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  • 1.
    Abreu-Vieira, Gustavo
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Fischer, Alexander W.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut. University of Hamburg, Germany.
    Mattsson, Charlotte
    de Jong, Jasper M. A.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Shabalina, Irina G.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Ryden, Mikael
    Laurencikiene, Jurga
    Arner, Peter
    Cannon, Barbara
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Nedergaard, Jan
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Petrovic, Natasa
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Cidea improves the metabolic profile through expansion of adipose tissue2015Ingår i: Nature Communications, ISSN 2041-1723, E-ISSN 2041-1723, Vol. 6, artikel-id 7433Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    In humans, Cidea (cell death-inducing DNA fragmentation factor alpha-like effector A) is highly but variably expressed in white fat, and expression correlates with metabolic health. Here we generate transgenic mice expressing human Cidea in adipose tissues (aP2-hCidea mice) and show that Cidea is mechanistically associated with a robust increase in adipose tissue expandability. Under humanized conditions (thermoneutrality, mature age and prolonged exposure to high-fat diet), aP2-hCidea mice develop a much more pronounced obesity than their wild-type littermates. Remarkably, the malfunctioning of visceral fat normally caused by massive obesity is fully overcome-perilipin 1 and Akt expression are preserved, tissue degradation is prevented, macrophage accumulation is decreased and adiponectin expression remains high. Importantly, the aP2-hCidea mice display enhanced insulin sensitivity. Our data establish a functional role for Cidea and suggest that, in humans, the association between Cidea levels in white fat and metabolic health is not only correlative but also causative.

  • 2. Acheva, Anna
    et al.
    Haghdoost, Siamak
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut. University of Caen Normandy, France.
    Sollazzo, Alice
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Launonen, Virpi
    Kamarainen, Meerit
    Presence of Stromal Cells Enhances Epithelial-to-Mesenchymal Transition (EMT) Induction in Lung Bronchial Epithelium after Protracted Exposure to Oxidative Stress of Gamma Radiation2019Ingår i: Oxidative Medicine and Cellular Longevity, ISSN 1942-0900, E-ISSN 1942-0994, Vol. 2019, artikel-id 4120379Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The aim of the study was to investigate the role of a microenvironment in the induction of epithelial-to-mesenchymal transition (EMT) as a sign of early stages of carcinogenesis in human lung epithelial cell lines after protracted low-dose rate gamma-radiation exposures. BEAS-2B and HBEC-3KT lung cell lines were irradiated with low-dose rate gamma-rays (Cs-137, 1.4 or 14 mGy/h) to 0.1 or 1 Gy with or without adding TGF-beta. TGF-beta-treated samples were applied as positive EMT controls and tested in parallel to find out if the radiation has a potentiating effect on the EMT induction. To evaluate the effect of the stromal component, the epithelial cells were irradiated in cocultures with stromal MRC-9 lung fibroblasts. On day 3 post treatment, the EMT markers: alpha-SMA, vimentin, fibronectin, and E-cadherin, were analyzed. The oxidative stress levels were evaluated by 8-oxo-dG analysis in both epithelial and fibroblast cells. The protracted exposure to low Linear Energy Transfer (LET) radiation at the total absorbed dose of 1 Gy was able to induce changes suggestive of EMT. The results show that the presence of the stromal component and its signaling (TGF-beta) in the cocultures enhances the EMT. Radiation had a minor cumulative effect on the TGF-beta-induced EMT with both doses. The oxidative stress levels were higher than the background in both epithelial and stromal cells post chronic irradiation (0.1 and 1 Gy); as for the BEAS-2B cell line, the increase was statistically significant. We suggest that the induction of EMT in bronchial epithelial cells by radiation requires more than single acute exposure and the presence of stromal component might enhance the effect through free radical production and accumulation.

  • 3.
    Alkemar, Gunnar
    Stockholms universitet, Naturvetenskapliga fakulteten, Wenner-Grens institut för experimentell biologi.
    Ribosome and ribosomal RNA Structure: An experimental and computational analysis of expansion segments in eukaryotic ribosomal RNA2008Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    Ribosomes are large ribonucleoprotein complexes which incorporate amino acids into peptide chains during translational process in all types of living cells. The eukaryotic ribosome is larger compared to its prokaryotic counterpart. The size differences are due to a larger protein part and that the rRNA contains eukaryote specific expansion segments (ES). Cryo-EM reconstruction has visualized many ES on the ribosomal surface which have given clues about function and structural features. However, the secondary structures of most ES are unknown or ill defined. In this thesis, the secondary and also to a certain extent the tertiary structures of several ES are determined by using computational methods and biochemical experimental techniques. The juxtaposition of ES6 close to ES3 in the Cryo-EM image of the yeast ribosome suggested that ES3 and ES6 might interact. A computational analysis of more than 2900 sequences shows that a complementary helical region of seven to nine contiguous base pairs can form between ES3 and ES6 in almost all analyzed sequences. Biochemical in situ experiments support the proposed interaction. Secondary structure models are presented for ES3 and ES6 in 18S rRNA and ES39 in 28S rRNA, where homologous structural elements could be modeled in the experimentally analyzed ribosomes from fungi, plants and mammals. The structure models were further supported by computational methods where the ES6 structure and the ES39 structure could be formed in more than 6000 and 900 sequences respectively. A tertiary structure model of ES3 and ES6 including the helical interaction is presented. An in vitro transcribed and folded ES6 sequence differed from that observed in situ, suggesting that chaperones, ribosomal proteins, and/or the tertiary rRNA interaction could be involved in the in vivo folding of ES6. An analysis of the similarities between ES39 structures suggests that it might be under selective constraint to preserve its secondary structure.

  • 4. Assadi, Ghazaleh
    et al.
    Vesterlund, Liselotte
    Bonfiglio, Ferdinando
    Mazzurana, Luca
    Cordeddu, Lina
    Schepis, Danika
    Mjösberg, Jenny
    Ruhrmann, Sabrina
    Fabbri, Alessia
    Vukojevic, Vladana
    Percipalle, Piergiorgio
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut. New York University Abu Dhabi, United Arab Emirates.
    Salomons, Florian A.
    Laurencikiene, Jurga
    Törkvist, Leif
    Halfvarson, Jonas
    D'Amato, Mauro
    Functional Analyses of the Crohn's Disease Risk Gene LACC12016Ingår i: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 11, nr 12, artikel-id e0168276Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Background Genetic variation in the Laccase (multicopper oxidoreductase) domain-containing 1 (LACC1) gene has been shown to affect the risk of Crohn's disease, leprosy and, more recently, ulcerative colitis and juvenile idiopathic arthritis. LACC1 function appears to promote fatty-acid oxidation, with concomitant inflammasome activation, reactive oxygen species production, and anti-bacterial responses in macrophages. We sought to contribute to elucidating LACC1 biological function by extensive characterization of its expression in human tissues and cells, and through preliminary analyses of the regulatory mechanisms driving such expression. Methods We implemented Western blot, quantitative real-time PCR, immunofluorescence microscopy, and flow cytometry analyses to investigate fatty acid metabolism-immune nexus (FAMIN; the LACC1 encoded protein) expression in subcellular compartments, cell lines and relevant human tissues. Gene-set enrichment analyses were performed to initially investigate modulatory mechanisms of LACC1 expression. A small-interference RNA knockdown in vitro model system was used to study the effect of FAMIN depletion on peroxisome function. Results FAMIN expression was detected in macrophage-differentiated THP-1 cells and several human tissues, being highest in neutrophils, monocytes/macrophages, myeloid and plasmacytoid dendritic cells among peripheral blood cells. Subcellular co-localization was exclusively confined to peroxisomes, with some additional positivity for organelle endomembrane structures. LACC1 co-expression signatures were enriched for genes involved in peroxisome proliferator-activated receptors (PPAR) signaling pathways, and PPAR ligands downregulated FAMIN expression in in vitro model systems. Conclusion FAMIN is a peroxisome-associated protein with primary role(s) in macrophages and other immune cells, where its metabolic functions may be modulated by PPAR signaling events. However, the precise molecular mechanisms through which FAMIN exerts its biological effects in immune cells remain to be elucidated.

  • 5.
    Attoff, Kristina
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Kertika, Dimitra
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Lundqvist, Jessica
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Oredsson, S.
    Forsby, Anna
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi. Stockholm Univ, Dept Neurochem, S-10691 Stockholm, Sweden.
    Acrylamide affects proliferation and differentiation of the neural progenitor cell line C17.2 and the neuroblastoma cell line SH-SY5Y2016Ingår i: Toxicology in Vitro, ISSN 0887-2333, E-ISSN 1879-3177, Vol. 35, s. 100-111Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Acrylamide is a well-known neurotoxic compound and people get exposed to the compound by food consumption and environmental pollutants. Since acrylamide crosses the placenta barrier, the fetus is also being exposed resulting in a risk for developmental neurotoxicity. In this study, the neural progenitor cell line C17.2 and the neuroblastoma cell line SH-SY5Y were used to study proliferation and differentiation as alerting indicators for developmental neurotoxicity. For both cell lines, acrylamide reduced the number of viable cells by reducing proliferation and inducing cell death in undifferentiated cells. Acrylamide concentrations starting at 10 fM attenuated the differentiation process in SH-SY5Y cells by sustaining cell proliferation and neurite outgrowth was reduced at concentrations from 10 pM. Acrylamide significantly reduced the number of neurons starting at 1 mu M and altered the ratio between the different phenotypes in differentiating C17.2 cell cultures. Ten micromolar of acrylamide also reduced the expression of the neuronal and astrocyte biomarkers. Although the neurotoxic concentrations in the femtomolar range seem to be specific for the SH-SY5Y cell line, the fact that micromolar concentrations of acrylamide seem to attenuate the differentiation process in both cell lines raises the interest to further investigations on the possible developmental neurotoxicity of acrylamide.

