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  • 1. Adamsson, I
    et al.
    Edlund, Charlotta
    Södertörn University, Avdelning Naturvetenskap.
    Seensalu, R
    Engstrand, L
    The use of AP-PCR and flaA-RFLP typing to investigate treatment failure in Helicobacter pylori infection2000In: Clinical Microbiology and Infection, ISSN 1198-743X, E-ISSN 1469-0691, Vol. 6, no 5, p. 265-267Article in journal (Refereed)
  • 2. Agianian, Bogos
    et al.
    Lesch, Christine
    Loseva, Olga
    Dushay, Mitchell S.
    Södertörn University, School of Life Sciences. Uppsala University.
    Preliminary characterization of hemolymph coagulation in Anopheles gambiae larvae2007In: Developmental and Comparative Immunology, ISSN 0145-305X, E-ISSN 1879-0089, Vol. 31, no 9, p. 879-888Article in journal (Refereed)
    Abstract [en]

    Hemolymph coagulation is a first response to injury, impeding infection, and ending bleeding. Little is known about its molecular basis in insects, but clotting factors have been identified in the fruit fly Drosophila melanogaster. Here, we have begun to study coagulation in the aquatic larvae of the malaria vector mosquito Anopheles gambiae using methods developed for Drosophila. A delicate clot was seen by light microscopy, and pullout and proteomic analysis identified phenoloxidase and apolipophorin-I as major candidate clotting factors. Electron microscopic analysis confirmed clot formation and revealed it contains fine molecular sheets, most likely a result of lipophorin assembly. Phenoloxidase appears to be more critical in clot formation in Anopheles than in Drosophila. The Anopheles larval clot thus differs in formation, structure, and composition from the clot in Drosophila, confirming the need to study coagulation in different insect species to learn more about its evolution and adaptation to different lifestyles.

  • 3. Agvald-Öhman, C
    et al.
    Wernerman, J
    Nord, C E
    Edlund, Charlotta
    Södertörn University, Avdelning Naturvetenskap.
    Anaerobic bacteria commonly colonize the lower airways of intubated ICU patients2003In: Clinical Microbiology and Infection, ISSN 1198-743X, E-ISSN 1469-0691, Vol. 9, no 5, p. 397-405Article in journal (Refereed)
    Abstract [en]

    Objectives To investigate respiratory tract colonization by aerobic and anaerobic bacteria in mechanically ventilated patients. Methods Bacterial colonization of the stomach and the respiratory tract was qualitatively and quantitatively analyzed over time in 41 consecutive mechanically ventilated patients in a Swedish intensive care unit (ICU), with special emphasis on elucidation of the role of anaerobic bacteria in the lower respiratory tract. Samples were taken from the oropharynx, gastric juice, subglottic space and trachea within 24 h (median 14 h) of intubation, and then every third day until day 18 and every fifth day until day 33. Results The patients were often heavily colonized with microorganisms not considered to belong to a healthy normal oropharyngeal and gastric flora on admission to the ICU. A majority harbored enterococci, coagulase-negative staphylococci and Candida spp. in at least one site on day 1. Anaerobic bacteria, mainly peptostreptococci and Prevotella spp., were isolated from subglottic and/or tracheal secretions in 59% of the patients. Different routes of tracheal colonization for different groups of microorganisms were found. Primary or concomitant colonization of the oropharynx with staphylococci, enterococci, enterobacteria and Candida was often seen, while Pseudomonas spp., other non-fermenting Gram-negative rods and several anaerobic species often primarily colonized the trachea, indicating exogenous or direct gastrointestinal routes of colonization. Conclusions Mechanically ventilated patients were heavily colonized in their lower airways by potential pathogenic microorganisms, including a high load of anaerobic bacteria. Different routes of colonization were shown for different species.

  • 4. Ahnström, G.
    et al.
    Nygren, Jonas
    Södertörn University, Avdelning Naturvetenskap. Karlinska Instiutet.
    Eriksson, S.
    The effect of dimethyl sulphoxide on the induction and repair of double-strand breaks in human cells after irradiation with γ-rays and accelerated ions: Rapid or slow repair may depend on accessibility of breaks in chromatin of different compactness2000In: International Journal of Radiation Biology, ISSN 0955-3002, E-ISSN 1362-3095, Vol. 76, no 4, p. 533-538Article in journal (Refereed)
    Abstract [en]

    Purpose: The repair of double-strand breaks (dsb) in mammalian cells is characterized by a rapid phase with a half-life of less than half an hour and a slower phase that lasts for many hours. The proportion of slow repair increase with LET and it has been suggested that the slow repair component consists of more complex damage and is more deleterious to the cells. To see if removal of OH radicals could remove part of the damage in complex dsb and make them easier to repair, human cells were irradiated in the presence of dimethyl sulphoxide (DMSO). Methods: Induction and repair of dsb were studied by neutral elution in human VH10 cells exposed to γ-rays, helium ions (mean LET 40 keV/μm) and 80 and 125 keV/μm monoenergetic nitrogen ions in the presence and absence of 2 M DMSO. Results: Incubation of cells exposed to γ-rays, 40 keV/μm helium and 80 keV/μm N ions demonstrated that scavenging of OH radicals by DMSO removed most of the rapid repair component. The response to DMSO was less marked after 125 keV/μm nitrogen ions, where about half of the repair was resistant to DMSO. Conclusions: It is unlikely that the complexity of dsb is responsible for the slow repair because the removal of OH radicals did not make the breaks easier to repair. Instead, it is suggested that rapid and slow repair can be explained on the basis of how different parts of the chromatin are accessible to repair enzymes.

  • 5.
    Alheim, Katarina
    et al.
    Södertörn University, Avdelning Naturvetenskap. Karolinska Institutet.
    Corness, J
    Samuelsson, M K R
    Bladh, L G
    Murata, T
    Nilsson, T
    Okret, S
    Identification of a functional glucocorticoid response element in the promoter of the cylcin-dependant kinase inhibitor p57(Kip2)2003In: Journal of Molecular Endocrinology, ISSN 0952-5041, E-ISSN 1479-6813, Vol. 30, no 3, p. 359-368Article in journal (Refereed)
    Abstract [en]

    Glucocorticoids are known regulators of the cell cycle, normally exerting an anti-proliferative effect. We have previously shown that glucocorticoids stimulate expression of p57(Kip2), a member of the Cip/Kip family of cyclin-dependent kinase inhibitors which, in some cell types, may account for the anti-proliferative responses seen after glucocorticoid treatment. The induction of p57(Kip2) involves primary transcriptional effects where no de novo protein synthesis is necessary, suggesting a direct interaction of the glucocorticoid receptor with the p57(Kip2) gene. In this study we have identified a functional glucocorticoid response element (GRE), located 5 kilo bases (kb) upstream of the transcription start site in the human P57(Kip2) promoter. This GRE was functional also when isolated, suggesting a direct transcriptional effect of the glucocorticoid receptor. Furthermore, mutation of this GRE abolished glucocorticoid induction of the reporter gene, whereas mutation of a nearby Sp1 site did not. Using electrophoretic mobility shift assays, we have shown that the -5 kb p57(Kip2) promoter GRE was able to compete with a well-known GRE for glucocorticoid receptor binding. Sequence comparisons with the mouse genome showed that this GRE is highly conserved, further strengthening the biological importance of this site. All these data emphasize the involvement of this GRE in the glucocorticoid-mediated induction of p57(Kip2) expression.

  • 6.
    Ali, Fuad
    Södertörn University College, School of Life Sciences.
    Developing electroporation as a method to obtain Stable Transformation in Drosophila melanogaster2008Independent thesis Advanced level (degree of Master (Two Years)), 20 credits / 30 HE creditsStudent thesis
    Abstract [en]

    In this project I have tried to obtain stable transformants of Drosophila melanogaster flies using electroporation. I have completed approximately 200 tests using different DNA concentrations, voltages and cuvettes, including a novel Petri dish cuvette which I developed and manufactured myself. I also developed new and more efficient procedures of egg collection and egg dechorionation. Although I was not  successful in obtaining true stable transformants, control experiments indicate that electroporation of DNA into embryos could be accomplished under the conditions used. The lack of stable transformants was probably due to failure of the electroporated DNA to integrate into the host genome. The reasons for why the DNA did not integrate was not further investigated in this study.

