umu.sePublications
Change search
Refine search result
6789101112 401 - 450 of 969
CiteExportLink to result list
Permanent link
Cite
Citation style
  • apa
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Other style
More styles
Language
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Other locale
More languages
Output format
  • html
  • text
  • asciidoc
  • rtf
Rows per page
  • 5
  • 10
  • 20
  • 50
  • 100
  • 250
Sort
  • Standard (Relevance)
  • Author A-Ö
  • Author Ö-A
  • Title A-Ö
  • Title Ö-A
  • Publication type A-Ö
  • Publication type Ö-A
  • Issued (Oldest first)
  • Issued (Newest first)
  • Created (Oldest first)
  • Created (Newest first)
  • Last updated (Oldest first)
  • Last updated (Newest first)
  • Disputation date (earliest first)
  • Disputation date (latest first)
  • Standard (Relevance)
  • Author A-Ö
  • Author Ö-A
  • Title A-Ö
  • Title Ö-A
  • Publication type A-Ö
  • Publication type Ö-A
  • Issued (Oldest first)
  • Issued (Newest first)
  • Created (Oldest first)
  • Created (Newest first)
  • Last updated (Oldest first)
  • Last updated (Newest first)
  • Disputation date (earliest first)
  • Disputation date (latest first)
Select
The maximal number of hits you can export is 250. When you want to export more records please use the Create feeds function.
  • 401.
    Johansson, Anna-Mia
    et al.
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Stenberg, Per
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Allgardsson, Anders
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Larsson, Jan
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    POF Regulates the Expression of Genes on the Fourth Chromosome in Drosophila melanogaster by Binding to Nascent RNA2012In: Molecular and Cellular Biology, ISSN 0270-7306, E-ISSN 1098-5549, Vol. 32, no 11, p. 2121-2134Article in journal (Other academic)
    Abstract [en]

    In Drosophila, two chromosome-wide compensatory systems have been characterized: the dosage compensation system that acts on the male X chromosome and the chromosome-specific regulation of genes located on the heterochromatic fourth chromosome. Dosage compensation in Drosophila is accomplished by hypertranscription of the single male X chromosome mediated by the male-specific lethal (MSL) complex. The mechanism of this compensation is suggested to involve enhanced transcriptional elongation mediated by the MSL complex, while the mechanism of compensation mediated by the painting of fourth (POF) protein on the fourth chromosome has remained elusive. Here, we show that POF binds to nascent RNA, and this binding is associated with increased transcription output from chromosome 4. We also show that genes located in heterochromatic regions spend less time in transition from the site of transcription to the nuclear envelope. These results provide useful insights into the means by which genes in heterochromatic regions can overcome the repressive influence of their hostile environment.

  • 402. Johansson, Bente B
    et al.
    Torsvik, Janniche
    Gundersen, Lise
    Vesterhus, Mette
    Ragvin, Anja
    Tjora, Erling
    Fjeld, Karianne
    Hoem, Dag
    Johansson, Stefan
    Rader, Helge
    Lindquist, Susanne
    Umeå University, Faculty of Medicine, Department of Clinical Sciences, Paediatrics.
    Hernell, Olle
    Umeå University, Faculty of Medicine, Department of Clinical Sciences, Paediatrics.
    Cnop, Miriam
    Saraste, Jaakko
    Flatmark, Torgeir
    Molven, Anders
    Njolstad, Pal R
    Diabetes and pancreatic exocrine dysfunction due to mutations in the carboxyl-ester lipase gene (CEL-MODY): a protein misfolding disease2011In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 286, no 40, p. 34593-34605Article in journal (Refereed)
    Abstract [en]