  • 6.
    Behm, Mikaela
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut. Stockholms universitet.
    Regulation of RNA Editing: The impact of inosine on the neuronal transcriptome2017Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    The transcriptome of the mammalian brain is extensively modified by adenosine to inosine (A-to-I) nucleotide conversion by two adenosine deaminases (ADAR1 and ADAR2). As adenosine and inosine have different base pairing properties, A-to-I RNA editing shapes the functional output of both coding and non-coding RNAs (ncRNAs) in the brain. The aim of this thesis was to identify editing events in small regulatory ncRNAs (miRNAs) and to determine their temporal and spatial editing status in the developing and adult mouse brain. To do this, we initially analyzed the editing status of miRNAs from different developmental time points of the mouse brain. We detected novel miRNA substrates subjected to A-to-I editing and found a general increase in miRNA editing during brain development, implicating a more stringent control of miRNAs as the brain matures. Most of the edited miRNAs were found to be transcribed as a single long consecutive transcript from a large gene cluster. However, maturation from this primary miRNA (pri-miRNA) transcript into functional forms of miRNAs is regulated individually, and might be influenced by the ADAR proteins in an editing independent matter. We also found that edited miRNAs were highly expressed at the synapse, implicating a role as local regulators of synaptic translation. We further show that the increase in editing during development is explained by a gradual accumulation of the ADAR enzymes in the nucleus. Specifically for ADAR2, we found a developmentally increasing interaction with two factors, importin-α4 and Pin1, that facilitate nuclear localization of the editing enzyme. We have also found that selectively edited stem loops often are flanked by other long stem loop structures that induce editing in cis. This may explain why multiple pri-miRNAs are edited within the same cluster. In conclusion, this thesis has significantly increased the understanding of the dynamics of both editing substrates and enzymes in the developing and mature brain.

  • 7.
    Behm, Mikaela
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Wahlstedt, Helene
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Widmark, Albin
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Eriksson, Maria
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Öhman, Marie
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Accumulation of nuclear ADAR2 regulates A-to-I RNA editing during neuronal development2017Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 130, s. 745-753Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Adenosine to inosine (A-to-I) RNA editing is important for a functional brain, and most known sites that are subject to selective RNA editing have been found to result in diversified protein isoforms that are involved in neurotransmission. In the absence of the active editing enzymes ADAR1 or ADAR2 (also known as ADAR and ADARB1, respectively), mice fail to survive until adulthood. Nuclear A-to-I editing of neuronal transcripts is regulated during brain development, with low levels of editing in the embryo and a dramatic increase after birth. Yet, little is known about the mechanisms that regulate editing during development. Here, we demonstrate lower levels of ADAR2 in the nucleus of immature neurons than in mature neurons. We show that importin-a4 (encoded by Kpna3), which increases during neuronal maturation, interacts with ADAR2 and contributes to the editing efficiency by bringing it into the nucleus. Moreover, we detect an increased number of interactions between ADAR2 and the nuclear isomerase Pin1 as neurons mature, which contribute to ADAR2 protein stability. Together, these findings explain how the nuclear editing of substrates that are important for neuronal function can increase as the brain develops. 

  • 8. Bell, Thomas J.
    et al.
    Eiríksdóttir, Emelía
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Langel, Ülo
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Eberwine, James
    PAIR technology: exon-specific RNA binding protein isolation in live cells2011Ingår i: Cell-penetrating peptides: Methods and Protocols / [ed] Ülo Langel, New York: Humana Press, 2011, s. 473-486Kapitel i bok, del av antologi (Refereegranskat)
    Abstract [en]

    RNA-binding proteins (RBPs) are fundamental regulatory proteins for all forms of transcriptional and posttranscriptional control of gene expression. However, isolating RBPs is technically challenging for investigators. Currently, the most widely used techniques to isolate RBPs are in vitro biochemical approaches. Although these approaches have been useful, they have several limitations. One key limitation to using in vitro biochemical approaches is that RBP–RNA interactions are isolated under nonbiological conditions. Here we review a novel experimental approach to identify RBPs called peptide nucleic acid (PNA)-assisted identification of RBPs (PAIR) technology (Zielinski et al., Proc Natl Acad Sci USA 103:1557–1562, 2006). This technology has two significant advantages over traditional approaches. (1) It overcomes the in vitro limitation of biochemical approaches by allowing investigators to isolate RBP–RNA interactions under in vivo conditions. (2) This technology is highly mRNA specific; it isolates RBPs in an exon-specific manner. By selectively targeting alternatively spliced exons with PAIR technology, investigators can isolate splice variant-specific and mRNA region-specific (5-UTR and 3-UTR) RBP complexes for any mRNA of interest.

  • 9.
    Bengtsson, Johanna
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    The impact of cytochrome P4501-inhibitors on aryl hydrocarbon receptor signaling2016Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    The aryl hydrocarbon receptor (AHR) best known as a ligand-activated transcription factor that mediates toxic responses to xenobiotics such as dioxins, is also activated by certain endogenous compounds. Activation of the AHR up-regulates transcription of a large number of genes, including those encoding members of the cytochrome P450 1 family of enzymes (CYP1s). Although the AHR has been shown to be involved in several normal processes, its physiological role remains elusive. The endogenous ligand 6-formylindolo[3,2-b]carbazole (FICZ), formed from tryptophan, is present in cell culture media and biological specimens. FICZ is an excellent substrate for CYP1 enzymes and together FICZ/AHR/CYP1A1 interactions constitute an auto regulatory feedback loop that controls AHR signaling. A vast number of compounds that inhibit CYP1 enzymes have been reported to be AHR activators, even though they have little or no affinity for the receptor. We hypothesized, that their agonistic effects are dependent on the presence of background levels of FICZ. To test this, AHR signaling in different cell systems exposed to FICZ and/or inhibitors was assessed by measuring EROD activity and CYP1A1 transcription. In addition to a commercial culture medium, a medium free of background levels of FICZ was used. Activation of AHR by of a diverse set of CYP1A1 inhibitors did require FICZ in the culture medium. Furthermore, the compounds tested both prolonged and potentiated FICZ-induced receptor signaling. On the basis of these observations we propose that a compound may activate AHR signaling indirectly by inhibiting CYP1A1 and thereby attenuating the metabolism of FICZ. This mechanism was confirmed for certain polyphenols and pharmaceuticals. Surprisingly, the activating capacity and potentiating effect of two pharmaceuticals on AHR signaling could not be explained by the mechanism proposed, and we speculated that in these cases the agonistic effect might involve interactions of the cellular antioxidant response with the basic transcription machinery. Together, our observations provide a mechanistic explanation as to how compounds that inhibit CYP1A1 can activate AHR signaling. They also indicate that the general perception of the binding pocket of AHR as promiscuous, is probably wrong. The fact that indirect activation of AHR may cause sustained signaling requires further studies in vivo not least, in order to prevent toxicity.

  • 10. Bergkvist, Johanna
    et al.
    Klawonn, Isabell
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för ekologi, miljö och botanik.
    Whitehouse, Martin J.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för geologiska vetenskaper. Swedish Museum of Natural History, Sweden.
    Lavik, Gaute
    Brüchert, Volker
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för geologiska vetenskaper.
    Ploug, Helle
    Turbulence simultaneously stimulates small-and large-scale CO2 sequestration by chain-forming diatoms in the sea2018Ingår i: Nature Communications, ISSN 2041-1723, E-ISSN 2041-1723, Vol. 9, artikel-id 3046Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Chain-forming diatoms are key CO2-fixing organisms in the ocean. Under turbulent conditions they form fast-sinking aggregates that are exported from the upper sunlit ocean to the ocean interior. A decade-old paradigm states that primary production in chain-forming diatoms is stimulated by turbulence. Yet, direct measurements of cell-specific primary production in individual field populations of chain-forming diatoms are poorly documented. Here we measured cell-specific carbon, nitrate and ammonium assimilation in two field populations of chain-forming diatoms (Skeletonema and Chaetoceros) at low-nutrient concentrations under still conditions and turbulent shear using secondary ion mass spectrometry combined with stable isotopic tracers and compared our data with those predicted by mass transfer theory. Turbulent shear significantly increases cell-specific C assimilation compared to still conditions in the cells/chains that also form fast-sinking, aggregates rich in carbon and ammonium. Thus, turbulence simultaneously stimulates small-scale biological CO2 assimilation and large-scale biogeochemical C and N cycles in the ocean.

  • 11.
    Bonath, Franziska
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut. Stockholms universitet, Science for Life Laboratory (SciLifeLab).
    Domingo-Prim, Judit
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Tarbier, Marcel
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut. Stockholms universitet, Science for Life Laboratory (SciLifeLab).
    Friedländer, Marc
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut. Stockholms universitet, Science for Life Laboratory (SciLifeLab).
    Visa, Neus
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Next-generation sequencing reveals two populations of damage- induced small RNAs at endogenous DNA double-strand breaksIngår i: Nucleic Acids Research, ISSN 0305-1048, E-ISSN 1362-4962Artikel i tidskrift (Refereegranskat)
  • 12. Brunetti, Dario
    et al.
    Torsvik, Janniche
    Dallabona, Cristina
    Teixeira, Pedro
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Sztromwasser, Pawel
    Fernandez-Vizarra, Erika
    Cerutti, Raffaele
    Reyes, Aurelio
    Preziuso, Carmela
    D'Amati, Giulia
    Baruffini, Enrico
    Goffrini, Paola
    Viscomi, Carlo
    Ferrero, Ileana
    Boman, Helge
    Telstad, Wenche
    Johansson, Stefan
    Glaser, Elzbieta
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Knappskog, Per M.
    Zeviani, Massimo
    Bindoff, Laurence A.
    Defective PITRM1 mitochondrial peptidase is associated with A amyloidotic neurodegeneration2016Ingår i: EMBO Molecular Medicine, ISSN 1757-4676, E-ISSN 1757-4684, Vol. 8, nr 3, s. 176-190Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Mitochondrial dysfunction and altered proteostasis are central features of neurodegenerative diseases. The pitrilysin metallopeptidase 1 (PITRM1) is a mitochondrial matrix enzyme, which digests oligopeptides, including the mitochondrial targeting sequences that are cleaved from proteins imported across the inner mitochondrial membrane and the mitochondrial fraction of amyloid beta (A). We identified two siblings carrying a homozygous PITRM1 missense mutation (c.548G>A, p.Arg183Gln) associated with an autosomal recessive, slowly progressive syndrome characterised by mental retardation, spinocerebellar ataxia, cognitive decline and psychosis. The pathogenicity of the mutation was tested invitro, in mutant fibroblasts and skeletal muscle, and in a yeast model. A Pitrm1(+/-) heterozygous mouse showed progressive ataxia associated with brain degenerative lesions, including accumulation of A-positive amyloid deposits. Our results show that PITRM1 is responsible for significant A degradation and that impairment of its activity results in A accumulation, thus providing a mechanistic demonstration of the mitochondrial involvement in amyloidotic neurodegeneration.