  • 7.
    Aliyu, Habibu
    et al.
    Centre for Microbial Ecology and Genomics, University of Pretoria, Pretoria, South Africa.
    De Maayer, Pieter
    School of Molecular and Cell Biology, University of the Witwatersrand, Johannesburg, South Africa.
    Sjöling, Sara
    Södertörn University, School of Natural Sciences, Technology and Environmental Studies, Environmental Science.
    Cowan, Donald A.
    Centre for Microbial Ecology and Genomics, University of Pretoria, Pretoria, South Africa.
    Metagenomic Analysis of Low-Temperature Environments2017In: Psychrophiles: From Biodiversity to Biotechnology / [ed] Rosa Margesin, Cham: Springer, 2017, p. 389-421Chapter in book (Refereed)
    Abstract [en]

    The Earth’s permanently cold biosphere is known to harbour abundant microbial biomass and represents a rich resource for the discovery of novel cold-adapted microorganisms, many of which form part of the ‘microbial dark matter’ which cannot be analysed using traditional culture-dependent approaches. The recent development of metagenomics and related multi-omics strategies has provided a means by which entire microbial communities can be studied directly, without the prerequisite of culturing. The advancement of the ‘omic’ methods is directly linked to recent progress in high-throughput sequencing, robust data processing capabilities and the application of cutting-edge analytical tools for high-throughput detection of biomolecules. The combined application of these tools and strategies has provided an unprecedented access to the structure and potential function of microbial communities in cold environments, providing increasingly comprehensive insights into the taxonomic richness and functional capacity of the indigenous microorganisms. Applications of ‘omic’ strategies have enhanced our understanding of psychrophilic adaptation mechanisms, revealing the versatility and adaptability of life in the ‘cryosphere’. In addition to the predicted roles of psychrophiles in biogeochemical cycling, recent multi-omic studies have further emphasised the importance of the ‘cryosphere’ in influencing global atmospheric conditions. Finally, metagenomic bioprospecting of cold environments has yielded a variety of novel bioactive molecules including novel ‘psychrozymes’, with a wide range of potential industrial and biotechnological applications. Here, we have provided an overview of recent developments in metagenomic technologies and their application in the study of the cold biosphere.

  • 8.
    Alkemar, Gunnar
    et al.
    Södertörn University, Avdelning Naturvetenskap. Stockholm Universtiy.
    Nygård, Odd
    Södertörn University, Avdelning Naturvetenskap.
    A possible tertiary rRNA interaction between expansion segments ES3 and ES6 in eukaryotic 40S ribosomal subunits2003In: RNA: A publication of the RNA Society, ISSN 1355-8382, E-ISSN 1469-9001, Vol. 9, no 1, p. 20-24Article in journal (Refereed)
    Abstract [en]

    Eukaryotic 16S-like ribosomal RNAs contain 12 so-called expansion segments, i.e., sequences not included in the RNA secondary structure core common to eubacteria, archaea, and eukarya. Two of these expansion segments, ES3 and ES6, are juxtaposed in the recent three-dimensional model of the eukaryotic 40S ribosomal subunit. We have analyzed ES3 and ES6 sequences from more than 2900 discrete eukaryotic species, for possible sequence complementarity between the two expansion segments. The data show that ES3 and ES6 could interact by forming a helix consisting of seven to nine contiguous base pairs in almost all analyzed species. We, therefore, suggest that ES3 and ES6 form a direct RNA-RNA contact in the ribosome.

  • 9.
    Alkemar, Gunnar
    et al.
    Stockholms universitet, Wenner-Grens institut.
    Nygård, Odd
    Södertörn University, School of Life Sciences.
    Comparative analysis of rRNA sequences from the large ribosomal subunit of more than 900 eukaryotic species reveals structural similarities in expansion segment ES39.Manuscript (preprint) (Other academic)
  • 10.
    Alkemar, Gunnar
    et al.
    Södertörn University, School of Life Sciences. Stockholm University.
    Nygård, Odd
    Södertörn University, School of Life Sciences.
    Probing the secondary structure of expansion segment ES6 in 18S ribosomal RNA2006In: Biochemistry, ISSN 0006-2960, E-ISSN 1520-4995, Vol. 45, no 26, p. 8067-8078Article in journal (Refereed)
    Abstract [en]

    Expansion segment ES6 in 18S ribosomal RNA is, unlike many other expansion segments, present in all eukaryotes. The available data suggest that ES6 is located on the surface of the small ribosomal subunit. Here we have analyzed the secondary structure of the complete ES6 sequence in intact ribosomes from three eukaryotes, wheat, yeast, and mouse, representing different eukaryotic kingdoms. The availability of the ES6 sequence for modification and cleavage by structure sensitive chemicals and enzymatic reagents was analyzed by primer extension and gel electrophoresis on an ABI 377 automated DNA sequencer. The experimental results were used to restrict the number of possible secondary structure models of ES6 generated by the folding software MFOLD. The modification data obtained from the three experimental organisms were very similar despite the sequence variation. Consequently, similar secondary structure models were obtained for the ES6 sequence in wheat, yeast, and mouse ribosomes. A comparison of sequence data from more than 6000 eukaryotes showed that similar structural elements could also be formed in other organisms. The comparative analysis also showed that the extent of compensatory base changes in the suggested helices was low. The in situ structure analysis was complemented by a secondary structure analysis of wheat ES6 transcribed and folded in vitro. The obtained modification data indicate that the secondary structure of the in vitro transcribed sequence differs from that observed in the intact ribosome. These results suggest that chaperones, ribosomal proteins, and/or tertiary rRNA interactions could be involved in the in vivo folding of ES6.

  • 11.
    Alkemar, Gunnar
    et al.
    Södertörn University, School of Chemistry, Biology, Geography and Environmental Science. Stockholm University.
    Nygård, Odd
    Södertörn University, School of Chemistry, Biology, Geography and Environmental Science.
    Secondary structure of two regions in expansion segments ES3 and ES6 with the potential of forming a tertiary interaction in eukaryotic 40S ribosomal subunits2004In: RNA: A publication of the RNA Society, ISSN 1355-8382, E-ISSN 1469-9001, Vol. 10, no 3, p. 403-411Article in journal (Refereed)
    Abstract [en]

    The 18S rRNA of the small eukaryotic ribosomal subunit contains several expansion segments. Electron microscopy data indicate that two of the largest expansion segments are juxtaposed in intact 40S subunits, and data from phylogenetic sequence comparisons indicate that these two expansion segments contain complementary sequences that could form a direct tertiary interaction on the ribosome. We have investigated the secondary structure of the two expansion segments in the region around the putative tertiary interaction. Ribosomes from yeast, wheat, and mouse-three organisms representing separate eukaryotic kingdoms-were isolated, and the structure of ES3 and part of the ES6 region were analyzed using the single-strand-specific chemical reagents CMCT and DMS and the double-strand-specific ribonuclease V1. The modification patterns were analyzed by primer extension and gel electrophoresis on an ABI 377 automated DNA sequencer. The investigated sequences were relatively exposed to chemical and enzymatic modification. This is in line with their indicated location on the surface at the solvent side of the subunit. The complementary ES3 and ES6 sequences were clearly inaccessible to single-strand modification, but available for cleavage by double-strand-specific RNase V1. The results are compatible with a direct helical interaction between bases in ES3 and ES6. Almost identical results were obtained with ribosomes from the three organisms investigated.

  • 12.
    Andrén, Elinor
    et al.
    Södertörn University, School of Natural Sciences, Technology and Environmental Studies, Environmental Science.
    Andrén, Thomas
    Södertörn University, School of Natural Sciences, Technology and Environmental Studies, Environmental Science.
    Syrefria bottnar - orsakade av klimat, människa eller både och?2014In: Havsutsikt, ISSN 1104-0513, no 2, p. 12-14Article in journal (Other (popular science, discussion, etc.))
  • 13. Antuch, W
    et al.
    Berndt, Kurt D
    Eidgenössische Technische Hochschule–Hönggerberg, Zürich, Switzerland.
    Chávez, M A
    Delfín, J
    Wüthrich, K
    The NMR solution structure of a Kunitz-type proteinase inhibitor from the sea anemone Stichodactyla helianthus.1993In: European Journal of Biochemistry, ISSN 0014-2956, E-ISSN 1432-1033, Vol. 212, p. 675-684Article in journal (Refereed)
    Abstract [en]

    The solution structure of a 55-amino-acid Kunitz-type proteinase inhibitor, ShPI, purified from the Caribbean sea anemone Stichodactyla helianthus, was determined by NMR spectroscopy. Nearly complete sequence-specific 1H-NMR assignments were obtained at pH 4.6 and 36 degrees C, and stereo-specific assignments were determined for 23 pairs of diastereotopic substituents. A data set of 666 upper distance limit constraints and 122 dihedral angle constraints collected on this basis was used as input for a structure calculation with the program DIANA. Following energy minimization with the program OPAL, the average root-mean-square diviation (RMSD) of the 20 DIANA conformers used to represent the solution structure relative to the mean structure is 61 pm for all backbone atoms N, C alpha and C', and 106 pm for all heavy atoms of residues 2-53. This high-quality solution structure of ShPI has a nearly identical molecular architecture as the bovine pancreatic trypsin inhibitor (BPTI), despite a mere 35% of sequence similarity between the two proteins. Exchange rates measured for 48 out of the 51 backbone amide protons showed that the positions of 20 slowly exchanging amide protons correlate well with hydrogen bonds involving these protons in the energy-minimized solution structure. The solution structure of ShPI is compared to the four homologous proteins for which the three-dimensional structure is also available.