    CEL-MODY, diabetes with pancreatic lipomatosis and exocrine dysfunction, is due to dominant frame-shift mutations in the acinar cell carboxyl-ester lipase gene (CEL). As Cel knock-out mice do not express the phenotype and the mutant protein has an altered, intrinsically disordered tandem repeat domain, we hypothesized that the disease mechanism might involve a negative effect of the mutant protein. In silico analysis showed that the pI of the tandem repeat was markedly increased from pH 3.3 in wild-type (WT) to 11.8 in mutant (MUT) human CEL. By stably over-expressing CEL-WT and CEL-MUT in HEK293 cells, we found similar glycosylation, ubiquitination, constitutive secretion and quality control of the two proteins. The CEL-MUT protein demonstrated, however, a high propensity to form aggregates found intracellularly and extracellularly. Different physico-chemical properties of the intrinsically disordered tandem repeat domains of WT and MUT proteins may contribute to different short-range and long-range interactions with the globular core domain and other macromolecules, including cell membranes. Thus, we propose that CEL-MODY is a protein misfolding disease caused by a negative gain-of-function effect of the mutant proteins in pancreatic tissues.

  • 403. Johansson, Björn P
    et al.
    Levander, Fredrik
    von Pawel-Rammingen, Ulrich
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Berggård, Tord
    Björck, Lars
    James, Peter
    The protein expression of Streptococcus pyogenes is significantly influenced by human plasma2005In: Journal of Proteome Research, ISSN 1535-3893, E-ISSN 1535-3907, Vol. 4, no 6, p. 2302-2311Article in journal (Refereed)
    Abstract [en]

    During the courser of infection, the common human pathogen Streptococcus pyogenes encounters plasma. We show that plasma causes S. pyogenes to rapidly remodel its cellular metabolism and virulence pathways. We also identified a variant of the major virulence factor, M1 protein, lacking 13 amino acids at the NH2-terminus in bacteria grown with plasma. The pronounced effect of plasma on protein expression, suggests this is an important adaptive mechanism with implications for S. pyogenes pathogenicity.

  • 404.
    Johansson, Henrik
    Umeå University, Faculty of Science and Technology, Plant Physiology.
    Gene regulation of UDP-glucose synthesis and metabolism in plants2003Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Photosynthesis captures light from the sun and converts it into carbohydrates, which are utilised by almost all living organisms. The conversion between the different forms of carbohydrates is the basis to form almost all biological molecules.

    The main intention of this thesis has been to study the role of UDP-glucose in carbohydrate synthesis and metabolism, and in particular the genes that encode UDP-glucose pyrophosphorylase (UGPase) and UDP-glucose dehydrogenase (UGDH) in plants and their regulation. UGPase converts glucose-1-phosphate to UDP-glucose, which can be utilised for sucrose synthesis, or cell wall polysaccharides among others. UGDH converts UDP-glucose to UDP-glucuronate, which is a precursor for hemicellulose and pectin. As model species I have been working with both Arabidopsis thaliana and poplar.

    Sequences for two full-length EST clones of Ugp were obtained from both Arabidopsis and poplar, the cDNAs in Arabidopsis correlate with two genes in the Arabidopsis genomic database.

    The derived protein sequences are 90-93% identical within each plants species and 80-83% identical between the two species.

    Studies on Ugp showed that the expression is up-regulated by Pi-deficiency, sucrose-feeding and by light exposure in Arabidopsis. Studies with Arabidopsis plants with mutations in sugar/ starch- and Pi-content suggested that the Ugp expression is modulated by an interaction of signals derived from Pi-deficiency, sugar content and light/ dark conditions, where the signals act independently or inhibiting each other, depending on conditions. Okadaic acid, a known inhibitor of certain classes of protein phosphatases, prevented the up-regulation of Ugp by Pi-deficiency and sucrose-feeding. In poplar, sucrose also up-regulated the expression of Ugp. When poplar and Arabidopsis were exposed to cold, an increase of Ugp transcript content was detected as well as an increase in UGPase protein and activity. In poplar, Ugp was found to be expressed in all tissues that were examined (differentiating xylem, phloem, apical leaves and young and mature leaves).