  • 13. Busayavalasa, Kiran
    et al.
    Chen, Xin
    Östlund Farrants, Ann-Kristin
    Stockholms universitet, Naturvetenskapliga fakulteten, Wenner-Grens institut, Avdelningen för cellbiologi.
    Wagner, Nicole
    Sabri, Nafiseh
    The nup155 mediated organisation of inner nuclear membrane proteins is independent of nup155 anchoring to the metazoan nuclear pore complex2012Ingår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 125, nr 18, s. 4214-4218Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The nuclear envelope (NE), an important barrier between the nucleus and the cytoplasm, is composed of three structures: the outer nuclear membrane, which is continuous with the ER, the inner nuclear membrane (INM), which interfaces with chromatin, and nuclear pore complexes (NPCs), which are essential for the exchange of macromolecules between the two compartments. The NPC protein Nup155 has an evolutionarily conserved role in the metazoan NE formation; but the in vivo analysis of Nup155 has been severely hampered by the essential function of this protein in cell viability. Here, we take advantage of the hypomorphicity of RNAi systems and use a combination of protein binding and rescue assays to map the interaction sites of two neighbouring NPC proteins Nup93 and Nup53 on Nup155, and to define the requirements of these interactions in INM protein organization. We show that different parts of Drosophila Nup155 have distinct functions: the Nup155 beta-propeller anchors the protein to the NPC, whereas the alpha-solenoid part of Nup155 is essential for the correct localisation of INM proteins lamin-B receptor (LBR) and otefin. Using chromatin extracts from semisynchronized cells, we also provide evidence that the Nup155 alpha-solenoid has a chromatin-binding activity that is stronger at the end of mitosis. Our results argue that the role of Nup155 in INM protein localisation is not mediated through the NPC anchoring activity of the protein and suggest that regions other than Nup155 beta-propeller are necessary for the targeting of proteins to the INM.

  • 14.
    Cannon, Barbara
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Nedergaard, Jan
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    What Ignites UCP1?2017Ingår i: Cell Metabolism, ISSN 1550-4131, E-ISSN 1932-7420, Vol. 26, nr 5, s. 697-698Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    We thought we knew how the heat-producing uncoupling protein 1 in brown adipose tissue was activated: by fatty acids released upon lipid droplet breakdown in the brown adipocytes. However, two studies in this issue (Schreiber et al., 2017; Shin et al., 2017) imply that this classical model may not be valid: heat can be produced in brown fat without intracellular lipolysis.

  • 15. Cedernaes, Jonathan
    et al.
    Schonke, Milena
    Orzechowski Westholm, Jakub
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik. Stockholms universitet, Science for Life Laboratory (SciLifeLab).
    Mi, Jia
    Chibalin, Alexander
    Voisin, Sarah
    Osler, Megan
    Vogel, Heike
    Hornaeus, Katarina
    Dickson, Suzanne L.
    Lind, Sara Bergstrom
    Bergquist, Jonas
    Schioth, Helgi B.
    Zierath, Juleen R.
    Benedict, Christian
    Acute sleep loss results in tissue-specific alterations in genome-wide DNA methylation state and metabolic fuel utilization in humans2018Ingår i: Science Advances, ISSN 0036-8156, E-ISSN 2375-2548, Vol. 4, nr 8, artikel-id eaar8590Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Curtailed sleep promotes weight gain and loss of lean mass in humans, although the underlying molecular mechanisms are poorly understood. We investigated the genomic and physiological impact of acute sleep loss in peripheral tissues by obtaining adipose tissue and skeletal muscle after one night of sleep loss and after one full night of sleep. We find that acute sleep loss alters genome-wide DNA methylation in adipose tissue, and unbiased transcriptome-, protein-, and metabolite-level analyses also reveal highly tissue-specific changes that are partially reflected by altered metabolite levels in blood. We observe transcriptomic signatures of inflammation in both tissues following acute sleep loss, but changes involving the circadian clock are evident only in skeletal muscle, and we uncover molecular signatures suggestive of muscle breakdown that contrast with an anabolic adipose tissue signature. Our findings provide insight into how disruption of sleep and circadian rhythms may promote weight gain and sarcopenia.

  • 16. Chiurchiu, Valerio
    et al.
    Leuti, Alessandro
    Dalli, Jesmond
    Jacobsson, Anders
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Battistini, Luca
    Maccarrone, Mauro
    Serhan, Charles N.
    Proresolving lipid mediators resolvin D1, resolvin D2, and maresin 1 are critical in modulating T cell responses2016Ingår i: Science Translational Medicine, ISSN 1946-6234, E-ISSN 1946-6242, Vol. 8, nr 353, artikel-id 353ra111Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Resolution of inflammation is a finely regulated process mediated by specialized proresolving lipid mediators (SPMs), including docosahexaenoic acid (DHA)-derived resolvins and maresins. The immunomodulatory role of SPMs in adaptive immune cells is of interest. We report that D-series resolvins (resolvin D1 and resolvin D2) and maresin 1 modulate adaptive immune responses in human peripheral blood lymphocytes. These lipid mediators reduce cytokine production by activated CD8(+) T cells and CD4(+) T helper 1 (T(H)1) and T(H)17 cells but do not modulate T cell inhibitory receptors or abrogate their capacity to proliferate. Moreover, these SPMs prevented naive CD4(+) T cell differentiation into T(H)1 and T(H)17 by down-regulating their signature transcription factors, T-bet and Rorc, in a mechanism mediated by the GPR32 and ALX/FPR2 receptors; they concomitantly enhanced de novo generation and function of Foxp3(+) regulatory T (T-reg) cells via the GPR32 receptor. These results were also supported in vivo in a mouse deficient for DHA synthesis (Elovl2(-/-)) that showed an increase in T(H)1/T(H)17 cells and a decrease in T-reg cells compared to wild-type mice. Additionally, either DHA supplementation in Elovl2(-/-) mice or in vivo administration of resolvin D1 significantly reduced cytokine production upon specific stimulation of T cells. These findings demonstrate actions of specific SPMs on adaptive immunity and provide a new avenue for SPM-based approaches to modulate chronic inflammation.

  • 17. Coenen-Stass, Anna M. L.
    et al.
    Sork, Helena
    Gatto, Sole
    Godfrey, Caroline
    Bhomra, Amarjit
    Krjutskov, Kaarel
    Hart, Jonathan R.
    Westholm, Jakub O.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik. Stockholms universitet, Science for Life Laboratory (SciLifeLab).
    O'Donovan, Liz
    Roos, Andreas
    Lochmueller, Hanns
    Puri, Pier Lorenzo
    EL Andaloussi, Samir
    Wood, Matthew J. A.
    Roberts, Thomas C.
    Comprehensive RNA-Sequencing Analysis in Serum and Muscle Reveals Novel Small RNA Signatures with Biomarker Potential for DMD2018Ingår i: Molecular Therapy - Nucleic Acids, ISSN 2162-2531, E-ISSN 2162-2531, Vol. 13, s. 1-15Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Extracellular small RNAs (sRNAs), including microRNAs (miRNAs), are promising biomarkers for diseases such as Duchenne muscular dystrophy (DMD), although their biological relevance is largely unknown. To investigate the relationship between intracellular and extracellular sRNA levels on a global scale, we performed sRNA sequencing in four muscle types and serum from wild-type, dystrophic mdx, and mdx mice in which dystrophin protein expression was restored by exon skipping. Differentially abundant sRNAs were identified in serum (mapping to miRNA, small nuclear RNA [snRNA], and PIWI-interacting RNA [piRNA] loci). One novel candidate biomarker, miR-483, was increased in both mdx serum and muscle, and also elevated in DMD patient sera. Dystrophin restoration induced global shifts in miRNA (including miR-483) and snRNA-fragment abundance toward wild-type levels. Specific serum piRNA-like sRNAs also responded to exon skipping therapy. Absolute miRNA expression in muscle was positively correlated with abundance in the circulation, although multiple highly expressed miRNAs in muscle were not elevated in mdx serum, suggesting that both passive and selective release mechanisms contribute to serum miRNA levels. In conclusion, this study has revealed new insights into the sRNA biology of dystrophin deficiency and identified novel DMD biomarkers.

  • 18. Darai-Ramqvist, Eva
    et al.
    Nilsonne, Gustav
    Stockholms universitet, Samhällsvetenskapliga fakulteten, Stressforskningsinstitutet. Karolinska Institutet, Sweden.
    Flores-Staino, Carmen
    Hjerpe, Anders
    Dobra, Katalin
    Microenvironment-dependent phenotypic changes in a SCID mouse model for malignant mesothelioma2013Ingår i: Frontiers in Oncology, ISSN 2234-943X, E-ISSN 2234-943X, Vol. 3, artikel-id 203Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Background and Aims: Malignant mesothelioma is an aggressive, therapy-resistant tumor. Mesothelioma cells may assume an epithelioid or a sarcomatoid phenotype, and presence of sarcomatoid cells predicts poor prognosis. In this study, we investigated differentiation of mesothelioma cells in a xenograft model, where mesothelioma cells of both phenotypes were induced to form tumors in severe combined immunodeficiency mice.