  • 14.
    Appelgren, Henrik
    et al.
    Södertörn University, Avdelning Naturvetenskap. Karolinska Institute.
    Kniola, Barbara
    Södertörn University, Avdelning Naturvetenskap. Karolinska Institute.
    Ekwall, Karl
    Södertörn University, Avdelning Naturvetenskap. Karolinska Institute.
    Distinct centromere domain structures with separate functions demonstrated in live fission yeast cells2003In: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 116, no 19, p. 4035-4042Article in journal (Refereed)
    Abstract [en]

    Fission yeast (Saccharomyces pombe) centromere DNA is organized in a central core region flanked on either side by a region of outer repeat (otr) sequences. The otr region is known to be heterochromatic and bound by the Swi6 protein whereas the central core region contains an unusual chromatin structure involving the histone H3 variant Cnp1 (S. pombe CENP-A). The central core is the base for formation of the kinetochore structure whereas the flanking region is important for sister centromere cohesion. We have previously shown that the ultrastructural domain structure of S. pombe centromeres in interphase is similar to that of human centromeres. Here we demonstrate that S. pombe centromeres are organized in cytologically distinct domains even in mitosis. Fluorescence in situ hybridization of fixed metaphase cells revealed that the otr regions of the centromere were still held together by cohesion even after the sister kinetochores had separated. In live cells, the central cores and kinetochores of sister chromosomes could be distinguished from one Another when they were subjected to mitotic tension. The function of the different centromeric domains was addressed. Transacting mutations affecting the kinetochore (nuf2) central core domain (mis6) and the heterochromatin domain (rik1) were analyzed in live cells. In interphase, both nuf2 and mis6 caused declustering of centromeres from the spindle pole body whereas centromere clustering was normal in rik1 despite an apparent decondensation defect. The declustering of centromeres in mis6 cells correlated with loss the Ndc80 kinetochore marker protein from the centromeres. Interestingly the declustered centromeres were still restricted to the nuclear periphery thus revealing a kinetochore-independent peripheral localization mechanism for heterochromatin. Time-lapse microscopy of live mis6 and nuf2-1 mutant cells in mitosis showed similar severe misaggregation phenotypes whereas the rik1 mutants showed a mild cohesion defect. Thus, S. pombe centromeres have two distinguishable domains even during mitosis, and our functional analyses support the previous observations that the kinetochore/central core and the heterochromatin domains have distinct functions both in interphase and mitosis.

  • 15.
    Arabi, Azadeh
    Södertörn University, School of Life Sciences. Karolinska Institutet.
    Regulation of the ribosomal RNA transcription by c-MYC oncoprotein2006Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    The transcription factor c-Myc is a key regulator of growth and proliferation. c-Myc levels are tightly controlled and deregulated c-Myc is often associated with human cancers. In our initial studies we observed that upon inhibition of proteasomes, excess c-Myc accumulates primarily in the nucleoli. After further investigation we could show that c-Myc binds to and activates RNA polymerase I-mediated transcription of the ribosomal RNA (rRNA) genes located in the nucleoli and that proteasomes are involved in this process. We demonstrate that upon an increase in c-Myc levels through either inhibition of the proteasomes or high expression, c-Myc accumulates in the nucleoli. The dynamics of the nucleoplasmic and the nucleolar c-Myc was studied in living cells expressing GFP-fused cMyc using the Fluorescent loss in photo-bleaching and the Fluorescent recovery after photobleaching techniques. We show that c-Myc is relatively stably associated with the nucleoli. In addition, we show that proteasomes accumulate and co-localise with nucleolar c-Myc. We further investigate the function of c-Myc in the nucleoli and show that c-Myc and Max interact in the nucleoli and are associated with the ribosomal DNA. Upon mitogenic stimulation of quiescent human lymphocytes c-Myc is recruited to the rRNA genes together with pol I. Association of c-Myc with the rDNA is also accompanied by an increase in rDNA histone acetylation and activation of rRNA transcription. Inhibition of c-Myc inhibits rRNA transcription. These results suggest that c-Myc plays a key role in regulating ribosome biogenesis and thus cell growth. We also show that proteasomes are required for activation of rRNA transcription, even though c-Myc levels increase in response to reduced proteasome activity. The role of proteasomes in rDNA transcription remains to be determined. We also investigate the role of c-Myc in regulation of the nucleolar organisation and induction of nucleolar alterations in cancer cells. Several types of human cancers with nucleolar alterations including cancers of blood, prostate and breast are also associated with deregulated levels of c-Myc. However, it is not known whether c-Myc contributes to the induction of nucleolar changes in these cancers. We show that despite high levels, c-Myc does not accumulate in the nucleoli in lymphoma and breast cancer cell lines. This is intriguing since nucleolar accumulation of excess c-Myc in other cell lines is associated with inhibition of rRNA transcription.

  • 16.
    Arabi, Azadeh
    et al.
    Södertörn University, School of Life Sciences. Karolinska Institutet.
    Berkson, Rachel
    Wright, Anthony P.
    Södertörn University, School of Life Sciences.
    Regulation of the nucleolar structure by the oncoprotein c-Myc and proteasomesManuscript (preprint) (Other academic)
  • 17.
    Arabi, Azadeh
    et al.
    Södertörn University, Avdelning Naturvetenskap. Karolinska Institute.
    Rustum, Cecilia
    Södertörn University, Avdelning Naturvetenskap. Stockholm University.
    Hallberg, Einar
    Södertörn University, Avdelning Naturvetenskap.
    Wright, Anthony P H
    Södertörn University, Avdelning Naturvetenskap. Karolinska Institute.
    Accumulation of c-Myc and proteasomes at the nucleoli of cells containing elevated c-Myc protein levels2003In: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 116, no 9, p. 1707-1717Article in journal (Refereed)
    Abstract [en]

    c-Myc is a predominately nuclear transcription factor that is a substrate for rapid turnover by the proteasome system. Cancer-related mutations in c-Myc lead to defects in its degradation and thereby contribute to the increase in its cellular level that is associated with the disease. Little is known about the mechanisms that target c-Myc to the proteasomes. By using a GFP fusion protein and live analysis we show that c-Myc shuttles between the nucleus and cytoplasm and thus it could be degraded in either compartment. Strikingly, at elevated levels of expression c-Myc accumulates at nucleoli in some cells, consistent with saturation of a nucleolus-associated degradation system in these cells. This idea is further supported by the observation that proteasome inhibitor treatment causes accumulation of c-Myc at the nucleoli of essentially all cells. Under these conditions c-Myc is relatively stably associated with the nucleolus, as would be expected if the nucleolus functions as a sequestration/degradation site for excess c-Myc. Furthermore, during elevated c-Myc expression or proteasome inhibition, nucleoli that are associated with c-Myc also accumulate proteasomes. c-Myc and proteasomes co-localise in intranucleolar regions distinct from the dense fibrillar component of the nucleolus. Based on these results we propose a model for c-Myc downregulation where c-Myc is sequestered at the nucleoli. Sequestration of c-Myc is accompanied by recruitment of proteasomes and may lead to subsequent degradation.

  • 18.
    Arabi, Azadeh
    et al.
    Södertörn University, School of Life Sciences. Karolinska Institutet.
    Wu, Siqin
    SLU.
    Ridderstråle, Karin
    SLU.
    Bierhoff, Holger
    German Cancer Research Center, Heidelberg, Germany.
    Shiue, Chiounan
    Södertörn University, School of Life Sciences. Karolinska Institutet.
    Fatyol, Karoly
    German Cancer Research Center, Heidelberg, Germany.
    Fahlén, Sara
    SLU.
    Hydbring, Per
    SLU.
    Söderberg, Ola
    Uppsala universitet.
    Grummt, Ingrid
    German Cancer Research Center, Heidelberg, Germany.
    Larsson, Lars-Gunnar
    SLU.
    Wright, Anthony P H
    Södertörn University, School of Life Sciences. Karolinska Institutet.
    c-Myc associates with ribosomal DNA and activates RNA polymerase I transcription2005In: Nature Cell Biology, ISSN 1465-7392, E-ISSN 1476-4679, Vol. 7, no 3, p. 303-310Article in journal (Refereed)
    Abstract [en]

    The c-Myc oncoprotein regulates transcription of genes that are associated with cell growth, proliferation and apoptosis(1). c-Myc levels are modulated by ubiquitin/proteasome-mediated degradation(1). Proteasome inhibition leads to c-Myc accumulation within nucleoli(2), indicating that c-Myc might have a nucleolar function. Here we show that the proteins c-Myc and Max interact in nucleoli and are associated with ribosomal DNA. This association is increased upon activation of quiescent cells and is followed by recruitment of the Myc cofactor TRRAP, enhanced histone acetylation, recruitment of RNA polymerase I (Pol I), and activation of rDNA transcription. Using small interfering RNAs (siRNAs) against c-Myc and an inhibitor of Myc - Max interactions, we demonstrate that c-Myc is required for activating rDNA transcription in response to mitogenic signals. Furthermore, using the ligand-activated MycER ( ER, oestrogen receptor) system, we show that c-Myc can activate Pol I transcription in the absence of Pol II transcription. These results suggest that c-Myc coordinates the activity of all three nuclear RNA polymerases, and thereby plays a key role in regulating ribosome biogenesis and cell growth.

  • 19. Archer, Amena
    et al.
    Lauter, Gilbert
    Södertörn University, School of Life Sciences.
    Hauptmann, Giselbert
    Södertörn University, School of Life Sciences.
    Mode, Agneta
    Gustafsson, Jan-Ake
    Transcriptional activity and developmental expression of liver X receptor (lxr) in zebrafish2008In: Developmental Dynamics, ISSN 1058-8388, E-ISSN 1097-0177, Vol. 237, no 4, p. 1090-1098Article in journal (Refereed)
    Abstract [en]

    Mammalian liver-X-receptors (LXRs) are transcription factors activated by oxysterols. They play an essential role in lipid and glucose metabolism. We have cloned the open reading frame of zebrafish lxr and describe its genomic organization. Zebrafish lxr encodes a 50-kDa protein with high sequence similarity to mammalian LXR alpha. In transfection assays, the encoded protein showed transcriptional activity in response to LXR-ligands. Treatment of adult zebrafish with the synthetic LXR ligand, GW3965, induced expression of genes involved in hepatic cholesterol and lipid pathways. Using qPCR and in situ hybridization, we found ubiquitous expression of lxr mRNA during the first 24 hr of development, followed by more restricted expression, particularly to the liver at 3dpf and the liver and intestine at 4dpf. In adult fish, all examined organs expressed lxr. In addition to a metabolic role of lxr, the temporal expression pattern suggests a developmental role in, e.g., the liver and CNS.

