| 1. |
- Werme, Karin, et al.
(författare)
-
Tick-borne encephalitis virus NS5 associates with membrane protein scribble and impairs interferon-stimulated JAK-STAT signalling.
- 2008
-
Ingår i: Cellular Microbiology. - Oxford : Hindawi Limited. - 1462-5814 .- 1462-5822. ; 10:3, s. 696-712
-
Tidskriftsartikel (refereegranskat)abstract
- 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.
|
|
| 2. |
- Gustafsson, Kerstin, et al.
(författare)
-
Direct and indirect effects of the fungicide azoxystrobin in outdoor brackish water microcosms
- 2010
-
Ingår i: Ecotoxicology. - : Springer Science and Business Media LLC. - 0963-9292 .- 1573-3017. ; 19:2, s. 431-444
-
Tidskriftsartikel (refereegranskat)abstract
- The effects of the strobilurin fungicide azoxystrobin were studied in brackish water microcosms, with natural plankton communities and sediment. Two experiments were conducted: Experiment 1 (nominal conc. 0, 15 and 60 mu g/L, 24-L outdoor microcosms for 21 days) and a second, follow-up, Experiment 2 (nominal conc. 0, 3, 7.5, 15 mu g/L, 4-L indoor microcosms for 12 days). The microcosms represent a simplified brackish water community found in shallow semi-enclosed coastal areas in agricultural districts in the Baltic Sea region. Measured water concentrations of the fungicide (Experiment 1) were, on average, 83 and 62% of nominal concentrations directly after application, and 25 and 30% after 21 days, for the low and high dose treatments, respectively, corresponding to mean DT50-values of 15.1 and 25.8 days, for low and high dose treatments, respectively. In Experiment 1, direct toxic effects on calanoid copepods at both test concentrations were observed. Similarly, in Experiment 2, the copepod abundance was significantly reduced at all tested concentrations. There were also significant secondary effects on zooplankton and phytoplankton community structure, standing stocks and primary production. Very few ecotoxicological studies have investigated effects of plant protection products on Baltic organisms in general and effects on community structure and function specifically. Our results show that azoxystrobin is toxic to brackish water copepods at considerably lower concentrations than previously reported from single species tests on freshwater crustaceans, and that direct toxic effects on this ecologically important group may lead to cascade effects altering lower food webs and ecosystem functioning.
|
|
| 3. |
- Wigerius, Michael, et al.
(författare)
-
Scribble controls NGF-mediated neurite outgrowth in PC12 cells
- 2013
-
Ingår i: European Journal of Cell Biology. - : Urban & Fischer. - 0171-9335 .- 1618-1298. ; 92:6-7, s. 213-221
-
Tidskriftsartikel (refereegranskat)abstract
- 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.
|
|
| 4. |
- Löfmark, Sonja, et al.
(författare)
-
Clindamycin-induced enrichment and long-term persistence of resistant Bacteroides spp. and resistance genes
- 2006
-
Ingår i: Journal of Antimicrobial Chemotherapy. - : Oxford University Press (OUP). - 0305-7453 .- 1460-2091. ; 58:6, s. 1160-1167
-
Tidskriftsartikel (refereegranskat)abstract
- Objectives: The aim was to study the long-term consequences of 1 week clindamycin administration regarding selection and persistence of resistance, resistance determinants and diversity of the Bacteroides spp. in the intestinal microflora. Methods: A total of 1306 Bacteroides isolates were collected from constitutively cultured faecal samples during a 2 year period from eight healthy volunteers. The strains were identified by biochemical and genotyping methods. MIC values were determined by the agar dilution method and presence of resistance genes was screened by real-time PCR. Results: Ecological changes in the intestinal microflora persisting up to 24 months were recorded after a 7 day clindamycin administration to four healthy volunteers. Compared to a control group, not exposed to clindamycin, an enrichment and stabilization of resistant Bacteroides strains and resistance determinants were discovered up to 2 years after clindamycin exposure. Conclusions: The results indicate that even a short-term antibiotic administration can cause long-term alterations in the commensal microbiota of individual subjects, detectable 2 years after dosing. The recorded selection and persistence of resistant strains and resistance genes, illustrates the importance of increasing our knowledge of the role of the abundant intestinal microbial community as a reservoir for spread of resistance.
|
|
| 5. |
- Kellner, Martin, 1978-, et al.