    Methods: Xenografts were established and thoroughly characterized using a comprehensive immunohistochemical panel, array comparative genomic hybridization (aCGH) of chromosome 3, fluorescent in situ hybridization, and electron microscopy.

    Results: Epithelioid and sarcomatoid cells gave rise to xenografts of similar epithelioid morphology. While sarcomatoid-derived xenografts had higher growth rates, the morphology and expression of differentiation-related markers was similar between xenografts derived from both phenotypes. aCGH showed a convergent genotype for both xenografts, resembling the original aggressive sarcomatoid cell sub-line.

    Conclusion: Human mesothelioma xenografts from sarcomatoid and epithelioid phenotypes converged to a similar differentiation state, and genetic analyses suggested that clonal selection in the mouse microenvironment was a major contributing factor. This thoroughly characterized animal model can be used for further studies of molecular events underlying tumor cell differentiation.

  • 19.
    Domingo Prim, Judit
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    The RNA exosome and the maintenance of genome integrity2018Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    The RNA exosome is a ribonucleolytic complex that acts on different RNA substrates and plays important roles in RNA metabolism. In recent years, the synthesis and the processing of RNA have been directly linked to the integrity of the genome. RNAs can either be the responsible for genomic instability or, on the contrary, can participate in the DNA damage response. Damage-induced RNAs (diRNAs) are short non-coding RNAs that have been implicated in the repair of DNA double-strand breaks (DSBs) by homologous recombination. The implication of specialized RNAs in DNA damage and repair led us to investigate whether the exosome was involved in DNA repair.

    In Paper I, we have shown by fluorescence microscopy and chromatin immunoprecipitation that the exosome catalytic subunit RRP6/EXOSC10 is recruited to DSBs in Drosophila and human cells. Depletion of this subunit or overexpression of a catalytically inactive mutant makes the cells more sensitive to radiation and unable to recruit the homologous recombination factor RAD51 to DSBs, which is consistent with RRP6/EXOSC10 playing a role in homologous recombination, both in insect and mammalian cells. The results obtained with the RRP6 inactive mutant also suggest that the ribonucleolytic activity of RRP6 is required for DNA repair. However, the mechanisms by which RNAs and the exosome are implicated in DNA repair need to be further investigated.

    In Paper II, we describe how transcription of DSB-flanking sequences by RNA polymerase II gives rise to damage-induced long non-coding RNAs that are processed into diRNAs. The direct detection of diRNAs had been elusive and their existence had been questioned, but our results show that damage-induced transcription and diRNA production occur at DSBs in endogenous, repetitive genomic sequences in mammalian cells. However, our exhaustive next-generation sequencing failed to detect diRNAs derived from DSBs in unique sequences. The diRNAs produced at repetitive loci bind to Argonaute and belong to two different subpopulations. One of them is Dicer-dependent and has a length of 21-22 nucleotides. The other one is not yet well characterized and is probably composed of degradation products from other ribonucleases.

    Finally, in Paper III, we have demonstrated that EXOSC10 is one of the ribonucleases involved in RNA degradation at DSBs. By strand-specific quantitative PCR and RNA-seq, we show that the levels of diRNA precursors and diRNAs are increased in the absence of EXOSC10. Moreover, EXOSC10-depleted cells fail to recruit RPA to DSBs, and this defect is restored by RNase A digestion. Depletion of EXOSC10 also results in extended DNA resected tracks, as shown by both single-molecule analysis of resected tracks and quantitative amplification of single-stranded DNA. These results suggest that EXOSC10 is involved in RNA degradation at DSBs to allow RPA recruitment and regulated resection.

    The work presented in this thesis supports the conclusion that damage-induced RNAs are synthesized de novo by RNA polymerase II at DSBs in mammalian cells. In repetitive genomic loci, these RNAs are processed into diRNAs that bind Argonaute. Regardless of whether diRNAs are functional or not, their precursors have to be degraded. The main function of the exosome, and more specifically EXOSC10, in the maintenance of the integrity of the genome is to degrade these transcripts in order to allow faithful repair of DNA double-strand breaks by homologous recombination.

  • 20.
    Domingo-Prim, Judit
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Endara-Coll, Martín
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Bonath, Franziska
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut. Stockholms universitet, Science for Life Laboratory (SciLifeLab).
    Jimeno, Sonia
    Friedländer, Marc
    Stockholms universitet, Science for Life Laboratory (SciLifeLab). Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Huertas, Pablo
    Visa, Neus
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    EXOSC10 is required for RPA assembly and controlled DNA resection at DNA dobule-strand breaksManuskript (preprint) (Övrigt vetenskapligt)
  • 21.
    Ekberg, Monica
    Stockholms universitet.
    Radical transfer in ribonucleotide reductase from Escherichia coli2000Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
  • 22.
    EL Andaloussi, Samir
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Lehto, Taavi
    Laboratory of Molecular Biotechnology, Institute of Technology, Tartu University, Tartu, Estonia.
    Lundin, Per
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Langel, Ülo
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Application of PepFect peptides for the delivery of splice-correcting oligonucleotides2011Ingår i: Cell-penetrating peptides: Methods and Protocols, New York: Humana Press, 2011, s. 361-373Kapitel i bok, del av antologi (Övrigt vetenskapligt)
    Abstract [en]

    One oligonucleotide-based approach that appear very promising for the treatment of different genetic disorders are based on so-called splice-correcting oligonucleotides (SCOs) that are exploited to manipulate splicing patterns. In order to increase the bioavailability, cell-penetrating peptides (CPPs) have readily been covalently conjugated to SCOs to facilitate cellular internalization. While being a successful strategy for the delivery of uncharged oligonucleotides (ONs), it is extremely difficult to generate covalent conjugates between commonly used negatively charged ON analogs and cationic CPPs. Furthermore, high concentrations of ONs in the micromolar range are often needed to obtain biological responses, most likely as a result of endosomal entrapment of material. Therefore, exploring other vectorization methods using CPPs with endosomolytic properties are highly desired. A method of using stearyl modified CPP (i.e., TP10) analogs, named PepFect3 and PepFect4, are being described for the transfection of antisense SCOs using a simple one-step co-incubation procedure. These peptides form complexes with SCOs and efficiently promote cellular uptake by facilitating endosomal escape. This chapter describes the methods of how to form and characterize these nanoparticles and the cellular assay used to address the delivery.

  • 23.
    EL Andaloussi, Samir
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Said Hassane, Fatouma
    Université Montpellier, Motpellier, France.
    Boisguerin, Prisca
    Université Montpellier, Motpellier, France.
    Sillard, Rannar
    University of Tartu, Institute of Technology, Tartu, Estonia.
    Langel, Ülo
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Lebleu, Bernard
    Université Montpellier, Motpellier, France.
    Cell-penetrating peptides-based strategies for the delivery of splice redirecting antisense oligonucleotides2011Ingår i: Therapeutic Oligonucleotides: Methods and Protocols / [ed] John Goodchild, New York: Humana Press, 2011, s. 75-89Kapitel i bok, del av antologi (Övrigt vetenskapligt)
    Abstract [en]

    Progress in our understanding of the molecular pathogenesis of human malignancies has provided therapeutic targets amenable to oligonucleotide (ON)-based strategies. Antisense ON-mediated splicing regulation in particular offers promising prospects since the majority of human genes undergo alternative splicing and since splicing defects have been found in many diseases. However, their implementation has been hampered so far by the poor bioavailability of nucleic acids-based drugs. Cell-penetrating peptides (CPPs) now appear as promising non-viral delivery vector for non-permeant biomolecules. We describe here new CPPs allowing the delivery of splice redirecting steric-block ON using either chemical conjugation or non-covalent complexation. We also describe a convenient and robust splice redirecting assay which allows the quantitative assessment of ON nuclear delivery.

  • 24. Elliott, Mark
    et al.
    Favre-Guilmard, Christine
    Liu, Sai Man
    Maignel, Jacquie
    Masuyer, Geoffrey
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Beard, Matthew
    Boone, Christopher
    Carre, Denis
    Kalinichev, Mikhail
    Lezmi, Stephane
    Mir, Imran
    Nicoleau, Camille
    Palan, Shilpa
    Perier, Cindy
    Raban, Elsa
    Zhang, Sicai
    Dong, Min
    Stenmark, Pål
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik. Lund University, Sweden.
    Krupp, Johannes
    Engineered botulinum neurotoxin B with improved binding to human receptors has enhanced efficacy in preclinical models2019Ingår i: Science Advances, E-ISSN 2375-2548, Vol. 5, nr 1, artikel-id eaau7196Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Although botulinum neurotoxin serotype A (BoNT/A) products are common treatments for various disorders, there is only one commercial BoNT/B product, whose low potency, likely stemming from low affinity toward its human receptor synaptotagmin 2 (hSyt2), has limited its therapeutic usefulness. We express and characterize two full-length recombinant BoNT/B1 proteins containing designed mutations E1191M/S1199Y (rBoNT/B1(MY)) and E1191Q/S1199W (rBoNT/B1(QW)) that enhance binding to hSyt2. In preclinical models including human-induced pluripotent stem cell neurons and a humanized transgenic mouse, this increased hSyt2 affinity results in high potency, comparable to that of BoNT/A. Last, we solve the cocrystal structure of rBoNT/B1(MY) in complex with peptides of hSyt2 and its homolog hSyt1. We demonstrate that neuronal surface receptor binding limits the clinical efficacy of unmodified BoNT/B and that modified BoNT/B proteins have promising clinical potential.