  • 20.
    Archer, Amena
    et al.
    BioNut, Karolinska Institutet.
    Srinivas Kitambi, Satish
    BioNut, Karolinska Institutet.
    Hallgren, Stefan
    Södertörn University, School of Life Sciences, Environmental science. Södertörn University, School of Life Sciences, Biology.
    Pedrelli, Matteo
    BioNut, Laboratoriemedicin, Karolinska Institutet.
    Olsén, Håkan
    Södertörn University, School of Life Sciences, Environmental science. Södertörn University, School of Life Sciences, Biology.
    Mode, Agneta
    BioNut, Karolinska Institutet.
    Gustafsson, Jan-Åke
    BioNut, Karolinska Institutet; Dept. Biology and Biochemistry, University of Houston.
    The Liver X-Receptor (Lxr) Governs Lipid Homeostasis in Zebrafish during Development2012In: Open journal of endocrine and metabolic diseases, ISSN 2165-7424, Vol. 2, no 4, p. 74-81Article in journal (Refereed)
    Abstract [en]

    The liver-X-receptors (LXRs) act as cholesterol sensors and participate in the regulation of lipid and cholesterol metabolism. The objective of this study was to determine the role of LXR during development using the zebrafish model. By in situ hybridization we showed distinct expression of lxr in the brain and the retina in the developing and adult zebrafish. Lxr ligand activation affected the expression of genes involved in lipid metabolism in zebrafish adult brain and eye as well as in zebrafish embryos. Morpholino knock down of lxr resulted in an overall impaired lipid deposition as determined by oil red O staining particularly in the head and around the eyes, and to significantly elevated levels of both total and free cholesterol in the yolk of lxr morphant embryos. The expression of genes involved in lipid and cholesterol metabolism was also changed in the lxr morphants. Furthermore, alcian blue staining revealed malformation of the pharyngeal skeleton in the lxr morphant. Our data show that Lxr is an important component of the regulatory network governing the lipid homeostasis during zebrafish development, which in turn may support a role of Lxr for normal development of the central nervous sytem, including the retina.

  • 21. Arnqvist, L
    et al.
    Dutta, P C
    Jonsson, Lisbeth
    Södertörn University, Avdelning Naturvetenskap.
    Sitbon, F
    Reduction of cholesterol and glycoalkaloid levels in transgenic potato plants by overexpression of a type 1 sterol methyltransferase cDNA2003In: Plant Physiology, ISSN 0032-0889, E-ISSN 1532-2548, Vol. 131, no 4, p. 1792-1799Article in journal (Refereed)
    Abstract [en]

    Transgenic potato (Solanum tuberosum cv Desiree) plants overexpressing a soybean (Glycine max) type 1 sterol methyltransferase (GmSMT1) cDNA were generated and used to study sterol biosynthesis in relation to the production of toxic glycoalkaloids. Transgenic plants displayed an increased total sterol level in both leaves and tubers, mainly due to increased levels of the 24-ethyl sterols isofucosterol and sitosterol. The higher total sterol level was due to increases in both free and esterified sterols. However, the level of free cholesterol, a nonalkylated sterol, was decreased. Associated with this was a decreased glycoalkaloid level in leaves and tubers, down to 41% and 63% of wild-type levels, respectively. The results show that glycoalkaloid biosynthesis can be down-regulated in transgenic potato plants by reducing the content of free nonalkylated sterols, and they support the view of cholesterol as a precursor in glycoalkaloid biosynthesis.

  • 22. Arnqvist, Lisa
    et al.
    Persson, Mattias
    Jonsson, Lisbeth
    Södertörn University, School of Life Sciences.
    Dutta, Paresh C.
    Sitbon, Folke
    Overexpression of CYP710A1 and CYP710A4 in transgenic Arabidopsis plants increases the level of stigmasterol at the expense of sitosterol2008In: Planta, ISSN 0032-0935, E-ISSN 1432-2048, Vol. 227, no 2, p. 309-317Article in journal (Refereed)
    Abstract [en]

    Sitosterol and stigmasterol are major sterols in vascular plants. An altered stigmasterol:sitosterol ratio has been proposed to influence the properties of cell membranes, particularly in relation to various stresses, but biosynthesis of stigmasterol is poorly understood. Recently, however, Morikawa et al. (Plant Cell 18:1008-1022, 2006) showed in Arabidopsis thaliana that synthesis of stigmasterol and brassicasterol is catalyzed by two separate sterol C-22 desaturases, encoded by the genes CYP710A1 and CYP710A2, respectively. The proteins belong to a small cytochrome P450 subfamily having four members, denoted by CYP710A1-A4, and are related to the yeast sterol C-22 desaturase Erg5p acting in ergosterol synthesis. Here, we report on our parallel investigation of the Arabidopsis CYP710A family. To elucidate the function of CYP710A proteins, transgenic Arabidopsis plants were generated overexpressing CYP710A1 and CYP710A4. Compared to wild-type plants, both types of transformant displayed a normal phenotype, but contained increased levels of free stigmasterol and a concomitant decrease in the level of free sitosterol. CYP710A1 transformants also displayed higher levels of esterified forms of stigmasterol, cholesterol, 24-methylcholesterol and isofucosterol. The results confirm the findings of Morikawa et al. (Plant Cell 18:1008-1022, 2006) regarding the function of CYP710A1 in stigmasterol synthesis, and show that CYP710A4 also has this capacity. Furthermore, our results suggest that an increased stigmasterol level alone is sufficient to stimulate esterification of other major sterols.

  • 23. Arteaga, H J
    et al.
    Hinkula, J
    van Dijk-Hard, I
    Dilber, M S
    Wahren, B
    Christensson, B
    Mohamed, Abdalla J
    Smith, C I E
    Choosing CCR5 or Rev siRNA in HIV-12003In: Nature Biotechnology, ISSN 1087-0156, E-ISSN 1546-1696, Vol. 21, no 3, p. 230-231Article in journal (Refereed)
  • 24.
    Articus, Kristina
    et al.
    Belgian Biodiversity Platform, Université Libre de Bruxelles.
    Wedin, Mats
    Naturhistoriska riksmuseet, Stockholm.
    Mattsson, Jan-Eric
    Södertörn University, Avdelning Naturvetenskap, Biology.
    Tibell, Leif
    Uppsala universitet.
    Grube, Martin
    Institute of plan sciences, University of Graz, Austria.
    Phylogenetic studies in Usnea.2000In: The Fourth IAL Symposium Progress and Problems in Lichenology at the Turn of the Millenium, Abstracts: 100. Barcelona., 2000Conference paper (Other academic)
  • 25.
    Arup, Ulf
    et al.
    University of Lund.
    Ekman, Stefan
    University of Lund.
    Lindblom, Louise
    University of Lund.
    Mattsson, Jan-Eric
    University of Lund.
    High performance thin layer chromatography (HPTLC), an improved technique for screening lichen substances.1993In: The Lichenologist, ISSN 0024-2829, E-ISSN 1096-1135, Vol. 25, p. 61-71Article in journal (Refereed)
  • 26.
    Asghar, Naveed
    et al.
    Södertörn University, School of Natural Sciences, Technology and Environmental Studies, Biology. Örebro universitet.
    Lee, Yi-Ping
    Umeå universitet.
    Nilsson, Emma
    Umeå universitet.
    Lindqvist, Rickard
    Umeå universitet.
    Melik, Wessam
    Örebro universitet.
    Kröger, Andrea
    6Helmholtz Centre for Infection Research, Braunschweig, Germany / University of Magdeburg, Magdenbrug, Germany.
    Överby, Anna K.
    Umeå universitet.
    Johansson, Magnus
    Örebro universitet.
    The role of the poly(A) tract in the replication and virulence of tick-borne encephalitis virus2016In: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, no 6, article id 39265Article in journal (Refereed)
    Abstract [en]

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

  • 27.
    Asghar, Naveed
    et al.
    Södertörn University, School of Natural Sciences, Technology and Environmental Studies, Biology.
    Lindblom, Pontus
    Melik, Wessam
    Södertörn University, School of Natural Sciences, Technology and Environmental Studies, Biology.
    Lindqvist, Richard
    Haglund, Mats
    Forsberg, Pia
    Overby, Anna K
    Andreassen, Ashild
    Lindgren, Per-Eric
    Johansson, Magnus
    Södertörn University, School of Natural Sciences, Technology and Environmental Studies, Biology. Örebro University.
    Tick-borne encephalitis virus sequenced directly from questing and blood-feeding ticks reveals quasispecies variance.2014In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 9, no 7, article id e103264Article in journal (Refereed)
    Abstract [en]