    By using antisense strategy, Arabidopsis plants that had a decrease in UGPase activity of up to 30% were obtained. In the antisense plants, the soluble carbohydrate content was reduced in the leaves by at least 50%; in addition the starch content decreased. Despite the changes in carbohydrate content, the growth rate of the antisense plants was not changed compared to wild type plants under normal growth conditions. However, in the antisense lines the UGPase activity and protein content in sliliques and roots increased, perhaps reflecting compensatory up-regulation of second Ugp gene. This correlates with a slightly larger molecular mass of UGPase protein in roots and siliques when compared to that in leaves. Maximal photosynthesis rates were similar for both wild type and antisense plants, but the latter had up to 40% lower dark respiration and slightly lower quantum yield than wild type plants.

    Two Ugdh cDNAs from poplar and one from Arabidopsis were sequenced. The highest Ugdh expression was found in xylem and younger leaves. Expression data from sugar and osmoticum feeding experiment in poplar suggested that the Ugdh expression is regulated via an osmoticumdependent pathway.

  • 405.
    Johansson, Inga-Maj
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Pea carbonic anhydrase: a kinetic study1994Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    The enzyme carbonic anhydrase (CA), catalysing the interconversion between CO2 and HCO3', has long been known to be present in plants as well as in animals. Several of the animal isozymes, but none of the plant CAs, have been extensively studied. When the first plant CA cDNA sequences were published in 1990, it was obvious that the animal and plant CAs represent evolutionarily distinct families with no significant sequence homology between the families.

    Pea CA is synthesised as a precursor and subsequently processed at the import into the chloroplast. When we purified CA from pea leaves two oligomeric forms with molecular masses around 230 kDa were obtained. One form was homogenous while the other form contained subunits of two different sizes. The larger subunit has an acidic and highly charged N-terminal extension, consisting of 37 residues. We propose that the sequence that precedes the cleavage site resulting in the large subunit represents the functional transit peptide, directing CA to the chloroplast. Neither the transit peptide nor the acidic 37-residue peptide were found to affect the folding, activity or oligomerisation of pea CA.

    Kinetic investigations showed that pea CA requires a reduced environment and high concentrations of buffer for maximal catalytic activity. High buffer concentrations result in a faster turnover of the enzyme (kcat) while the efficiency (kcatlKm) is not affected. This is consistent with a ping-pong mechanism with the buffer as the second substrate. Both kcat and kcatlKm increase with pH but the dependences cannot be described by simple titration curves. SCN' is an uncompetitive inhibitor at high pH and a noncompetitive inhibitor at neutral and low pH. This is in accordance with the mechanistic model, previously proposed for human CAM, involving a zincbound water molecule as a catalytic group. In this model, the carbon dioxide - bicarbonate interconversion, reflected by kcatlKm, is temporally separated from a rate limiting proton-transfer step. At high pH, solvent hydrogen isotope effects obtained for pea CA agree with this scheme, while they do not fit at neutral and low pH.

    Site-specific mutations of cysteine residues at positions 165, 269 and 272 were difficult to study, either because strong deviations from Michaelis-Menten kinetics were observed, or because the mutants were found in inclusion bodies. However, the mutant H208A was found to be a very efficient enzyme with the highest kcatlKm value obtained for any CA so far, 2.9-108 M'1s '1. With the H208A mutant an increased dependence on high buffer concentrations at low pH was obtained. At high pH, the mutant is more efficient than the unmutated enzyme. The H208A mutant is also more prone to oxidation than the wild-type enzyme.

  • 406. Johansson, J.
    et al.
    Berg, T.
    Kurzejamska, E.
    Pang, M-F
    Tabor, V.
    Jansson, Malin
    Umeå University, Faculty of Medicine, Department of Surgical and Perioperative Sciences, Surgery.
    Roswall, P.
    Pietras, K.
    Sund, Malin
    Umeå University, Faculty of Medicine, Department of Surgical and Perioperative Sciences, Surgery.
    Religa, P.
    Fuxe, J.
    MiR-155-mediated loss of C/EBP beta shifts the TGF-beta response from growth inhibition to epithelial-mesenchymal transition, invasion and metastasis in breast cancer2013In: Oncogene, ISSN 0950-9232, E-ISSN 1476-5594, Vol. 32, no 50, p. 5614-5624Article in journal (Refereed)
    Abstract [en]