(författare)
-
Developmental exposure to the SSRI citalopram causes long-lasting behavioural effects in the three-spined stickleback (Gasterosteus aculeatus)
- 2018
-
Ingår i: Ecotoxicology. - : Springer Science and Business Media LLC. - 0963-9292 .- 1573-3017. ; 27:1, s. 12-22
-
Tidskriftsartikel (refereegranskat)abstract
- Selective Serotonin re-uptake inhibitors (SSRIs) are a class of psychotropic drugs used to treat depression in both adolescents and pregnant or breast-feeding mothers as well as in the general population. Recent research on rodents points to long-lasting behavioural effects of pre- and perinatal exposure to SSRIs which last into adulthood. In fish however, studies on effects of developmental exposure to SSRIs appears to be non-existent. In order to study effects of developmental SSRI exposure in fish, three-spine sticklebacks were exposed to 1.5 A mu g/l of the SSRI citalopram in the ambient water for 30 days, starting two days post-fertilisation. After approximately 100 days of remediation in clean water the fish were put through an extensive battery of behavioural tests. Feeding behaviour was tested as the number of bites against a piece of food and found to be increased in the exposed fish. Aggression levels were measured as the number of bites against a mirror image during 10 min and was also found to be significantly increased in the exposed fish. Novel tank behaviour and locomotor activity was tested in an aquarium that had a horizontal line drawn half-way between the bottom and the surface. Neither the latency to the first transition to the upper half, nor the number of transitions or the total time spent in the upper half was affected by treatment. Locomotor activity was significantly reduced in the exposed fish. The light/dark preference was tested in an aquarium where the bottom and walls were black on one side and white on the other. The number of transitions to the white side was significantly reduced in the exposed fish but there was no effect on the latency to the first transition or the total time spent in the white half. The results in the current study indicate that developmental SSRI exposure causes long-lasting behavioural effects in fish and contribute to the existing knowledge about SSRIs as environmental pollutants.
|
|
| 6. |
- Broman, Elias, 1985-, et al.
(författare)
-
Long-Term Pollution Does Not Inhibit Denitrification and DNRA by Adapted Benthic Microbial Communities
- 2023
-
Ingår i: Microbial Ecology. - : Elsevier. - 0095-3628 .- 1432-184X. ; 86, s. 2357-2372
-
Tidskriftsartikel (refereegranskat)abstract
- Denitrification in sediments is a key microbial process that removes excess fixed nitrogen, while dissimilatory nitrate reduction to ammonium (DNRA) converts nitrate to ammonium. Although microorganisms are responsible for essential nitrogen (N) cycling, it is not yet fully understood how these microbially mediated processes respond to toxic hydrophobic organic compounds (HOCs) and metals. In this study, we sampled long-term polluted sediment from the outer harbor of Oskarshamn (Baltic Sea), measured denitrification and DNRA rates, and analyzed taxonomic structure and N-cycling genes of microbial communities using metagenomics. Results showed that denitrification and DNRA rates were within the range of a national reference site and other unpolluted sites in the Baltic Sea, indicating that long-term pollution did not significantly affect these processes. Furthermore, our results indicate an adaptation to metal pollution by the N-cycling microbial community. These findings suggest that denitrification and DNRA rates are affected more by eutrophication and organic enrichment than by historic pollution of metals and organic contaminants.
|
|
| 7. |
- Brooks, Andrew J, et al.
(författare)
-
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 signals
- 2002
-
Ingår i: Journal of Biological Chemistry. - 0021-9258 .- 1083-351X. ; 277:39, s. 36399-36407
-
Tidskriftsartikel (refereegranskat)abstract
- 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.
|
|
| 8. |
- Ellencrona, Karin, et al.
(författare)
-
Flavivirus NS5 associates with host-cell proteins zonula occludens-1 (ZO-1) and regulating synaptic membrane exocytosis-2 (RIMS2) via an internal PDZ binding mechanism
- 2009
-
Ingår i: Biological chemistry (Print). - Berlin : Walter de Gruyter. - 1431-6730 .- 1437-4315. ; 390:4, s. 319-323
-
Tidskriftsartikel (refereegranskat)abstract
- 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.
|
|
| 9. |
- Johansson, Magnus, et al.
(författare)
-
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, NS3
- 2001
-
Ingår i: Journal of General Virology. - : Microbiology Society. - 0022-1317 .- 1465-2099. ; 82:Pt 4, s. 735-745
-
Tidskriftsartikel (refereegranskat)abstract
- 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.
|
|
| 10. |
- Wigerius, Michael, et al.
(författare)
-
Rac1 and Scribble are targets for the arrest of neurite outgrowth by TBE virus NS5
- 2010
-
Ingår i: Molecular and Cellular Neuroscience. - : Elsevier BV. - 1044-7431 .- 1095-9327. ; 44:3, s. 260-271
-
Tidskriftsartikel (refereegranskat)abstract
- 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, βPIX 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 βPIX 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.
|
|