  • 25.
    Fischer, Alexander W.
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut. University Medical Center Hamburg-Eppendorf, Germany.
    Shabalina, Irina G.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Mattsson, Charlotte L.
    Abreu-Vieira, Gustavo
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Cannon, Barbara
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Nedergaard, Jan
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Petrovic, Natasa
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    UCP1 inhibition in Cidea-overexpressing mice is physiologically counteracted by brown adipose tissue hyperrecruitment2017Ingår i: American Journal of Physiology. Endocrinology and Metabolism, ISSN 0193-1849, E-ISSN 1522-1555, Vol. 312, nr 1, s. e72-E87Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Cidea is a gene highly expressed in thermogenesis- competent (UCP1-containing) adipose cells, both brown and brite/beige. Here, we initially demonstrate a remarkable adipose-depot specific regulation of Cidea expression. In classical brown fat, Cidea mRNA is expressed continuously and invariably, irrespective of tissue recruitment. However, Cidea protein levels are regulated posttranscriptionally, being conspicuously induced in the thermogenically recruited state. In contrast, in brite fat, Cidea protein levels are regulated at the transcriptional level, and Cidea mRNA and protein levels are proportional to tissue briteness. Although routinely followed as a thermogenic molecular marker, Cidea function is not clarified. Here, we employed a gain-of-function approach to examine a possible role of Cidea in the regulation of thermogenesis. We utilized transgenic aP2-hCidea mice that overexpress human Cidea in all adipose tissues. We demonstrate that UCP1 activity is markedly suppressed in brown-fat mitochondria isolated from aP2-hCidea mice. However, mitochondrial UCP1 protein levels were identical in wildtype and transgenic mice. This implies a regulatory effect of Cidea on UCP1 activity, but as we demonstrate that Cidea itself is not localized to mitochondria, we propose an indirect inhibitory effect. The Cidea-induced inhibition of UCP1 activity (observed in isolated mitochondria) is physiologically relevant since the mice, through an appropriate homeostatic compensatory mechanism, increased the total amount of UCP1 in the tissue to exactly match the diminished thermogenic capacity of the UCP1 protein and retain unaltered nonshivering thermogenic capacity. Thus, we verified Cidea as being a marker of thermogenesis-competent adipose tissues, but we conclude that Cidea, unexpectedly, functions molecularly as an indirect inhibitor of thermogenesis.

  • 26.
    Gaudry, Michael J.
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Jastroch, Martin
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Molecular evolution of uncoupling proteins and implications for brain function2019Ingår i: Neuroscience Letters, ISSN 0304-3940, E-ISSN 1872-7972, Vol. 696, s. 140-145Artikel, forskningsöversikt (Refereegranskat)
    Abstract [en]

    Uncoupling proteins (UCPs) belong to the mitochondrial anion carrier superfamily and catalyze important metabolic functions at the mitochondrial inner membrane. While the thermogenic role of UCP1 in brown fat of eutherian mammals is well established, the molecular functions of UCP1 in ectothermic vertebrates and of other UCP paralogs remain less clear. Here, we critically discuss the existence of brain UCPs and their potential roles. Applying phylogenetic classification of novel UCPs, we summarize the evidence for brain UCP1 among vertebrates, the role of UCP2 in specific brain areas, and the existence of brain-specific UCPs. The phylogenetic analyses and discussion on functional data should alert the scientific community that the molecular function of so-called UCP1 homologues is by far not clarified and possibly relates to neither thermogenesis nor mitochondrial uncoupling.

  • 27.
    Gañez Zapater, Antoni
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Gene regulation by chromatin remodelling complexes: SWI/SNF complex in mRNA processing and B-WICH complex in ribosomal gene expression2018Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    The aim of this project is to investigate the roles of chromatin remodelling complexes in gene regulation. It is focused on two groups of chromatin complexes: the mammalian BRG1 and BRM SWI/SNF complexes and the ISWI-containing B-WICH complex.

    Study 1 investigates the role of SWI/SNF complexes in alternative splicing. We show that the presence of the ATPase core subunits Brg1 and Brm influence the alternative splicing outcome of a subset of genes. We show that Brg1 and Brm interact with several splicing related factors in the nascent RNA, and that the recruitment of some of these factors to their target sites is regulated by the presence of Brg1 and Brm. We propose that SWI/SNF ATPases can modulate the interactions of RNA binding factors to the nascent RNA and in that way alter alternative splicing outcome.

    Study 2 focuses on SWI/SNF complexes and their influence on cleavage and polyadenylation of mRNA. We show that Brg1 and Brm interact with subunits of the cleavage and polyadenylation complexes in the nascent mRNA. SWI/SNF complexes facilitate the recruitment of the cleavage and polyadenylation complex to the polyadenylation site in a subset of genes, and this results in a more efficient cleavage and polyadenylation.

    Study 3 shows that B-WICH is required for ribosome gene transcriptional activation upon glucose stimulation. WSTF and SNF2h, two of the B-WICH subunits, are needed to establish an active chromatin state in the RNA pol I gene promoter when the glucose concentration is raised after a period of deprivation. We propose that it counteracts the silent, poised chromatin state imposed by the silencing chromatin remodelling complex NuRD to allow for the RNA pol I machinery to bind to the promoter.

    These studies show that the influence of chromatin remodelling complexes upon gene expression is important for remodelling nucleosomes at the promoter, for alternative splicing, cleavage and polyadenylation and transcriptional initiation. These complexes work together with other chromatin remodelling factors, interact with other complexes and regulate their activity by affecting their recruitment dynamics.

  • 28.
    Gañez Zapater, Antoni
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Mackowiak, Sebastian
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Guo, Yuan
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Jordán-Pla, Antonio
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Visa, Neus
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Östlund-Farrants, Ann-Kristin
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    SWI/SNF subunits Brg1 and Brm regulate alternative splicing by interacting with RNA binding proteins in the nascent RNA.Manuskript (preprint) (Övrigt vetenskapligt)
  • 29. Gałecki, Maciej
    et al.
    Tartas, Adrianna
    Szymanek, Agata
    Sims, Emma
    Lundholm, Lovisa
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Sollazzo, Alice
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Cheng, Lei
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Fujishima, Yohei
    Yoshida, Mitsuaki A.
    Żygierewicz, Jarosław
    Wojcik, Andrzej
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut. Jan Kochanowski University, Poland.
    Brzozowska-Wardecka, Beata
    Precision of scoring radiation-induced chromosomal aberrations and micronuclei by unexperienced scorers2019Ingår i: International Journal of Radiation Biology, ISSN 0955-3002, E-ISSN 1362-3095, Vol. 95, nr 9, s. 1251-1258Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Purpose: Dose assessment plays an important role in case of radiological accidents and can be performed by scoring structural changes of chromosome morphology induced in cells by ionizing radiation. The results of such a test are biased by scorer experience, therefore, simple to learn assays are recommended to be used when fast analysis of a large amount of data is needed. The aim of this study was to compare the performance of two radiobiological assays - chromosomal aberrations and micronuclei - by unexperienced scorers with the reference values generated by an expert.

    Materials and methods: Each participant of an EU-funded two-week radiobiology course was asked to score Chinese hamster ovary cells exposed to gamma radiation up to 4 Gy. The congruence of students' and expert's scores at each dose and the coherence of the dose-response curve parameters between the students were investigated.

    Results: Micronucleus test tended to be faster and easier to learn than scoring chromosomal aberrations. However, both assays carried out by inexperienced students showed reasonable dose-response curves.

    Conclusions: In the case of a large radiological accident involving many casualties, the unexperienced scorers would support the process of biodosimetric triage by cytogenetic biological dosimetry.

  • 30.
    Geörg, Miriam
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Maudsdotter, Lisa
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Tavares, Raquel
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Jonsson, Ann-Beth
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Meningococcal resistance to antimicrobial peptides is mediated by bacterial adhesion and host cell RhoA and Cdc42 signalling2013Ingår i: Cellular Microbiology, ISSN 1462-5814, E-ISSN 1462-5822, Vol. 15, nr 11, s. 1938-1954Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Antimicrobial peptides (AMPs) constitute an essential part of the innate immune defence. Pathogenic bacteria have evolved numerous strategies to withstand AMP-mediated killing. The influence of host epithelia on bacterial AMP resistance is, however, still largely unknown. We found that adhesion to pharyngeal epithelial cells protected Neisseria meningitidis, a leading cause of meningitis and sepsis, from the human cathelicidin LL-37, the cationic model amphipathic peptide (MAP) and the peptaibol alamethicin, but not from polymyxin B. Adhesion to primary airway epithelia resulted in a similar increase in LL-37 resistance. The inhibition of selective host cell signalling mediated by RhoA and Cdc42 was found to abolish the adhesion-induced LL-37 resistance by a mechanism unrelated to the actin cytoskeleton. Moreover, N. meningitidis triggered the formation of cholesterol-rich membrane microdomains in pharyngeal epithelial cells, and host cell cholesterol proved to be essential for adhesion-induced resistance. Our data highlight the importance of Rho GTPase-dependent host cell signalling for meningococcal AMP resistance. These results indicate that N. meningitidis selectively exploits the epithelial microenvironment in order to protect itself from LL-37.

  • 31. Gottipati, Ponnari
    et al.
    Cassel, Tobias N.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för genetik, mikrobiologi och toxikologi.
    Savolainen, Linda
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för genetik, mikrobiologi och toxikologi.
    Helleday, Thomas
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för genetik, mikrobiologi och toxikologi.
    Transcription-associated recombination is dependent on replication in Mammalian cells2008Ingår i: Molecular and Cellular Biology, ISSN 0270-7306, E-ISSN 1098-5549, Vol. 28, nr 1, s. 154-64Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Transcription can enhance recombination; this is a ubiquitous phenomenon from prokaryotes to higher eukaryotes. However, the mechanism of transcription-associated recombination in mammalian cells is poorly understood. Here we have developed a construct with a recombination substrate in which levels of recombination can be studied in the presence or absence of transcription. We observed a direct enhancement in recombination when transcription levels through the substrate were increased. This increase in homologous recombination following transcription is locus specific, since homologous recombination at the unrelated hprt gene is unaffected. In addition, we have shown that transcription-associated recombination involves both short-tract and long-tract gene conversions in mammalian cells, which are different from double-strand-break-induced recombination events caused by endonucleases. Transcription fails to enhance recombination in cells that are not in the S phase of the cell cycle. Furthermore, inhibition of transcription suppresses induction of recombination at stalled replication forks, suggesting that recombination may be involved in bypassing transcription during replication.