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

  • 28.
    Asghar, Naveed
    et al.
    Södertörn University, School of Natural Sciences, Technology and Environmental Studies, Biology. Örebro universitet.
    Petersson, Mona
    Södertörn University, School of Natural Sciences, Technology and Environmental Studies, Geography.
    Johansson, Magnus
    Södertörn University, School of Natural Sciences, Technology and Environmental Studies, Biology. Örebro univarsitet.
    Dinnétz, Patrik
    Södertörn University, School of Natural Sciences, Technology and Environmental Studies, Environmental Science.
    Local land-scape effects on population dynamics of Ixodes ricinus2016In: Geospatial Health, ISSN 1827-1987, Vol. 11, p. 283-289, article id 487Article in journal (Refereed)
  • 29.
    Asghar, Naveed
    et al.
    Södertörn University, School of Natural Sciences, Technology and Environmental Studies, Biology. Örebro universitet.
    Pettersson, John H-O
    Norwegian Institute of Public Health, Oslo, Norway / Statens veterinärmedicinska anstalt.
    Dinnétz, Patrik
    Södertörn University, School of Natural Sciences, Technology and Environmental Studies, Environmental Science.
    Andreassen, Åshild
    Norwegian Institute of Public Health, Oslo, Norway.
    Johansson, Magnus
    Örebro universitet.
    Deep sequencing analysis of tick-borne encephalitis virus from questing ticks at natural foci reveals similarities between quasispecies pools of the virus2017In: Journal of General Virology, ISSN 0022-1317, E-ISSN 1465-2099, Vol. 98, no 3, p. 413-421Article in journal (Refereed)
    Abstract [en]

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

  • 30.
    Askarian Nameghi, Shahnaz
    Södertörn University College, School of Life Sciences.
    Genotyping Escherichia coli isolates by Pulsed-Field Gel Electrophoresis2007Independent thesis Advanced level (degree of Magister), 20 points / 30 hpStudent thesis
    Abstract [en]

    Transmission of bacterial strains between patients is a serious problem in hospitals and with the increasing rate of antibiotic resistance the problem has farther escalated. Enterobacteriaceae produced extended-spectrum beta-lactamses (ESBLs), especially Escherichia coli (E-coli), are increasingly important nosocomial pathogens (7, 8). These bacteria are often multiple resistant and are responsible for many intestinal infections and urinary tract infections (2, 5). With the more frequent use of invasive devices in hospital care, these types of nosocomial infections have increased, particularly in seriously ill patients.

    In order to diminish transmission of bacterial strains between patients and to study the epidemiology of these bacteria, it is of great importance to develop rapid and specific methods to be able to subtype on strain-level, i.e. to create a fingerprint of the isolates. The method may be based on phenotypic or genotypic characteristics of the microorganism. Any typing method must have high reproducibility and discrimination power to differentiate unrelated strains and also to demonstrate relationship of organisms deriving from the same source. In the present project, a Pulsed-Field Gel Electrophoresis (PFGE) assay for genotyping clinical E. coli isolates was used. PFGE can be used as a genotyping tool and is widely used to type bacteria and trace nosocomial infection. However, the method is time-consuming and relatively expensive in compare with other methods like PCR. In this study, a total of 93 strains were collected. The study was aimed to investigate the genotypes of the collected isolates and to identify and potential the outbreak strains.

    The isolates investigated were genotypically diverse shown by a variety of PFGE banding patterns. However, clusters of closely related isolates involved in outbreaks were also identified.

    In conclusion, when analyzing a large number of strains, a combination of a rapid phenotyping or genotyping method and a powerful genotyping method like PFGE would be an appropriate strategy for studying clonal relationship among isolates e.g. for detecting cross-transmission of nosocomial pathogens.

  • 31.
    Asnicar, Davide
    et al.
    University of Padova, Italy; University of Gothenburg, Sweden.
    Ašmonaitė, Giedrė
    University of Gothenburg, Sweden.
    Birgersson, Lina
    University of Gothenburg, Sweden.
    Kvarnemo, Charlotta
    University of Gothenburg, Sweden.
    Svensson, Ola
    Södertörn University, School of Natural Sciences, Technology and Environmental Studies, Mathematics Teaching. University of Gothenburg, Sweden.
    Sturve, Joachim
    University of Gothenburg, Sweden.
    Sand Goby: An Ecologically Relevant Species for Behavioural Ecotoxicology2018In: Fishes, E-ISSN 2410-3888, Vol. 3, no 1, article id 13Article in journal (Refereed)
    Abstract [en]

    Locomotion-based behavioural endpoints have been suggested as suitable sublethal endpoints for human and environmental hazard assessment, as well as for biomonitoring applications. Larval stages of the sand goby (Pomatoschistus minutus) possess a number of attractive qualities for experimental testing that make it a promising species in behavioural ecotoxicology. Here, we present a study aimed at developing a toolkit for using the sand goby as novel species for ecotoxicological studies and using locomotion as an alternative endpoint in toxicity testing. Exposure to three contaminants (copper (Cu), di-butyl phthalate (DBP) and perfluorooctanoic acid (PFOA) was tested in the early life stages of the sand goby and the locomotion patterns of the larvae were quantified using an automatic tracking system. In a photo-motor test, sand goby larvae displayed substantially higher activity in light than in dark cycles. Furthermore, all tested compounds exerted behavioural alterations, such as hypo- and hyperactivity. Our experimental results show that sand goby larvae produce robust and quantifiable locomotive responses, which could be used within an ecotoxicological context for assessing the behavioural toxicity of environmental pollutants, with particular relevance in the Nordic region. This study thus suggests that sand goby larvae have potential as an environmentally relevant species for behavioural ecotoxicology, and as such offer an alternative to standard model species.

  • 32.
    Awebro, Kenneth
    Södertörn University College, School of Gender, Culture and History, History.
    Valfångst2007In: Människan och faunan / [ed] Håkan Tunón, Mattias Iwarsson, Stephen Manktelow, Stockholm: Wahlström & Widstrand , 2007Chapter in book (Other academic)
  • 33. Backhed, F
    et al.
    Normark, S
    Schweda, Elke K H
    Södertörn University, Avdelning Naturvetenskap.
    Oscarson, S
    Richter-Dahlfors, A
    Structural requirements for TLR4-mediated LPS signalling: a biological role for LPS modifications2003In: Microbes and infection, ISSN 1286-4579, E-ISSN 1769-714X, Vol. 5, no 12, p. 1057-1063Article in journal (Refereed)
    Abstract [en]

    Cells of the mucosal lining are the first to encounter invading bacteria during infection, and as such, they have developed numerous ways of detecting microbial intruders. Recently, we showed that epithelial cells recognize lipopolysaccharide (LPS) through the CD14-Toll-like receptor (TLR)-4 complex. Here, we identify the substructures of LPS that are recognized by the TLR4 receptor complex. In contrast to lipid A, the O-antigen does not mediate an inflammatory response; rather it interferes with the lipid A recognition. An Escherichia coli strain genetically modified to express penta-acylated lipid A not only showed reduced immunogenicity, but was also found to inhibit proinflammatory signalling induced by wild-type E. coli (hexa-acylated lipid A) as well as LPS from other bacteria of the Enterobacteriaceae family. Furthermore, penta-acylated LPS from Pseudomonas aeruginosa acted as an antagonist to hexa-acylated E. coli LPS, as did E. coli, as shown by its inhibitory effect on IL-8 production in stimulated cells. Hypo-acylated lipidA, such as that of P. aeruginosa, is found in several species within the gut microflora as well as in several bacteria causing chronic infections. Thus, our results suggest that the composition of the microflora may be important in modulating pro-inflammatory signalling in epithelial cells under normal as well as pathologic conditions.

  • 34.
    Backman, Agneta
    et al.
    Södertörn University, School of Chemistry, Biology, Geography and Environmental Science. Karolinska Institutet.
    Jansson, Janet K
    Södertörn University, School of Chemistry, Biology, Geography and Environmental Science. SLU.
    Degradation of 4-chlorophenol at low temperature and during extreme temperature fluctuations by Arthrobacter chlorophenolicus A62004In: Microbial Ecology, ISSN 0095-3628, E-ISSN 1432-184X, Vol. 48, no 2, p. 246-253Article in journal (Refereed)
    Abstract [en]

    Low average temperatures and temperature fluctuations in temperate soils challenge the efficacy of microbial strains used for clean up of pollutants. In this study, we investigated the cold tolerance of Arthrobacter chlorophenolicus A6, a microorganism previously shown to degrade high concentrations of 4-chlorophenol at 28degreesC. Luciferase activity from a luc-tagged derivative of the strain (A6L) was used to monitor the metabolic status of the population during 4-chlorophenol degradation. The A6L strain could degrade 200-300 mug mL(-1) 4-chlorophenol in pure cultures incubated at 5degreesC, although rates of degradation, growth and the metabolic status of the cells were lower at 5degreesC compared to 28degreesC. When subjected to temperature fluctuations between 5 and 28degreesC, A6L continued to degrade 4-chlorophenol and remained active. In soil microcosm experiments, the degradation rates were significantly faster the first week at 28degreesC, compared to 5degreesC. However, this difference was no longer seen after 7 days, and equally low 4-chlorophenol concentrations were reached after 17 days at both temperatures. During 4-chlorophenol degradation in soil, CFU and luciferase activity values remained constant at both 5 and 28degreesC. However, once most of the 4-chlorophenol was degraded, both values decreased by 1-1.5 logarithmic values at 28degreesC, whereas they remained constant at 5degreesC, indicating a high survival of the cells at low temperatures. Because of the ability of A. chlorophenolicus A6 to degrade high concentrations of 4-chlorophenol at 5degreesC, together with its tolerance to temperature fluctuations and stress conditions found in soil, this strain is a promising candidate for bioaugmentation of chlorophenol-contaminated soil in temperate climates.

