatitis B and D virus (HBV/HDV) infections can cause cancer. Current HBV therapy using nucleoside analogues (NAs) is life-long and reduces but does not eliminate the risk of cancer. A hallmark of chronic hepatitis B is a dysfunctional HBV-specific T cell response. We therefore designed an immunotherapy driven by naïve healthy T cells specific for the HDV antigen (HDAg) to bypass the need for HBV-specific T cells in order to prime PreS1-specific T cells and PreS1 antibodies blocking HBV entry.

    METHODS: Ten combinations of PreS1 and/or HDAg sequences were evaluated for induction of PreS1 antibodies and HBV- and HDV- specific T cells in vitro and in vivo. Neutralization of HBV by PreS1-specific murine and rabbit antibodies was evaluated in cell culture, and rabbit anti-PreS1 were tested for neutralization of HBV in mice repopulated with human hepatocytes.

    RESULTS: The best vaccine candidate induced T cells to PreS1 and HDAg, and PreS1 antibodies blocking HBV entry in vitro. Importantly, adoptive transfer of PreS1 antibodies prevented, or modulated, HBV infection after a subsequent challenge in humanized mice.

    CONCLUSION: We here describe a novel immunotherapy for chronic HBV/HDV that targets viral entry to complement NAs and coming therapies inhibiting viral maturation.

  • 22. Melik, W
    et al.
    Nilsson, A S
    Johansson, Magnus
    Detection strategies of tick-borne encephalitis virus in Swedish Ixodes ricinus reveal evolutionary characteristics of emerging tick-borne flaviviruses2007In: Archives of Virology, ISSN 0304-8608, E-ISSN 1432-8798, Vol. 152, no 5, p. 1027-1034Article in journal (Refereed)
    Abstract [en]

    The flaviviral tick-borne encephalitis virus (TBEV) is a human pathogen having significant impact on public health. The geographical distribution of TBEV and TBEV-like viruses is increasing, which makes it important to characterise the natural virus populations. Here we present four RT-PCR strategies designed for detection of broad types of tick-borne flaviviruses. Sequence information on more than 32% of a TBEV genome was generated from a small pool of ticks collected in the Stockholm archipelago on the island of Torö. The sequences were characterised and compared with those of other tick-borne flaviviruses, which classified the virus as Western European TBEV.

  • 23.
    Melik, Wessam
    et al.
    School of Life Sciences, Södertörn University, Huddinge, Sweden.
    Ellencrona, Karin
    School of Life Sciences, Södertörn University, Huddinge, Sweden.
    Wigerius, Michael
    School of Life Sciences, Södertörn University, Huddinge, Sweden; Department of Pharmacology, Faculty of Medicine, Dalhousie University, Halifax NS, Canada.
    Hedström, Christer
    School of Life Sciences, Södertörn University, Huddinge, Sweden.
    Elväng, Annelie
    School of Life Sciences, Södertörn University, Huddinge, Sweden.
    Johansson, Magnus
    Örebro University, School of Medicine, Örebro University, Sweden. School of Life Sciences, Södertörn University, Huddinge, Sweden.
    Two PDZ binding motifs within NS5 have roles in Tick-borne encephalitis virus replication2012In: Virus Research, ISSN 0168-1702, E-ISSN 1872-7492, Vol. 169, no 1, p. 54-62Article in journal (Refereed)
    Abstract [en]

    The flavivirus genus includes important human neurotropic pathogens like Tick-borne encephalitis virus (TBEV) and West-Nile virus (WNV). Flavivirus replication occurs at replication complexes, where the NS5 protein provides both RNA cap methyltransferase and RNA-dependent RNA polymerase activities. TBEVNS5 contains two PDZ binding motifs (PBMs) important for specific targeting of human PDZ proteins including Scribble, an association important for viral down regulation of cellular defense systems and neurite outgrowth. To determine whether the PBMs of TBEVNS5 affects virus replication we constructed a DNA based sub-genomic TBEV replicon expressing firefly luciferase. The PBMs within NS5 were mutated individually and in concert and the replicons were assayed in cell culture. Our results show that the replication rate was impaired in all mutants, which indicates that PDZ dependent host interactions influence TBEV replication. We also find that the C-terminal PBMs present in TBEVNS5 and WNVNS5 are targeting various human PDZ domain proteins. TBEVNS5 has affinity to Zonula occludens-2 (ZO-2), GIAP C-terminus interacting protein (GIPC), calcium/calmodulin-dependent serine protein kinase (CASK), glutamate receptor interacting protein 2, (GRIP2) and Interleukin 16 (IL-16). A different pattern was observed for WNVNS5 as it associate with a broader repertoire of putative host PDZ proteins.