  • 32.
    Gustafsson, Helena
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Tamm, Christoffer
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Forsby, Anna
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Signalling pathways for insulin-like growth factor type 1-mediated expression of uncoupling protein 3.2004Ingår i: J Neurochem, ISSN 0022-3042, Vol. 88, nr 2, s. 462-8Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Uncoupling protein 3 (UCP3) is a mitochondrial protein with antioxidant properties and its regulation by factors promoting cell-survival may be important for protection of, for instance, neurons in states of oxidative stress. In the present study, we investigated regulatory pathways for UCP3 expression mediated by the neuroprotective hormone insulin-like growth factor type 1 (IGF-1) in human neuroblastoma SH-SY5Y cells. Northern blot analysis and RT-PCR showed that treatment with 10 nm IGF-1 increased the UCP3 mRNA levels 2.5-fold after 5 h. Co-incubation with the phosphatidylinositol 3 (PI3)-kinase inhibitor LY294002 prohibited IGF-1-mediated induction of both UCP3 mRNA and protein in a concentration-dependent manner, with a complete blockage at 1 microm, as shown by RT-PCR and western blot analyses. The mitogen-activated protein (MAP) kinase kinase 1 (MKK1 or MEK) inhibitor PD98059 also decreased the UCP3 mRNA expression at 10 microm, however, this concentration only partly inhibited the protein expression. We conclude that IGF-1 enhanced UCP3 expression at transcriptional level, primarily through the PI3-kinase-dependent pathway and partly through the MAP kinase pathway.

  • 33. Gustafsson, Nina M. S.
    et al.
    Färnegårdh, Katarina
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för organisk kemi. Stockholms universitet, Science for Life Laboratory (SciLifeLab). Kancera AB, Sweden.
    Bonagas, Nadilly
    Ninou, Anna Huguet
    Groth, Petra
    Wiita, Elisee
    Jönsson, Mattias
    Hallberg, Kenth
    Lehto, Jemina
    Pennisi, Rosa
    Martinsson, Jessica
    Norström, Carina
    Hollers, Jessica
    Schultz, Johan
    Andersson, Martin
    Markova, Natalia
    Marttila, Petra
    Kim, Baek
    Norin, Martin
    Olin, Thomas
    Helleday, Thomas
    Targeting PFKFB3 radiosensitizes cancer cells and suppresses homologous recombination2018Ingår i: Nature Communications, ISSN 2041-1723, E-ISSN 2041-1723, Vol. 9, artikel-id 3872Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The glycolytic PFKFB3 enzyme is widely overexpressed in cancer cells and an emerging anticancer target. Here, we identify PFKFB3 as a critical factor in homologous recombination (HR) repair of DNA double-strand breaks. PFKFB3 rapidly relocates into ionizing radiation (IR)-induced nuclear foci in an MRN-ATM-gamma H2AX-MDC1-dependent manner and co-localizes with DNA damage and HR repair proteins. PFKFB3 relocalization is critical for recruitment of HR proteins, HR activity, and cell survival upon IR. We develop KAN0438757, a small molecule inhibitor that potently targets PFKFB3. Pharmacological PFKFB3 inhibition impairs recruitment of ribonucleotide reductase M2 and deoxynucleotide incorporation upon DNA repair, and reduces dNTP levels. Importantly, KAN0438757 induces radiosensitization in transformed cells while leaving non-transformed cells unaffected. In summary, we identify a key role for PFKFB3 enzymatic activity in HR repair and present KAN0438757, a selective PFKFB3 inhibitor that could potentially be used as a strategy for the treatment of cancer.

  • 34.
    Gustafsson, Robert
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Zhang, Sicai
    Masuyer, Geoffrey
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Dong, Min
    Stenmark, Pål
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Crystal Structure of Botulinum Neurotoxin A2 in Complex with the Human Protein Receptor SV2C Reveals Plasticity in Receptor Binding2018Ingår i: Toxins, ISSN 2072-6651, E-ISSN 2072-6651, Vol. 10, nr 4, artikel-id 153Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Botulinum neurotoxins (BoNTs) are a family of highly dangerous bacterial toxins, with seven major serotypes (BoNT/A-G). Members of BoNTs, BoNT/A1 and BoNT/B1, have been utilized to treat an increasing number of medical conditions. The clinical trials are ongoing for BoNT/A2, another subtype of BoNT/A, which showed promising therapeutic properties. Both BoNT/A1 and BoNT/A2 utilize three isoforms of synaptic vesicle protein SV2 (SV2A, B, and C) as their protein receptors. We here present a high resolution (2.0 angstrom) co-crystal structure of the BoNT/A2 receptor-binding domain in complex with the human SV2C luminal domain. The structure is similar to previously reported BoNT/A-SV2C complexes, but a shift of the receptor-binding segment in BoNT/A2 rotates SV2C in two dimensions giving insight into the dynamic behavior of the interaction. Small differences in key residues at the binding interface may influence the binding to different SV2 isoforms, which may contribute to the differences between BoNT/A1 and BoNT/A2 observed in the clinic.

  • 35.
    Guterstam, Peter
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    EL Andaloussi, Samir
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Langel, Ülo
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Characterization of cellular internalization pathways for CPP-mediated oligonucleotide delivery2011Ingår i: Cell-penetrating peptides: Methods and Protocols / [ed] Ülo Langel, New York: Humana Press, 2011, s. 219-230Kapitel i bok, del av antologi (Övrigt vetenskapligt)
    Abstract [en]

    The methods for evaluating internalization pathways of cellular CPP-mediated ON delivery utilizing a pre-mRNA splice correction assay and fluorescence-based quantification are described. Examples for characterization of CPP uptake routes, employing various endocytosis inhibitors, and special treatment conditions are demonstrated. The methods are developed to characterize cellular delivery of pre-mRNA splice switching peptide nucleic acids conjugated to CPPs by disulfide bond.

  • 36. Göranson, Sofie Paues
    et al.
    Thålin, Charlotte
    Lundström, Annika
    Hållström, Lars
    Lasselin, Julie
    Stockholms universitet, Samhällsvetenskapliga fakulteten, Stressforskningsinstitutet. Karolinska Institutet, Sweden.
    Wallén, Håkan
    Soop, Anne
    Mobarrez, Fariborz
    Circulating H3Cit is elevated in a human model of endotoxemia and can be detected bound to microvesicles2018Ingår i: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 8, artikel-id 12641Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Early diagnosis of sepsis is crucial since prompt interventions decrease mortality. Citrullinated histone H3 (H3Cit), released from neutrophil extracellular traps (NETs) upon binding of platelets to neutrophils following endotoxin stimulation, has recently been proposed a promising blood biomarker in sepsis. Moreover, microvesicles (MVs), which are released during cell activation and apoptosis and carry a variety of proteins from their parental cells, have also been shown to be elevated in sepsis. In a randomized and placebo-controlled human model of endotoxemia (lipopolysaccharide injection; LPS), we now report significant LPS-induced elevations of circulating H3Cit in 22 healthy individuals. We detected elevations of circulating H3Cit by enzyme-linked immunosorbent assay (ELISA), as well as bound to MVs quantified by flow cytometry. H3Cit-bearing MVs expressed neutrophil and/or platelet surface markers, indicating platelet-neutrophil interactions. In addition, in vitro experiments revealed that H3Cit can bind to phosphatidylserine exposed on platelet derived MVs. Taken together; our results demonstrate that NETs can be detected in peripheral blood during endotoxemia by two distinct H3Cit-specific methods. Furthermore, we propose a previously unrecognized mechanism by which H3Cit may be disseminated throughout the vasculature by the binding to MVs.

  • 37.
    Hamisi, Mariam
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Botaniska institutionen.
    Díez, Beatriz
    Institut de Ciéncies del Mar (ICM), CMIMA-CSIC, Barcelona.
    Lyimo, Thomas
    University of Dar es Salaam, Tanzania.
    Ininbergs, Karolina
    Stockholms universitet, Naturvetenskapliga fakulteten, Botaniska institutionen.
    Bergman, Birgitta
    Stockholms universitet, Naturvetenskapliga fakulteten, Botaniska institutionen.
    Cyanobacteria associated with the phyllosphere of the seagrass Cymodocea rotundata: Diversity, diel nifH expression and nitrogenase activity: Diversity, nifH expression and activity in seagrassManuskript (preprint) (Övrigt vetenskapligt)
    Abstract [en]

    Tropical seagrass ecosystems are highly productive and extremely important for sustaining marine life. As seagrasses are associated with complex assemblages of poorly examined epiphytic microbes, we proposed that nitrogen-fixing microorganisms may contribute to the productivity. The morphological and genetic diversity (based on the 16S rRNA and nifH genes) of cyanobacteria and diel variations in nifH gene expression, NifH protein levels and nitrogenase (nitrogen-fixing) activity were examined in the phyllosphere of Cymodocea rotundata of coastal areas of the western Indian Ocean (Tanzania). The 16S rRNA and nifH gene analyses during two consecutive years (October-November, 2007 and 2008) revealed the dominance of a mixed cyanobacterial community. Most sequences represented free-living uncultured cyanobacteria previously reported as benthic in the region, clearly separated from marine planktonic phylotypes, while a few sequences clustered with cyanobacterial symbionts of diatoms. Appreciable, but varying nitrogenase activities were found on a diel as well as monthly basis, with the highest activity encountered, 358 ± 232 and 258 ± 139 nmol C2H4 g-1 h-1, in November. On a diel basis, nifH gene expression coincided with the NifH protein level (Oct 2008) and nitrogenase activity. At day time, nifH gene expression primarily originated from heterocystous phylotypes, while from non-heterocystous filamentous phylotypes (mainly Oscillatoriales) at night. The data suggest that a variety of diazotrophic cyanobacteria are common among the epiphytes on Cymodocea and we propose that these may represent a valuable source of ‘new’ nitrogen in the often oligotrophic, but ecologically important seagrass ecosystems.