  • 35.
    Backman, Agneta
    et al.
    Södertörn University, School of Chemistry, Biology, Geography and Environmental Science. Karolinska Institutet.
    Maraha, Ninwe
    Södertörn University, School of Chemistry, Biology, Geography and Environmental Science. Karolinska Institutet.
    Jansson, Janet K
    Södertörn University, School of Chemistry, Biology, Geography and Environmental Science. SLU.
    Impact of temperature on the physiological status of a potential bioremediation inoculant, Arthrobacter chlorophenolicus A62004In: Applied and Environmental Microbiology, ISSN 0099-2240, E-ISSN 1098-5336, Vol. 70, no 5, p. 2952-2958Article in journal (Refereed)
    Abstract [en]

    Arthrobacter chlorophenolicus A6 (A6) can degrade large amounts of 4-chlorophenol in soil at 5 and 28degreesC. In this study, we investigated the effects of temperature on the physiological status of this bacterium in pure culture and in soil. A derivative of A6 tagged with the gfp gene (encoding green fluorescent protein [GFP]) was used to specifically quantify A6 cells in soil. In addition, cyano-ditolyl-tetrazoliumchloride was used to stain GFP-fluorescent cells with an active electron transfer system ("viable cellis") whereas propidium iodide (PI) was used to stain cells with damaged membranes ("dead cells"). Another derivative of the strain (tagged with the firefly luciferase gene [luc]) was used to monitor the metabolic activity of the cell population, since the bioluminescence phenotype is dependent on cellular energy reserves. When the cells were incubated in soil at 28degreesC, the majority were stained with PI, indicating that they had lost their cell integrity. In addition, there was a corresponding decline in metabolic activity and in the ability to be grown in cultures on agar plates after incubation in soil at 28degreesC, indicating that the cells were dying under those conditions. When the cells were incubated in soil at 5degreesC, by contrast, the majority of the cells remained intact and a large fraction of the population remained metabolically active. A similar trend towards better cell survival at lower temperatures was found in pure-culture experiments. These results make A. chlorophenolicus A6 a good candidate for the treatment of chlorophenol-contaminated soil in cold climates.

  • 36. Bakhiet, M
    et al.
    Tjernlund, A
    Mousa, A
    Gad, Annica
    Södertörn University, Avdelning Naturvetenskap. Karolinska Institute.
    Strömblad, Staffan
    Södertörn University, Avdelning Naturvetenskap. Karolinska Institute.
    Kuziel, W A
    Seiger, A
    Andersson, J
    RANTES promotes growth and survival of human first-trimester forebrain astrocytes2001In: Nature Cell Biology, ISSN 1465-7392, E-ISSN 1476-4679, Vol. 3, no 2, p. 150-157Article in journal (Refereed)
    Abstract [en]

    We have examined the role of alpha and beta chemokines in the promotion of the ontogenetic development of the brain. RANTES was expressed preferentially in human fetal astrocytes in an age-dependent manner. Astrocytes from 5-week-old brains showed high proliferation and reduced survival, whereas 10-week-old astrocytes exhibited opposite effects. These effects were suppressed by anti-RANTES or anti-RANTES receptor antibodies and were enhanced by recombinant RANTES. RANTES induced tyrosine phosphorylation of several cellular proteins and nuclear translocation of STAT-1 in astrocytes. Interferons (IFN-gamma) was required for RANTES effects because RANTES induced IFN-gamma, and only 10-week-old astrocytes expressed the IFN-gamma receptor. Blocking of IFN-gamma with antibody reversed the effects of RANTES, indicating that cytokine/chemokine networks are critically involved in brain development.

  • 37. Balogun, H. A.
    et al.
    Vasconcelos, N. -M
    Lindberg, Robert
    Södertörn University, School of Life Sciences.
    Haeggström, M.
    Moll, K.
    Chen, Q.
    Wahlgren, M.
    Berzins, K.
    Immunogenicity and antigenic properties of Pf332-C231, a fragment of a non-repeat region of the Plasmodium falciparum antigen Pf3322009In: Vaccine, ISSN 0264-410X, E-ISSN 1873-2518, Vol. 28, no 1, p. 90-97Article in journal (Refereed)
    Abstract [en]

    Antigen Pf332, a megadalton protein has been shown to be associated with the membrane of infected erythrocytes Detailed functional studies on the antigen have remained hampered by the cross-reactive nature of antibodies generated to Pf332 PB32-C231, identified in the C-terminal region of Pf332 was cloned and antibodies against the C231 fragment were shown to react with intact Pf332 antigen by both immunofluorescence and immunoblotting analyses Antibodies to C231 inhibited in vitro Plasmodium falciparum growth efficiently In addition. human sera from malaria-exposed individuals reacted with recombinant C231 We show that Pf332-C231 represents a functional domain and is expected to facilitate further studies on Pf332 as a potential target for protective immune responses and the function of the antigen.

  • 38. Ban, L.
    et al.
    Didon, Andrea
    Södertörn University, School of Life Sciences.
    Jonsson, L. M. V.
    Glinwood, R.
    Delp, Gabriele
    Södertörn University, School of Life Sciences.
    An improved detection method for the Rhopalosiphum padi virus (RhPV) allows monitoring of its presence in aphids and movement within plants2007In: Journal of Virological Methods, ISSN 0166-0934, E-ISSN 1879-0984, Vol. 142, no 1-2, p. 136-142Article in journal (Refereed)
    Abstract [en]

    Rhopalosiphumpadi virus (RhPV) is an insect RNA virus that infects aphids, reducing their lifespan and fecundity. It can be transmitted vertically between aphids and horizontally via the plant. An improved detection method for the virus in aphids and plants using RT-PCR was developed; this allowed individual aphids to be tested for RhPV. Testing of R. padi aphids collected from different sites in Sweden revealed the presence of RhPV in wild aphid populations for the first time in Europe. Virus could be detected in several life stages of R. padi, including sexual individuals and eggs, establishing an over-wintering route for the virus. Using RT-PCR, systemic transport of the virus in plants was tracked. Virus spread from the aphid feeding site to all parts of the plant, including roots, within 7 days, and could be acquired by virus-free aphids feeding on the same plant.

  • 39. Ban, Liping
    et al.
    Ahmed, Elham
    Ninkovic, Velemir
    Delp, Gabriele
    Södertörn University, School of Life Sciences.
    Glinwood, Robert
    Infection with an insect virus affects olfactory behaviour and interactions with host plant and natural enemies in an aphid2008In: Entomologia Experimentalis et Applicata, ISSN 0013-8703, E-ISSN 1570-7458, Vol. 127, no 2, p. 108-117Article in journal (Refereed)
    Abstract [en]

    Aphid ecology and population dynamics are affected by a series of factors including behavioural responses to ecologically relevant chemical cues, capacity for population growth, and interactions with host plants and natural enemies. Using the aphid Rhopalosiphum padi (L.) (Homoptera: Aphididae), we showed that these factors were affected by infection with Rhopalosiphum padi virus (RhPV). Uninfected aphids were attracted to odour of uninfected aphids on the host plant, an aggregation mechanism. However, infected aphids were not attracted, and neither infected nor uninfected aphids were attracted to infected aphids on the plant. Infected aphids did not respond to methyl salicylate, a cue denoting host suitability. Infected aphids were more behaviourally sensitive to aphid alarm pheromone, and left the host plant more readily in response to it. RhPV reduced the lifespan and population growth rate of the aphid. The predacious ladybird, Coccinella septempunctata (L.) (Coleoptera: Coccinellidae), consumed more infected aphids than uninfected aphids in a 24-h period, and the aphid parasitoid Aphidius ervi Haliday (Hymenoptera: Aphidiidae) attacked more infected than uninfected aphids. However, the proportion of mummies formed was lower with infected aphids. The results represent further evidence that associated organisms can affect the behaviour and ecology of their aphid hosts.

  • 40. Bao, W J
    et al.
    Thullberg, M
    Zhang, H Q
    Onischenko, A
    Strömblad, Staffan
    Södertörn University, Avdelning Naturvetenskap.
    Cell attachment to the extracellular matrix induces proteasomal degradation of p21(CIP1) via Cdc42/Rac1 signaling2002In: Molecular and Cellular Biology, ISSN 0270-7306, E-ISSN 1098-5549, Vol. 22, no 13, p. 4587-4597Article in journal (Refereed)
    Abstract [en]

    The cyclin-dependent kinase 2 (Cdk2) inhibitors p21(CIP1) and p27(KIP1) are negatively regulated by anchorage during cell proliferation, but it is unclear how integrin signaling may affect these Cdk2 inhibitors. Here, we demonstrate that integrin ligation led to rapid reduction of p21(CIp1) and p27(KIP1) protein levels in three distinct cell types upon attachment to various extracellular matrix (ECM) proteins, including fibronectin (FN), or to immobilized agonistic anti-integrin monoclonal antibodies. Cell attachment to FN did not rapidly influence p21(CIp1) mRNA levels, while the protein stability of p21(CIp1) was decreased. Importantly, the down-regulation of p21(CIP1) and p27(KIP1) was completely blocked by three distinct proteasome inhibitors, demonstrating that integrin ligation induced proteasomal degradation of these Cdk2 inhibitors. Interestingly, ECM-induced proteasomal proteolysis of a ubiquitination-deficient p21(CIP1) mutant (p21K6R) also occurred, showing that the proteasomal degradation of p21(CIP1) was ubiquitin independent. Concomitant with our finding that the small GTPases Cdc42 and Rac1 were activated by attachment to FN, constitutively active (ca) Cdc42 and ca Rac1 promoted down-regulation of p21(CIP1). However, dominant negative (dn) Cdc42 and do Rac1 mutants blocked the anchorage-induced degradation of p21(CIP1), suggesting that an integrin-induced Cdc42/Rac1 signaling pathway activates proteasomal degradation of p21(CIP1). Our results indicate that integrin-regulated proteasomal proteolysis might contribute to anchorage-dependent cell cycle control.