  • 24.
    Rytkonen, Paulina
    et al.
    Department of Life Sciences, Södertörn University, Huddinge, Sweden.
    Bonow, Madeleine
    Department of Life Sciences, Södertörn University, Huddinge, Sweden.
    Johansson, Magnus
    Örebro University, School of Medicine, Örebro University, Sweden. Department of Life Sciences, Södertörn University, Huddinge, Sweden.
    Persson, Ylva
    The National Veterinary Institute, Uppsala, Sweden.
    Goat cheese production in Sweden: a pioneering experience in the re-emergence of local food2013In: Acta Agriculturae Scandinavica - Section B, ISSN 0906-4710, E-ISSN 1651-1913, Vol. 63, no Suppl. 1, p. 38-46Article in journal (Refereed)
    Abstract [en]

    The re-emergence and modernization of traditional goat-cheese production in Jämtland led to the articulation of a localized agri-food system that represents the frontline of the return and reinforcement of local food in Sweden. Already in the 1970s, some initiatives were undertaken to formalize the productive activities of this branch and to improve the product quality. The most important project was the articulation of a cooperative that, unlike all other Swedish cooperatives, engaged its members in the development of a joint trademark, development of a standardized assortment, common marketing efforts and finding creative solutions for infrastructure problems. Despite the overall success, we also found some downsides. Producing goat cheese requires that at least two people are involved, because the workload often leads to body injuries and illness for people working alone. By studying the institutional frameworks, rules and regulations, the economic function and entrepreneurial dynamics, and the dynamics of knowledge and competences, the article highlights how and why farm dairies in Jamtland became reinforced and modernized. This grasps both the actions of individual economic agents and their interaction with their environment. A special emphasis was put on the role of regional authorities in this process. Even though many obstacles have been removed and the trade has found successful ways to solve strategic issues concerning product development and marketing, there are still important structural shortcomings that might decrease the profitability and endanger the future development of the trade. There is a lack of experience and infrastructure to solve more complex problems like animal healthand the potential risks related to the consumption of unpasteurized cheese and the increasing incidence of Tick-Borne Encephalitis (TBE).

  • 25.
    Tran, Pham Tue Hung
    et al.
    Örebro University, School of Medical Sciences.
    Asghar, Naveed
    Örebro University, School of Medical Sciences.
    Karlsson, Anders
    Department of Clinical Microbiology, Sahlgrenska University Hospital, Gothenburg, Sweden; Nanoxis Consulting AB, Gothenburg, Sweden.
    Karlsson, Roger
    Department of Clinical Microbiology, Sahlgrenska University Hospital, Gothenburg, Sweden; Nanoxis Consulting AB, Gothenburg, Sweden.
    Johansson, Magnus
    Örebro University, School of Medical Sciences.
    Melik, Wessam
    Örebro University, School of Medical Sciences.
    Screening of host proteins interacting with Kunjin, Langat, Zikareplication complex2019In: 16th Smögen Summer Symposium on Virology, 2019Conference paper (Refereed)
    Abstract [en]