  • 38.
    Hoeppner, Marc Patrick
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylärbiologi och funktionsgenomik.
    The deep evolutionary roots of non-coding RNA - a comparative genomics approach2011Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
    Abstract [en]

    Non-coding RNAs (ncRNA) are a diverse group of genes that do not encode proteins but function exclusively on the level of RNA and were originally suggested to be remnants of a pre-DNA stage of life known as the RNA world. More recent work, however, has uncovered a rich repertoire of previously unknown families with possible consequences for our understanding of the origin and evolution of the modern RNA infrastructure. The main goal of this thesis was therefore to re-examine the evolutionary history of RNAs and theories regarding the transition from an RNA world in light of recent advances in molecular and computational biology.

    Using comparative genomics approaches and sequence data from all domains of life, my work shows that the majority of known RNAs exhibit a highly domain-specific distribution, compatible with an ongoing emergence rather than deep ancestry. Focusing on small nucleolar RNAs (snoRNA), I find that the eukaryote ancestor possessed a complex snoRNA infrastructure, but that intronic snoRNAs are mobile over larger evolutionary time scales. The latter has consequences for predictions made by the Introns-first hypothesis, a framework to explain the emergence of introns in an RNA world and which we revisited in light of advances in our understanding of the evolutionary dynamics of introns.

    A more in-depth analysis of ncRNA mobility across vertebrates found intronic copies of both snoRNAs and miRNAs to be more stable than intergenic ones, suggesting that this arrangement may be a consequence of co-expression. Also, snoRNAs are frequently located in highly expressed genes, in line with their role in ribosome biogenesis. Finally, a closer examination of the genomic distribution of two essential ncRNAs, snoRNA U3 and the spliceosomal RNA U1 shows that both are present in numerous copies across vertebrate genomes. Using next-generation sequencing data, I tested whether this is the result of genetic drift or a requirement for having many copies.

  • 39.
    Hoeppner, Marc Patrick
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylärbiologi och funktionsgenomik.
    Gardner, Paul P.
    Wellcome Trust Sanger Institute, Wellcome Trust Genome Campus.
    Poole, Anthony M.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylärbiologi och funktionsgenomik.
    Comparative analysis of RNA families reveals distinct repertoires for each domain of lifeManuskript (preprint) (Övrigt vetenskapligt)
    Abstract [en]

    Some RNAs may date back to an RNA-rich period in the early evolution of life, butmany RNAs are thought to have more recent evolutionary origins. To chart the broadevolutionary history of known RNA families, we performed comparative genomicanalysis of over 3 million RNA annotations spanning 1446 families from the Rfam 10database. We report that 99% of known RNA families are restricted to a singledomain of life, revealing discrete repertoires for each domain. For the 1% of RNAfamilies/clans present in more than one domain, over half show evidence ofhorizontal gene transfer (HGT), and only six RNAs directly trace to the LastUniversal Common Ancestor (LUCA). These results indicate that cellular RNAinfrastructure evolves in a domain-specific manner.

  • 40.
    Hoeppner, Marc Patrick
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylärbiologi och funktionsgenomik.
    Poole, Anthony M.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylärbiologi och funktionsgenomik.
    Comparative Genomics of Eukaryotic Small Nucleolar RNAs Reveals Deep Evolutionary Roots Amidst Ongoing Intragenomic MobilityManuskript (preprint) (Övrigt vetenskapligt)
    Abstract [en]

    Small nucleolar (sno)RNAs are required for posttranscriptional processing andmodification of ribosomal, spliceosomal and messenger RNAs. There are two broadclasses (C/D and H/ACA), both of which have been characterized in eukaryotes andarchaea. The association with ribosomal RNA processing and modification has led tothe suggestion that snoRNAs are evolutionarily ancient, and date back to the RNAworld. That numerous snoRNAs have been identified in the introns of ribosomalprotein genes has led to alternate views on the origin of this organization. Oneproposal is that intronic snoRNAs predate their surrounding protein-coding exons,the latter being recruited as messenger RNA following the origin of geneticallyencodedprotein synthesis. Another is that intron position reflects selection forcoexpression of snoRNAs and ribosomal components. To gain a clearer insight intothe antiquity of individual snoRNA families and the stability of their genomic location,we examined the evolutionary history of snoRNA families across 44 eukaryotegenomes. Our analysis reveals that dozens of snoRNA families can be traced backto the Last Eukaryotic Common Ancestor (LECA). However, none of the snoRNA1families placed in the LECA are sufficiently similar to characterized archaeal sno-likeRNAs, for us to confidently place specific snoRNA families in the common ancestorof archaea and eukaryotes. In agreement with earlier studies, we can tracenumerous introns to the LECA. However, snoRNAs housed within such positionallyconserved introns are not themselves orthologs. Morevover, our comparativegenomics analysis argues against evolutionarily-stable association betweensnoRNAs and individual host genes — analysis of host gene expression dataindicates that the primary requirement being for hosting intronic snoRNAs is a broadexpression profile. Consistent with mobility over antiquity, we report a case ofdemonstrable intronic snoRNA gain, where an evolutionarily ancient snoRNA hasmigrated into the intron of a mammalian mitochondrial ribosomal protein gene.Together, these data best fit a model wherein snoRNAs are intragenomically mobile,frequently residing in the introns of broadly-expressed protein-coding genes.

  • 41.
    Hoeppner, Marc Patrick
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylärbiologi och funktionsgenomik.
    Poole, Anthony M.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylärbiologi och funktionsgenomik.
    Maintenance of redundant small RNA gene copies over evolutionarytimescales via a retrotransposition motor?Manuskript (preprint) (Övrigt vetenskapligt)
    Abstract [en]

    We analysed the stability of duplicated, essential RNAs on the backdrop of theirexpression profiles to test whether the data is compatible with functional redundancy ordiversification. Under the former model, the expectation is that copies are equallyexpressed across tissues and subject to high turn-over. The latter model, in contrast,predicts that sub- or neofunctionalization following duplication may lead to a range ofcomplementary expression profiles across tissues. By example of the spliceosomal RNAU1 and snoRNA U3, we find that only few loci are stable over the course of vertebrateevolution and that the majority of copies show little or no expression. We conclude thatthese findings are most compatible with the redundancy model. Interestingly, the deepestloci are associated with a testis-expressed gene, suggesting a possible driving forcebehind the ongoing proliferation that we observe.

  • 42. Holthaus, Lisa
    et al.
    Lamp, Daniel
    Gavrisan, Anita
    Sharma, Virag
    Ziegler, Anette-Gabriele
    Jastroch, Martin
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut. German Research Center for Environmental Health, Germany.
    Bonifacio, Ezio
    CD4(+) T cell activation, function, and metabolism are inhibited by low concentrations of DMSO2018Ingår i: JIM - Journal of Immunological Methods, ISSN 0022-1759, E-ISSN 1872-7905, Vol. 463, s. 54-60Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Dimethyl sulfoxide (DMSO) is a polar organic solvent used in a wide range of biological applications. DMSO is routinely used as a cryoprotectant for long-term cell freezing as well as to dissolve peptides and drugs for immune cell functional assays. Here, human CD4(+) T cell activation, cytokine production, proliferation, and metabolism were investigated after stimulation in the presence of 0.01% to 1%, DMSO, representing concentrations commonly used in vitro. Surface expression of the activation markers CD69, CD25 and CD154 after polyclonal activation of CD4(+) T cells was inhibited by 0.25% or higher concentrations of DMSO. The frequencies of IL-21(+), IL-4(+), and IL-22(+) CD4(+) T cells, following polyclonal activation were variably inhibited by DMSO at concentrations ranging from 0.25% to 1%, whereas IFN gamma(+) cells were unaffected. CD4(+) T cell proliferation after anti-CD3 or antigen stimulation was inhibited by 0.5% DMSO and abolished by 1% DMSO. After polyclonal stimulation, glucose uptake was inhibited in the presence of 1% DMSO, but only minor effects on CD4+ T cell respiration were observed. Consistent with the immune effects, the gene expression of early signaling and activation pathways were inhibited in CD4+ T cells in the presence of 1% DMSO. Our study revealed that DMSO at concentrations generally used for in vitro studies of T cells impacts multiple features of T cell function. Therefore, we urge care when adding DMSO-containing preparations to T cell cultures.

  • 43. Huque, Yasmin
    Radical generation and transfer in ribonucleotide reductase2001Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
  • 44. Högstrand, Kari
    et al.
    Lindvall, Jessica M.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik. Stockholms universitet, Science for Life Laboratory (SciLifeLab).
    Sundblad, Anne
    Grandien, Alf
    Transformation of mature mouse B cells into malignant plasma cells in vitro via introduction of defined genetic elements2019Ingår i: European Journal of Immunology, ISSN 0014-2980, E-ISSN 1521-4141, Vol. 49, nr 3, s. 454-461Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    An experimental system where defined alterations in gene function or gene expression levels in primary B cells would result in the development of transformed plasma cells in vitro would be useful in order to facilitate studies of the underlying molecular mechanisms of plasma cell malignancies. Here, such a system is described in which primary murine B cells rapidly become transformed into surface CD138(+), IgM(-/low), CD19(-) IgM-secreting plasma cells as a result of expression of the transcription factors IRF4 and MYC together with simultaneous expression of BMI1, mutated p53 or silencing of p19(Arf), and suppression of intrinsic apoptosis through expression of BCLXL. Analysis of gene expression patterns revealed that this combination of transforming genes resulted in expression of a number of genes previously associated with terminally differentiated B cells (plasma cells) and myeloma cells, whereas many genes associated with mature B cells and B-cell lymphomas were not expressed. Upon transplantation, the transformed cells preferentially localized to the bone marrow, presenting features of a plasma cell malignancy of the IgM isotype. The present findings may also be applicable in the development of novel methods for production of monoclonal antibodies.