  • 41.
    Bartish, Galyna
    Södertörn University, School of Life Sciences. Stockholms universitet, Wenner-Grens institut.
    Elongation factor 2: A key component of the translation machinery in eukaryotes: Properties of yeast elongation factor 2 studied in vivo2008Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Synthesis of proteins is performed by the ribosome, a large ribonucleoprotein complex. Apart from the ribosome, numerous protein factors participate in this process. Elongation factor 2 (eEF2) is one of these factors. eEF2 is an essential protein with a mol. mass of about 100 kDa. The amino acid sequence of eEF2 is highly conserved in different organisms. eEF2 from S. cerevisiae contains 842 amino acids. The role of eEF2 in protein synthesis is to participate in the translocation of tRNAs from the A- and P-sites on the ribosome to the P- and E-sites. This movement of tRNAs is accompanied by a simultaneous movement of mRNA by one codon. eEF2 consists of six domains referred to as domains G, G′ and II-V, belongs to the G-protein super-family and possesses all structural motifs characterizing proteins in this family. eEF2 binds to the ribosome in complex with GTP. After GTP hydrolysis and translocation, it leaves the ribosome bound to GDP. The rate of protein synthesis in the cell can be regulated by phosphorylation of eEF2. Phosphorylation occurs on two threonine residues, situated in the G domain of the factor. Phosphorylation of eEF2 is catalysed by Rck2-kinase in yeast which is activated in response to osmotic stress. Despite the high degree of conservation of the threonine residues, they are not essential for yeast cell under normal growth conditions. However, under mild osmotic stress the growth rate of the cells lacking threonine residues was decreased. Region where threonine residues are located, called Switch I. Cryo-EM reconstruction shows that this region adopts ordered conformation when the eEF2•GTP complex is bound to the ribosome but became structurally disordered upon GTP hydrolysis. Mutagenesis of individual amino acids in Switch I resulted in both functional and non-functional eEF2 depending on the site of mutation and the substituting amino acid. Both functional and non-functional Switch I mutants were able to bind to the ribosome, indicating that mutations did not abolish the capacity of the factor to bind GTP. Yeast eEF2 with Switch I region from E. coli was able to substitute the wild type protein in vivo, though the growth rate of these cells was severely impaired. The eEF2-dependent GTP hydrolysis can be activated by ribosome from heterologous sources as seen in vitro. However, eEF2 from A. thaliana, D. melanogaster and S. solfataricus could not substi-tute yeast eEF2 in vivo. This may indicate additional roles of eEF2 in the yeast cell, apart from translocation itself.

  • 42.
    Bartish, Galyna
    et al.
    Södertörn University, School of Life Sciences. Stockholm University.
    Moradi, Hossein
    Södertörn University, School of Life Sciences. Stockholm University.
    Nygård, Odd
    Södertörn University, School of Life Sciences.
    Amino acids Thr56 and Thr58 are not essential for elongation factor 2 function in yeast2007In: The FEBS Journal, ISSN 1742-464X, E-ISSN 1742-4658, Vol. 274, no 20, p. 5285-5297Article in journal (Refereed)
    Abstract [en]

    Yeast elongation factor 2 is an essential protein that contains two highly conserved threonine residues, T56 and T58, that could potentially be phosphorylated by the Rck2 kinase in response to environmental stress. The importance of residues T56 and T58 for elongation factor 2 function in yeast was studied using site directed mutagenesis and functional complementation. Mutations T56D, T56G, T56K, T56N and T56V resulted in nonfunctional elongation factor 2 whereas mutated factor carrying point mutations T56M, T56C, T56S, T58S and T58V was functional. Expression of mutants T56C, T56S and T58S was associated with reduced growth rate. The double mutants T56M/T58W and T56M/T58V were also functional but the latter mutant caused increased cell death and considerably reduced growth rate. The results suggest that the physiological role of T56 and T58 as phosphorylation targets is of little importance in yeast under standard growth conditions. Yeast cells expressing mutants T56C and T56S were less able to cope with environmental stress induced by increased growth temperatures. Similarly, cells expressing mutants T56M and T56M/T58W were less capable of adapting to increased osmolarity whereas cells expressing mutant T58V behaved normally. All mutants tested were retained their ability to bind to ribosomes in vivo. However, mutants T56D, T56G and T56K were under-represented on the ribosome, suggesting that these nonfunctional forms of elongation factor 2 were less capable of competing with wild-type elongation factor 2 in ribosome binding. The presence of nonfunctional but ribosome binding forms of elongation factor 2 did not affect the growth rate of yeast cells also expressing wild-type elongation factor 2.

  • 43.
    Bartish, Galyna
    et al.
    Södertörn University, School of Life Sciences. Stockholm University.
    Nygård, Odd
    Södertörn University, School of Life Sciences.
    Importance of individual amino acids in the Switch I region in eEF2 studied by functional complementation in S. cerevisiae2008In: Biochimie, ISSN 0300-9084, E-ISSN 1638-6183, Vol. 90, no 5, p. 736-748Article in journal (Refereed)
    Abstract [en]

    Elongation factor 2 (eEF2) is a member of the G-protein super family. G-proteins undergo conformational changes associated with binding of the guanosine nucleotide and hydrolysis of the bound GTP. These structural rearrangements affects the Switch I region (also known as the Effector loop). We have studied the role of individual amino acids in the Switch I region (amino acids 25-73) of S. cerevisiae eEF2 using functional complementation in yeast. 21 point mutations in the Switch I region were created by site-directed mutagenesis. Mutants K49R, E52Q, A53G, F55Y, K60R, Q63A, T68S, 169M and A73G were functional while mutants R54H, F55N, D57A, D57E, D57S, R59K, R59M, Q63E, R65A, R65N, T68A and T68M were inactive. Expression of mutants K49R, A53G, Q63A, 169M and A73G was associated with markedly decreased growth rates and yeast cells expressing mutants A53G and 169M became temperature sensitive. The functional capacity of eEF2 in which the major part Switch I (amino acids T56 to 169) was converted into the homologous sequence found in EF-G from E. coli was also studied. This protein chimera could functionally replace yeast eEF2 in vivo. Yeast cells expressing this mutant grew extremely slowly, showed increased cell death and became temperature sensitive. The ability of the mutant to replace authentic eEF2 in vivo indicates that the structural rearrangement of Switch I necessary for eEF2 function is similar in eukaryotes and bacteria. The effect of two point mutations in the P-loop was also studied. Mutant A25G but not A25V could functionally replace yeast eEF2 even if cells expressing the mutant grew slowly. The A25G mutation converted the consensus sequences AXXXXGK[T/S] in eEF2 to the corresponding motif GXXXXGK[T/S] found in all other G-proteins, suggesting that the alanine found in the P-loop of peptidyltranslocases are not essential for function.

  • 44.
    Bartish, Galyna
    et al.
    Stockholms universitet.
    Nygård, Odd
    Södertörn University, School of Life Sciences.
    The functional importance of the N- and C-terminal regions in elongation factor 2 from S. cerevisiaeManuscript (preprint) (Other academic)
  • 45. Bauer, S H J
    et al.
    Månsson, Martin
    Södertörn University, Avdelning Naturvetenskap.
    Hood, D W
    Richards, J C
    Moxon, E R
    Schweda, Elke K H
    Södertörn University, Avdelning Naturvetenskap.
    A rapid and sensitive procedure for determination of 5-N-acetyl neuraminic acid in lipopolysaccharides of Haemophilus influenzae: a survey of 24 non-typeable H-influenzae strains2001In: Carbohydrate Research, ISSN 0008-6215, E-ISSN 1873-426X, Vol. 335, no 4, p. 251-260Article in journal (Refereed)
    Abstract [en]

    In view of the importance of 5-N-acetyl neuraminic acid in bacterial pathogenesis, a sensitive, reproducible and reliable method for the determination of 5-N-acetyl neuraminic acid levels in lipopolysaccharide (LPS) is described and applied to 24 different non-typeable Haemophilus influenzae (NTHi) strains. The method involves analysis by high-performance anion-exchange chromatography with pulsed amperometric detection (HPAEC-PAD) of terminal 5-N-acetyl neuraminic acid residues released by neuraminidase treatment of O-deacylated LPS. The procedure is relatively fast and the instrumental effort is moderate. The results of the procedure were compared with data obtained by H-1 NMR and electrospray ionisation-mass spectrometry (ESI-MS). The analysis of LPS from 24 NTHi strains showed that 5-N-acetyl neuraminic acid was found to be a common constituent of LPS in NTHi. Only one strain (NTHi 432) did not show any sialylation. Molar ratios (LPS /5-N-acetyl neuraminic acid) ranged between 5/1 and 500/1. Several strains in which no 5-N-acetyl neuraminic acid could be determined by other methods including 1H NMR and ESI-MS were shown to contain 5-N-acetyl neuraminic acid by this HPAEC-PAD procedure. The method was applied to determine levels of terminal 5-N-acetyl neuraminic acid in LPS from NTHi strains grown under different conditions and mutant strains containing inactive LPS biosynthetic genes.