    During breast cancer progression, transforming growth factor-beta (TGF-beta) switches from acting as a growth inhibitor to become a major promoter of epithelial-mesenchymal transition (EMT), invasion and metastasis. However, the mechanisms involved in this switch are not clear. We found that loss of CCAAT-enhancer binding protein beta (C/EBP beta), a differentiation factor for the mammary epithelium, was associated with signs of EMT in triple-negative human breast cancer, and in invasive areas of mammary tumors in MMTV-PyMT mice. Using an established model of TGF-beta-induced EMT in mouse mammary gland epithelial cells, we discovered that C/EBP beta was repressed during EMT by miR-155, an oncomiR in breast cancer. Depletion of C/EBP beta potentiated the TGF-beta response towards EMT, and contributed to evasion of the growth inhibitory response to TGF-beta. Furthermore, loss of C/EBP beta enhanced invasion and metastatic dissemination of the mouse mammary tumor cells to the lungs after subcutaneous injection into mice. The mechanism by which loss of C/EBP beta promoted the TGF-beta response towards EMT, invasion and metastasis, was traced to a previously uncharacterized role of C/EBP beta as a transcriptional activator of genes encoding the epithelial junction proteins E-cadherin and coxsackie virus and adenovirus receptor. The results identify miR-155-mediated loss of C/EBP beta as a mechanism, which promotes breast cancer progression by shifting the TGF-beta response from growth inhibition to EMT, invasion and metastasis.

  • 407.
    Johansson, Marcus J O
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Determining if an mRNA is a Substrate of Nonsense-Mediated mRNA Decay in Saccharomyces cerevisiae2017In: Methods in Molecular Biology, ISSN 1064-3745, E-ISSN 1940-6029, Vol. 1507, p. 169-177Article in journal (Refereed)
    Abstract [en]

    Nonsense-mediated mRNA decay (NMD) is a conserved eukaryotic quality control mechanism which triggers decay of mRNAs harboring premature translation termination codons. In this chapter, I describe methods for monitoring the influence of NMD on mRNA abundance and decay rates in Saccharomyces cerevisiae. The descriptions include detailed methods for growing yeast cells, total RNA isolation, and Northern blotting. Although the chapter focuses on NMD, the methods can be easily adapted to assess the effect of other mRNA decay pathways.

  • 408.
    Johansson, Marcus J O
    et al.
    Umeå University, Faculty of Science and Technology, Molecular Biology (Faculty of Science and Technology).
    Byström, Anders S
    Umeå University, Faculty of Science and Technology, Molecular Biology (Faculty of Science and Technology).
    Dual function of the tRNA(m(5)U54)methyltransferase in tRNA maturation2002In: RNA: A publication of the RNA Society, ISSN 1355-8382, E-ISSN 1469-9001, Vol. 8, no 3, p. 324-335Article in journal (Refereed)
    Abstract [en]

    A 5-methyluridine (m(5)U) residue at position 54 is a conserved feature of bacterial and eukaryotic tRNAs. The methylation of U54 is catalyzed by the tRNA(m5U54)methyltransferase, which in Saccharomyces cerevisiae is encoded by the nonessential TRM2 gene. In this study, we identified four different strains with mutant forms of tRNA(Ser)CGA. The absence of the TRM2 gene in these strains decreased the stability of tRNA(Ser)CGA and induced lethality. Two alleles of TRM2 encoding catalytically inactive tRNA(m5U54)methyltransferases were able to stabilize tRNA(Ser)CGA in one of the mutants, revealing a role for the Trm2 protein per se in tRNA maturation. Other tRNA modification enzymes interacting with tRNA(Ser)CGA in the maturation process, such as Pus4p, Trm1 p, and Trm3p were essential or important for growth of the tRNA(Ser)CGA mutants. Moreover, Lhp1p, a protein binding RNA polymerase III transcripts, was required to stabilize the mutant tRNAs. Based on our results, we suggest that tRNA modification enzymes might have a role in tRNA maturation not necessarily linked to their known catalytic activity.