    During infection and eclipse time, Flaviviruses induce invagination of the endoplasmic reticulum (ER) membrane to form compartments, protecting their viral replication complex. The rearrangements of ER membrane require modifications in ER membrane lipid constituents or binding of proteins to bend the membrane. Indeed, it has been implicated that both KUNV and DENV NS1, NS2A, NS4A, NS4B proteins could induce membrane remodelings. However, it is not well known whether host proteins can also participate in the formation and maintenance of these compartments.In this project, we aimed to identify host proteins interacting with Kunjin, Langat, Zika replication complex. These proteins may function for ER invagination during Flavivirus infection. We used human adenocarcinoma epithelial A549 cells as a cell model, mosquito-borne Zika, Kunjin virus, and tick-borne Langat virus as virus models. After virus infections, the ER membranes from infected and non-infected cells were harvested using ultracentrifuge with a sucrose gradient. Proteins from these ERs were identified using mass spectrometry. We compared the differences between the ER proteomes to identify host candidate proteins that can cause the RC formation. To narrows the list of true candidate proteins, we attempted to enrich the RC-containing fractions by doing co-immuno precipitation. We are doing TMT-MS to identify and quantify the host proteins from Co-IP elutions. The functions of these proteins will be characterized by using molecular techniques.

  • 26.
    Tran, Pham Tue Hung
    et al.
    Örebro University, School of Medical Sciences.
    Asghar, Naveed
    Örebro University, School of Medical Sciences.
    Karlsson, Roger
    Department ofClinical Microbiology, Sahlgrenska University Hospital, Gothenburg, Sweden; NanoxisConsulting AB, Gothenburg, Sweden.
    Karlsson, Anders
    Department ofClinical Microbiology, Sahlgrenska University Hospital, Gothenburg, Sweden; NanoxisConsulting AB, Gothenburg, Sweden.
    Johansson, Magnus
    Örebro University, School of Medical Sciences.
    Melik, Wessam
    Örebro University, School of Medical Sciences.
    Identification and characterization of host proteins inducing the endoplasmic reticulum invagination during Flavivirus infection2019In: Positive-Strand RNA Viuses, 2019, p. 280-280Conference paper (Refereed)
    Abstract [en]

    When Flaviviruses infect host cells, they can induce invagination of endoplasmic reticulum (ER) membrane to form vesicle-like compartments. These unique structures are hypothetical to facilitate the viral replication by reducing diffusion of virus replication machinery and viral RNA, providing a scaffold to anchor the replication complex, and protecting viral RNA from host cell intrinsic surveillance. 

    The rearrangements of ER membrane to form these replication compartments (RCs) require modifications in its lipid constituents or binding of proteins to the membrane. Flaviviruses, indeed, use their proteins to generate RCs. It has been implicated that both KUNV and DENV viral NS1, NS2A, NS4A, NS4B proteins could induce membrane remodelings. However, it is recondite whether host proteins can also participate in the formation and maintenance of RCs.

    In this project, we aimed to identify and characterize of host proteins inducing RC generation during Flavivirus infections. We used A549 as a cell model, and mosquito-borne Zika and Kunjin virus, and tick-borne Langat virus as virus models. After virus infections, ER membranes were harvested using ultracentrifuge with a sucrose gradient. Proteins from these ERs were identified using mass spectrometry. We compared the differences between the ER proteomes of infected cells and non-infected cells to identify host candidate proteins that can cause the RC formation.  We are attempting to enrich the RC-containing fractions and identifying proteins here, which narrows the list of true candidate proteins. The candidate proteins then will be characterized by using molecular techniques such as gene knock down, overexpression, and microscopy techniques.

    Download full text (pdf)
    Identification and characterization of host proteins inducing the endoplasmic reticulum invagination during Flavivirus infection
  • 27.
    Tran, Pham Tue Hung
    et al.
    Örebro University, School of Medical Sciences.
    Asghar, Naveed
    Örebro University, School of Medical Sciences.
    Karlsson, Roger
    Department ofClinical Microbiology, Sahlgrenska University Hospital, Gothenburg, Sweden; NanoxisConsulting AB, Gothenburg, Sweden.
    Karlsson, Anders
    Department ofClinical Microbiology, Sahlgrenska University Hospital, Gothenburg, Sweden; NanoxisConsulting AB, Gothenburg, Sweden.
    Johansson, Magnus
    Örebro University, School of Medical Sciences.
    Melik, Wessam
    Örebro University, School of Medical Sciences.
    Screening of host proteins interacting with Kunjin, Langat, Zika replication complex2019In: Positive-Strand Rna Viuses, 2019Conference paper (Refereed)
    Abstract [en]

    When Flaviviruses infect host cells, they can induce invagination of endoplasmic reticulum (ER) membrane to form vesicle-like compartments. These unique structures are hypothetical to facilitate the viral replication by reducing diffusion of virus replication machinery and viral RNA, providing a scaffold to anchor the replication complex, and protecting viral RNA from host cell intrinsic surveillance. 