  • 46. Beckman, M
    et al.
    Kihlmark, Madeleine
    Södertörn University, School of Chemistry, Biology, Geography and Environmental Science.
    Iverfeldt, K
    Hallberg, Einar
    Södertörn University, School of Chemistry, Biology, Geography and Environmental Science.
    Degradation of GFP-labelled POM121, a non-invasive sensor of nuclear apoptosis, precedes clustering of nuclear pores and externalisation of phosphatidylserine2004In: Apoptosis (London), ISSN 1360-8185, E-ISSN 1573-675X, Vol. 9, no 3, p. 363-368Article in journal (Refereed)
    Abstract [en]

    The nuclear pore membrane protein POM121 is specifically degraded during apoptosis by a caspase-3-dependent process enabling early detection of apoptosis in living cells expressing POM121-GFP. Here we further investigated temporal aspects of apoptotic degradation of POM121-GFP. We demonstrate that decreased POM121-GFP fluorescence precedes annexin V-labelling of apoptotic cells. This indicates that degradation of the nuclear pore complex starts prior to redistribution of plasma membrane phosphatidylserine, which serves as a signal for phagocytotic elimination of apoptotic cells. Furthermore, a caspase-resistant GFP-labelled mutant of POM121 resisted degradation even in late apoptosis and was detected in clustered nuclear pores. Thus, it can be concluded that loss of POM121-GFP is a specific sensor of the activation of caspase-3-dependent proteolysis at the nuclear pores.

  • 47.
    Beckman, Marie
    et al.
    Södertörn University, School of Life Sciences, Molecular biology.
    Freeman, Craig
    Parish, Christopher R.
    Small, David H.
    Activation of cathepsin D by glycosaminoglycans2009In: The FEBS Journal, ISSN 1742-464X, E-ISSN 1742-4658, Vol. 276, no 24, p. 7343-7352Article in journal (Refereed)
    Abstract [en]

    We have previously shown that heparin can increase the activity of the proenzyme form of Alzheimer's beta-site amyloid precursor protein cleaving enzyme 1 (BACE1). Cathepsin D (CD) is a member of the aspartic protease family and has sequence similarity to BACE1. Therefore, we examined whether heparin and other glycosaminoglycans (GAGs) can influence the activity of CD. Heparin and other GAGs were found to stimulate the activity of recombinant proCD. Desulfation of heparin almost abolished the stimulation, indicating that sulfate groups were important for the stimulatory effect. In addition, the stimulation was dependent on the length of the GAG chain, as larger GAGs were more potent in their ability to stimulate proCD than shorter fragments. In the presence of heparin, limited autocatalytic proteolysis of the proenzyme was increased, suggesting that heparin increases the activity of proCD by accelerating the conversion of proCD, which has little activity, to pseudoCD, an active form lacking residues 1-26 of the prodomain. Furthermore, the activity of spleen-derived mature CD, which lacks the entire 44 amino acid residue prodomain, was also increased by heparin, indicating that the catalytic domain of CD contains at least one region to which GAGs bind and stimulate enzyme activity. Because heparin also stimulated the activity of pseudoCD, proenzyme activation was probably accelerated by the interaction of heparin with the catalytic domain of pseudoCD. However, it is possible that heparin may also activate the proenzyme directly. On the basis of this study, we propose that GAGs may regulate CD activity in vivo.

  • 48. Belyaev, I Y
    et al.
    Eriksson, S
    Nygren, Jonas
    Södertörn University, Avdelning Naturvetenskap. Karolinska Institute / Stockholm University.
    Torudd, J
    Harms-Ringdahl, M
    Effects of ethidium bromide on DNA loop organisation in human lymphocytes measured by anomalous viscosity time dependence and single cell gel electrophoresis1999In: Biochimica et Biophysica Acta - General Subjects, ISSN 0304-4165, E-ISSN 1872-8006, Vol. 1428, no 2-3, p. 348-356Article in journal (Refereed)
    Abstract [en]

    The effects of ethidium bromide (EtBr) on human lymphocytes were studied by the method of anomalous viscosity time dependence (AVTD) and by the comet assay. EtBr at low concentrations increased the maximum viscosity and time of radial migration as measured with AVTD at neutral conditions of lysis. A pronounced relaxation of DNA loops was observed with the neutral comet assay. The maximal comet length corresponded to 2 Mb DNA loops. At high concentrations of EtBr, 2. mg/ml, significant reduction in AVTD below control level was seen that suggested hypercondensation of chromatin. The hypercondensation was directly observed with the neutral comet assay. EtBr did not induce DNA strand breaks as measured by the alkaline comet assay. The hypercondensed nuclei could be decondensed by irradiation with gamma-rays or exposure to light. The data provide evidence that EtBr at high concentrations resulted in hypercondensation of chromatin below control level. The comet assay confirmed that the increase in AVTD peaks deals with relaxation of loops and AVTD decrease is caused by chromatin condensation. The prediction of the AVTD theory for a correlation between time of radial migration and condensation of chromatin was verified. Further, the data show that the comet assay at neutral conditions of lysis is rather sensitive to DNA loop relaxation in the absence of DNA damage. Finally, donor specificity was found for the hypercondensation.

  • 49. Benach, J
    et al.
    Filling, C
    Oppermann, U C T
    Roversi, P
    Bricogne, G
    Berndt, Kurt D
    Södertörn University, Avdelning Naturvetenskap. Karolinska Institutet.
    Jörnvall, H
    Ladenstein, R
    Structure of bacterial 3 beta/17 beta-hydroxysteroid dehydrogenase at 1.2 angstrom resolution: A model for multiple steroid recognition2002In: Biochemistry, ISSN 0006-2960, E-ISSN 1520-4995, Vol. 41, no 50, p. 14659-14668Article in journal (Refereed)
    Abstract [en]

    The enzyme 3beta/17beta-hydroxysteroid dehydrogenase (3beta/17beta-HSD) is a steroid-inducible component of the Gram-negative bacterium Conramonas testosteroni. It catalyzes the reversible reduction/ dehydrogenation of the oxo/beta-hydroxy groups at positions 3 and 17 of steroid compounds, including hormones and isobile acids. Crystallographic analysis at 1.2 Angstrom resolution reveals the enzyme to have nearly identical subunits that form a tetramer with 222 symmetry. This is one of the largest oligomeric structures refined at this resolution. The subunit consists of a monomer with a single-domain structure built around a seven-stranded beta-sheet flanked by six alpha-helices. The active site contains a Ser-Tyr-Lys triad, typical for short-chain dehydrogenases/reductases (SDR). Despite their highly diverse substrate specificities, SDR members show a close to identical folding pattern architectures and a common catalytic mechanism. In contrast to other SDR apostructures determined, the substrate binding loop is well-defined. Analysis of structure-activity relationships of catalytic cleft residues, docking analysis of substrates and inhibitors, and accessible surface analysis explains how 3beta/17beta-HSD accommodates steroid substrates of different conformations.

  • 50. Bensch, Staffan
    et al.
    Grahn, Mats
    Södertörn University, School of Life Sciences, Biology. Södertörn University, School of Life Sciences, Environmental science.
    Müller, Nils
    Gay, Laurene
    Åkesson, Susanne
    Genetic, morphological, and feather isotope variation of migratory willow warblers show gradual divergence in a ring.2009In: Molecular Ecology, ISSN 0962-1083, E-ISSN 1365-294X, Vol. 18, no 14, p. 3087-96Article in journal (Refereed)
    Abstract [en]

    The circular distribution of the willow warbler Phylloscopus trochilus around the Baltic Sea shares many features with the classic examples of ring species; however, the system is much younger. It has previously been shown that a secondary contact zone is located in central Scandinavia, where there are narrow clines for several morphological traits coincident with a migratory divide. Here we analyse multiple traits and genes from > 1700 males captured on breeding territories at 77 sites spread around the Baltic Sea to test the following hypothesis. If the secondary contact zone in Scandinavia is a result of divergence in two allopatric refuge populations during the last glaciation, we expect to find a similar secondary contact zone somewhere else around the circular distribution. Our results show that the trait clines were wider and displaced from each other along the eastern side of the Baltic Sea. Analyses of 12 microsatellite loci confirmed that the genome is very similar between the terminal forms (F(ST) = 0). Two AFLP-derived markers filtered out from a genomic scan instead appear to be maintained by selection. These markers exhibited steep clines at the secondary contact zone in Scandinavia, but as for the phenotypic traits, had vastly different cline centres east of the Baltic Sea. The trait clines along the ring distribution outside the Scandinavian secondary contact zone thus seem to have been shaped by independent action of selection or drift during the process of postglacial colonization.

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