  • 45. Imai, Kenichiro
    et al.
    Hayat, Sikander
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Sakiyama, Noriyuki
    Fujita, Naoya
    Tomii, Kentaro
    Elofsson, Arne
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Horton, Paul
    Localization prediction and structure-based in silico analysis of bacterial proteins: with emphasis on outer membrane proteins.2013Ingår i: Data Mining for Systems Biology / [ed] Hiroshi Mamitsuka, Charles DeLisi, Minoru Kanehisa, New York: Humana Press, 2013, Vol. 939, s. 115-140Kapitel i bok, del av antologi (Refereegranskat)
    Abstract [en]

    In this chapter, we first discuss protein localization in bacteria and evaluate some localization prediction tools on an independent dataset. Next, we focus on β-barrel outer membrane proteins (BOMPs), describing and evaluating new tools for BOMP detection and topology prediction. Finally, we apply general protein structure prediction methods on these proteins to show that the structure of most BOMPs in E. coli can be modeled reliably.

  • 46. Isidor, Marie S.
    et al.
    Winther, Sally
    Basse, Astrid L.
    Petersen, M. Christine H.
    Cannon, Barbara
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Nedergaard, Jan
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Hansen, Jacob B.
    An siRNA-based method for efficient silencing of gene expression in mature brown adipocytes2016Ingår i: Adipocyte, ISSN 2162-3945, E-ISSN 2162-397X, Vol. 5, nr 2, s. 175-185Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Brown adipose tissue is a promising therapeutic target for opposing obesity, glucose intolerance and insulin resistance. The ability to modulate gene expression in mature brown adipocytes is important to understand brown adipocyte function and delineate novel regulatory mechanisms of non-shivering thermogenesis. The aim of this study was to optimize a lipofection-based small interfering RNA (siRNA) transfection protocol for efficient silencing of gene expression in mature brown adipocytes. We determined that a critical parameter was to deliver the siRNA to mature adipocytes by reverse transfection, i.e. transfection of non-adherent cells. Using this protocol, we effectively knocked down both high-and low-abundance transcripts in a model of mature brown adipocytes (WT-1) as well as in primary mature mouse brown adipocytes. A functional consequence of the knockdown was confirmed by an attenuated increase in uncoupled respiration (thermogenesis) in response to beta-adrenergic stimulation of mature WT-1 brown adipocytes transfected with uncoupling protein 1 siRNA. Efficient gene silencing was also obtained in various mouse and human white adipocyte models (3T3-L1, primary mouse white adipocytes, hMADS) with the ability to undergo browning. In summary, we report an easy and versatile reverse siRNA transfection protocol to achieve specific silencing of gene expression in various models of mature brown and browning-competent white adipocytes, including primary cells.

  • 47. Jensen, Lasse D.
    et al.
    Hot, Belma
    Ramsköld, Daniel
    Germano, Raoul F. V.
    Yokota, Chika
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik. Ludwig Institute for Cancer Research Ltd, Sweden.
    Giatrellis, Sarantis
    Lauschke, Volker M.
    Hubmacher, Dirk
    Li, Minerva X.
    Hupe, Mike
    Arnold, Thomas D.
    Sandberg, Rickard
    Frisén, Jonas
    Trusohamn, Marta
    Martowicz, Agnieszka
    Wisniewska-Kruk, Joanna
    Nyqvist, Daniel
    Adams, Ralf H.
    Apte, Suneel S.
    Vanhollebeke, Benoit
    Stenman, Jan M.
    Kele, Julianna
    Disruption of the Extracellular Matrix Progressively Impairs Central Nervous System Vascular Maturation Downstream of beta-Catenin Signaling2019Ingår i: Arteriosclerosis, Thrombosis and Vascular Biology, ISSN 1079-5642, E-ISSN 1524-4636, Vol. 39, nr 7, s. 1432-1447Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Objective- The Wnt/beta-catenin pathway orchestrates development of the blood-brain barrier, but the downstream mechanisms involved at different developmental windows and in different central nervous system (CNS) tissues have remained elusive. Approach and Results- Here, we create a new mouse model allowing spatiotemporal investigations of Wnt/beta-catenin signaling by induced overexpression of Axin1, an inhibitor of beta-catenin signaling, specifically in endothelial cells (Axin1(iEC)-(OE)). AOE (Axin1 overexpression) in Axin1(iEC)-(OE) mice at stages following the initial vascular invasion of the CNS did not impair angiogenesis but led to premature vascular regression followed by progressive dilation and inhibition of vascular maturation resulting in forebrain-specific hemorrhage 4 days post-AOE. Analysis of the temporal Wnt/beta-catenin driven CNS vascular development in zebrafish also suggested that Axin1(iEC)-(OE) led to CNS vascular regression and impaired maturation but not inhibition of ongoing angiogenesis within the CNS. Transcriptomic profiling of isolated, beta-catenin signaling-deficient endothelial cells during early blood-brain barrier-development (E11.5) revealed ECM (extracellular matrix) proteins as one of the most severely deregulated clusters. Among the 20 genes constituting the forebrain endothelial cell-specific response signature, 8 (Adamtsl2, Apod, Ctsw, Htra3, Pglyrp1, Spock2, Ttyh2, and Wfdc1) encoded bona fide ECM proteins. This specific beta-catenin-responsive ECM signature was also repressed in Axin1(iEC)-(OE) and endothelial cell-specific beta-catenin-knockout mice (Ctnnb1-KOiEC) during initial blood-brain barrier maturation (E14.5), consistent with an important role of Wnt/beta-catenin signaling in orchestrating the development of the forebrain vascular ECM. Conclusions- These results suggest a novel mechanism of establishing a CNS endothelium-specific ECM signature downstream of Wnt-beta-catenin that impact spatiotemporally on blood-brain barrier differentiation during forebrain vessel development.

  • 48. Jevtusevskaja, Jekaterina
    et al.
    Krolov, Katrin
    Tulp, Indrek
    Langel, Ülo
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi. University of Tartu, Estonia.
    The effect of main urine inhibitors on the activity of different DNA polymerases in loop-mediated isothermal amplification2017Ingår i: Expert Review of Molecular Diagnostics, ISSN 1473-7159, E-ISSN 1744-8352, Vol. 17, nr 4, s. 403-410Artikel, forskningsöversikt (Refereegranskat)
    Abstract [en]

    Background: The use of rapid amplification methods to detect pathogens in biological samples is mainly limited by the amount of pathogens present in the sample and the presence of inhibiting substances. Inhibitors can affect the amplification efficiency by either binding to the polymerase, interacting with the DNA, or interacting with the polymerase during primer extension. Amplification is performed using DNA polymerase enzymes and even small changes in their activity can influence the sensitivity and robustness of molecular assaysMethods: The main purpose of this research was to examine which compounds present in urine inhibit polymerases with strand displacement activity. To quantify the inhibition, we employed quantitative loop-mediated isothermal amplificationResults: The authors found that the presence of BSA, Mg 2+, and urea at physiologically relevant concentrations, as well as acidic or alkaline conditions did not affect the activity of any of the tested polymerases. However, addition of salt significantly affected the activity of the tested polymerases.Conclusion: These findings may aid in the development of more sensitive, robust, cost effective isothermal amplification based molecular assays suitable for both point-of-care testing and on-site screening of pathogens directly from unprocessed urine which avoid the need for long and tedious DNA purification steps prior to amplification.

  • 49.
    Johnsson, Anna-Karin
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Wenner-Grens institut, Avdelningen för cellbiologi.
    Karlsson, Roger
    Stockholms universitet, Naturvetenskapliga fakulteten, Wenner-Grens institut, Avdelningen för cellbiologi.
    Microtubule-dependent localization of profilin I mRNA to actin polymerization sites in serum-stimulated cells2010Ingår i: European Journal of Cell Biology, ISSN 0171-9335, E-ISSN 1618-1298, Vol. 89, nr 5, s. 394-401Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Specific localization of messenger RNA (mRNA) appears to be a general mechanism to accumulate certain proteins to subcellular compartments for participation in local processes, thereby maintaining cell polarity under strict spatiotemporal control. Transportation of mRNA with associated protein components (RNP granules) by the actin microfilament or the microtubule systems is one important mechanism to achieve this locally distributed protein production. Here we provide evidence for a microtubule-dependent localization of mRNA encoding the actin regulatory protein profilin to sites in mouse embryonic fibroblasts, which express enhanced actin polymerization.

  • 50.
    Kanatani, Sachie
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Fuks, Jonas M.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Olafsson, Einar B.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Westermark, Linda
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Chambers, Benedict
    Varas-Godoy, Manuel
    Uhlén, Per
    Barragan, Antonio
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Voltage-dependent calcium channel signaling mediates GABA(A) receptor-induced migratory activation of dendritic cells infected by Toxoplasma gondii2017Ingår i: PLoS Pathogens, ISSN 1553-7366, E-ISSN 1553-7374, Vol. 13, nr 12, artikel-id e1006739Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The obligate intracellular parasite Toxoplasma gondii exploits cells of the immune system to disseminate. Upon T. gondii-infection,. Upsilon-aminobutyric acid (GABA)/GABAA receptor signaling triggers a hypermigratory phenotype in dendritic cells (DCs) by unknown signal transduction pathways. Here, we demonstrate that calcium (Ca2+) signaling in DCs is indispensable for T. gondii-induced DC hypermotility and transmigration in vitro. We report that activation of GABAA receptors by GABA induces transient Ca2+ entry in DCs. Murine bone marrow-derived DCs preferentially expressed the L-type voltage-dependent Ca2+ channel (VDCC) subtype Cav1.3. Silencing of Cav1.3 by short hairpin RNA or selective pharmacological antagonism of VDCCs abolished the Toxoplasma-induced hypermigratory phenotype. In a mouse model of toxoplasmosis, VDCC inhibition of adoptively transferred Toxoplasma-infected DCs delayed the appearance of cell-associated parasites in the blood circulation and reduced parasite dissemination to target organs. The present data establish that T. gondii-induced hypermigration of DCs requires signaling via VDCCs and that Ca2+ acts as a second messenger to GABAergic signaling via the VDCC Cav1.3. The findings define a novel motility-related signaling axis in DCs and unveil that interneurons and DCs share common GABAergic motogenic pathways. T. gondii employs GABAergic non-canonical pathways to induce host cell migration and facilitate dissemination.

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