    The rearrangements of ER membrane to form these replication compartments (RCs) require modifications in its lipid constituents or binding of proteins to the membrane. Flaviviruses, indeed, use their proteins to generate RCs. It has been implicated that both KUNV and DENV viral NS1, NS2A, NS4A, NS4B proteins could induce membrane remodelings. However, it is recondite whether host proteins can also participate in the formation and maintenance of RCs.

    In this project, we aimed to identify and characterize of host proteins inducing RC generation during Flavivirus infections. We used A549 as a cell model, and mosquito-borne Zika and Kunjin virus, and tick-borne Langat virus as virus models. After virus infections, ER membranes were harvested using ultracentrifuge with a sucrose gradient. Proteins from these ERs were identified using mass spectrometry. We compared the differences between the ER proteomes of infected cells and non-infected cells to identify host candidate proteins that can cause the RC formation.  We are attempting to enrich the RC-containing fractions and identifying proteins here, which narrows the list of true candidate proteins. The candidate proteins then will be characterized by using molecular techniques such as gene knock down, overexpression, and microscopy techniques.

    Download full text (pdf)
    Screening of host proteins interacting with Kunjin, Langat, Zikareplication complex
  • 28. Vasudevan, S G
    et al.
    Johansson, Magnus
    Brooks, A J
    Llewellyn, L E
    Jans, D A
    Characterisation of inter- and intra-molecular interactions of the dengue virus RNA dependent RNA polymerase as potential drug targets2001In: Il Farmaco, ISSN 0014-827X, E-ISSN 1879-0569, Vol. 56, no 1-2, p. 33-36Article in journal (Refereed)
    Abstract [en]

    Our research is directed towards enhancing the understanding of the molecular biology of dengue virus replication with the ultimate goal being to develop novel antiviral strategies based on preventing critical inter- or intra-molecular interactions required for the normal virus life cycle. The viral RNA-dependent RNA polymerase (NS5) and the viral helicase (NS3) interaction offers a possible target for inhibitors to bind and prevent replication. In this study the yeast-two hybrid system was used to show that a small region of NS5 interacts with NS3, and also with the cellular nuclear transport receptor importin-beta. Furthermore, intramolecular interaction between the two putative domains of NS5 can also be detected by the yeast two-hybrid assay. We have also modified the colony lift assay for the beta-galactosidase reporter activity in intact yeast cells which reflects the strength of interaction between two proteins to a microtiter plate format. This assay offers a unique opportunity to screen for small molecule compounds that block physiologically important interactions.

  • 29.
    Wallenhammar, Amélie
    et al.
    Örebro University, School of Medical Sciences.
    Lindqvist, Richard
    Department of Clinical Microbiology, Virology, Laboratory for Molecular Infection Medicine Sweden (MIMS), Umeå University, Umeå, Sweden.
    Asghar, Naveed
    Örebro University, School of Medical Sciences.
    Gunaltay, Sezin
    School of Medical Sciences, Faculty of Medicine and Health, Örebro University, Örebro, Sweden; Lydia Becker Institiute of Immunology and Inflammation, Faculty of Biology, Medicine and Health, Manchester Academic Health Science Centre, University of Manchester, Manchester, UK.
    Fredlund, Hans
    Department of Laboratory Medicine, Faculty of Medicine and Health, Örebro University, Örebro, Sweden.
    Davidsson, Åke
    Örebro University, School of Medical Sciences.
    Andersson, Sören
    Örebro University, School of Medical Sciences.
    Överby, Anna K.
    Department of Clinical Microbiology, Virology, Laboratory for Molecular Infection Medicine Sweden (MIMS), Umeå University, Umeå, Sweden.
    Johansson, Magnus
    Örebro University, School of Medical Sciences.
    Revealing new tick-borne encephalitis virus foci by screening antibodies in sheep milk2020In: Parasites & Vectors, ISSN 1756-3305, E-ISSN 1756-3305, Vol. 13, no 1, article id 185Article in journal (Refereed)
    Abstract [en]

    BACKGROUND: Tick distribution in Sweden has increased in recent years, with the prevalence of ticks predicted to spread towards the northern parts of the country, thus increasing the risk of tick-borne zoonoses in new regions. Tick-borne encephalitis (TBE) is the most significant viral tick-borne zoonotic disease in Europe. The disease is caused by TBE virus (TBEV) infection which often leads to severe encephalitis and myelitis in humans. TBEV is usually transmitted to humans via tick bites; however, the virus can also be excreted in the milk of goats, sheep and cattle and infection may then occur via consumption of unpasteurised dairy products. Virus prevalence in questing ticks is an unreliable indicator of TBE infection risk as viral RNA is rarely detected even in large sample sizes collected at TBE-endemic areas. Hence, there is a need for robust surveillance techniques to identify emerging TBEV risk areas at early stages.

    METHODS: Milk and colostrum samples were collected from sheep and goats in Örebro County, Sweden. The milk samples were analysed for the presence of TBEV antibodies by ELISA and validated by western blot in which milk samples were used to detect over-expressed TBEV E-protein in crude cell extracts. Neutralising titers were determined by focus reduction neutralisation test (FRNT). The stability of TBEV in milk and colostrum was studied at different temperatures.

    RESULTS: In this study we have developed a novel strategy to identify new TBEV foci. By monitoring TBEV antibodies in milk, we have identified three previously unknown foci in Örebro County which also overlap with areas of TBE infection reported during 2009-2018. In addition, our data indicates that keeping unpasteurised milk at 4 °C will preserve the infectivity of TBEV for several days.

    CONCLUSIONS: Altogether, we report a non-invasive surveillance technique for revealing risk areas for TBE in Sweden, by detecting TBEV antibodies in sheep milk. This approach is robust and reliable and can accordingly be used to map TBEV "hotspots". TBEV infectivity in refrigerated milk was preserved, emphasising the importance of pasteurisation (i.e. 72 °C for 15 s) prior to consumption.

  • 30.
    Werme, Karin
    et al.
    Södertörn University College, Huddinge, Sweden; Stockholm University, Stockholm, Sweden.
    Wigerius, Michael
    Södertörn University College, Huddinge, Sweden; Stockholm University, Stockholm, Sweden.
    Johansson, Magnus
    Södertörn University College, Huddinge, Sweden.
    Tick-borne encephalitis virus NS5 associates with membrane protein scribble and impairs interferon-stimulated JAK-STAT signalling.2008In: Cellular Microbiology, ISSN 1462-5814, E-ISSN 1462-5822, Vol. 10, no 3, p. 696-712Article in journal (Refereed)
    Abstract [en]

    Tick-borne encephalitis virus (TBEV) NS5 protein is a multifunctional RNA-dependent RNA polymerase that is indispensable for viral replication. TBEV is considered to be highly neurovirulent and can cause lethal encephalitis. In this study, we demonstrate a novel interaction between TBEV NS5 and the PDZ protein scribble (hScrib) affecting interferon (IFN) type I and II mediated JAK-STAT signalling. The sequence of TBEV NS5 interacting with hScrib was identified using extensive site-directed mutagenesis analysis. Two consecutive mutations in the methyltransferase (MTase) domain of NS5 were found to disrupt binding to hScrib. Colocalization studies with hScrib demonstrated that TBEV NS5 was present at the plasma membrane of mammalian cells. To address the role of viral interference with the IFN response, NS5 proteins were expressed in IFN-stimulated cells. While TBEV NS5 substantially blocked phosphorylation of STAT1, a mutated NS5 protein defective in hScrib binding failed to inhibit JAK-STAT signalling correctly. Furthermore, hScrib knock-down resulted in re-localization of NS5 to intracellular locations and abrogated the impaired STAT1 phosphorylation. These results define the TBEV NS5 protein in concert with hScrib as an antagonist of the IFN response, by demonstrating a correlation between the association and JAK-STAT interference.

  • 31.
    Wigerius, Michael
    et al.
    Södertörn University, Huddinge, Sweden; Dalhousie University, Halifax, Canada.
    Asghar, Naveed
    Södertörn University, Huddinge, Sweden.
    Melik, Wessam
    Södertörn University, Huddinge, Sweden.
    Johansson, Magnus
    Örebro University, School of Health and Medical Sciences, Örebro University, Sweden.
    Scribble controls NGF-mediated neurite outgrowth in PC12 cells2013In: European Journal of Cell Biology, ISSN 0171-9335, E-ISSN 1618-1298, Vol. 92, no 6-7, p. 213-221Article in journal (Refereed)
    Abstract [en]

    Neurite outgrowth is mediated by dynamic changes of the cytoskeleton and is largely controlled by Rho GTPases and their regulators. Here, we show that the polarity protein Scribble controls PC12 cell neurite outgrowth in response to nerve growth factor. Scribble knockdown decreases neurite numbers and increases neurite length. This effect is linked to TrkA the cognate receptor for NGF as pharmacological inhibition of phosphorylated TrkA (pTrkA) reduces Scribble expression. Moreover, Scribble forms a complex with the MAPK components ERK1/2 in a growth factor dependent manner. In RNAi experiments where Scribble expression is efficiently depleted sustained ERK1/2 phosphorylation is reduced. Conversely, siRNA with intermediate Scribble silencing efficiency fails to match this effect indicating that ERK1/2 activation depends on basic Scribble protein levels. Finally, Scribble translocates to the plasma membrane in response to growth factor where it complexes with HRas and Rac1 suggesting that the phenotype activated by loss of Scribble may be a result of altered GTPase activity. Together, these results demonstrate a novel role for Scribble in neurite outgrowth of PC12 cells. (c) 2013 Elsevier GmbH. All rights reserved.

  • 32. Wigerius, Michael
    et al.
    Melik, Wessam
    Elväng, Annelie
    Johansson, Magnus
    Södertörns högskola, Huddinge, Sweden.
    Rac1 and Scribble are targets for the arrest of neurite outgrowth by TBE virus NS52010In: Molecular and Cellular Neuroscience, ISSN 1044-7431, E-ISSN 1095-9327, Vol. 44, no 3, p. 260-271Article in journal (Refereed)
    Abstract [en]

    Tick-borne encephalitis virus (TBEV) causes extensive CNS disease in humans known as TBE, however, relatively little is known of the molecular mechanisms for its progress. Here, we now show that TBEV produces defects in neuronal development of PC12 cells through a function of the viral NS5 protein. The methyltransferase domain of NS5 is critical and sufficient for restriction of nerve growth factor induced neurite outgrowth. This effect is reversed by expression of NS5 mutants unable to bind Scribble and unexpectedly, in Scribble depleted cells with binding-competent NS5. Furthermore, we also demonstrate that the Rho GTPase Rac1 and the guanine nucleotide-exchange factor, betaPIX are outcompeted by NS5 for binding to Scribble, linking to effects on neurite outgrowth by TBEV. Together, these findings provide the first experimental evidence that Rac1 and betaPIX are indirect targets of NS5 acting through the multifunctional polarity protein Scribble to oppose neuronal differentiation. In conclusion, our results offer a potential mechanism by which TBEV alters neuronal circuitry and opens new avenues for therapeutic interventions.

1 - 32 of 32
CiteExportLink to result list
Permanent link
Cite
Citation style
  • apa
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Other style
More styles
Language
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Other locale
More languages
Output format
  • html
  • text
  • asciidoc
  • rtf