  • 409.
    Johansson, Marcus J O
    et al.
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Byström, Anders S
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Transfer RNA modifications and modifying enzymes in Saccharomyces cerevisiae.2005In: Fine-Tuning of RNA Functions by Modification and Editing. / [ed] Henri Grosjean, Springer Berlin/Heidelberg, 2005, p. 87-120Chapter in book (Refereed)
    Abstract [en]

    Transfer RNAs are adaptor molecules, which decode mRNA into protein and, thereby, play a central role in gene expression. During the maturation of a primary tRNA transcript, specific subsets of the four normal nucleosides adenosine, cytidine, guanosine, and uridine are modified. The formation of a modified nucleoside can require more than one gene product and may involve several enzymatic steps. In the last few years, the identification of gene products required for formation of modified nucleosides in tRNA has dramatically increased. In this review, proteins involved in modification of cytoplasmic tRNAs in Saccharomyces cerevisiae are described, emphasizing phenotypic characteristics of modification deficient strains and genetic approaches used to determine the in vivo role of modified nucleosides/modifying enzymes.

  • 410.
    Johansson, Marcus J O
    et al.
    University of Massachusetts Medical School, Department of Molecular Genetics and Microbiology,.
    He, Feng
    University of Massachusetts Medical School, Department of Molecular Genetics and Microbiology,.
    Spatrick, Phyllis
    University of Massachusetts Medical School, Department of Molecular Genetics and Microbiology,.
    Li, Chunfang
    University of Massachusetts Medical School, Department of Molecular Genetics and Microbiology,.
    Jacobson, Allan
    University of Massachusetts Medical School, Department of Molecular Genetics and Microbiology,.
    Association of yeast Upf1p with direct substrates of the NMD pathway.2007In: Proceedings of the National Academy of Sciences of the United States of America, ISSN 0027-8424, E-ISSN 1091-6490, Vol. 104, no 52, p. 20872-7Article in journal (Refereed)
    Abstract [en]

    Nonsense-mediated mRNA decay (NMD) is a surveillance mechanism that detects and degrades transcripts containing premature translation termination codons. Gene expression profiling experiments have shown that inactivation of the NMD pathway leads to the accumulation of both aberrant, nonsense-containing mRNAs, and many apparently wild-type transcripts. Such increases in transcript steady-state levels could arise from direct changes in the respective mRNA half-lives, or indirectly, as a consequence of the stabilization of transcripts encoding specific regulatory proteins. Here, we distinguished direct from indirect substrates by virtue of their association with the Saccharomyces cerevisiae Upf1 protein. Analyses of this dataset, and its comparison to the sets of transcripts that respectively increase or decrease in abundance when NMD is either inactivated or reactivated, indicate that the number of direct NMD substrates is larger than previously thought and that low abundance, alternatively transcribed mRNAs, i.e., mRNAs whose 5' ends are derived from previously unannotated 5' flanking sequences, comprise a significant class of direct substrates. Using thiamine metabolism as an example, we also show that apparent NMD-regulated cellular pathways may actually reflect the detection of low-abundance alternative transcripts under conditions where a pathway is repressed.

  • 411.
    Johansson, Marcus J O
    et al.
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Xu, Fu
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Byström, Anders S
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Elongator-a tRNA modifying complex that promotes efficient translational decoding2018In: Biochimica et Biophysica Acta, ISSN 0006-3002, E-ISSN 1878-2434, Vol. 1861, no 4, p. 401-408Article in journal (Refereed)
    Abstract [en]

    Naturally occurring modifications of the nucleosides in the anticodon region of tRNAs influence their translational decoding properties. Uridines present at the wobble position in eukaryotic cytoplasmic tRNAs often contain a 5-carbamoylmethyl (ncm5) or 5-methoxycarbonylmethyl (mcm5) side-chain and sometimes also a 2-thio or 2'-O-methyl group. The first step in the formation of the ncm5 and mcm5 side-chains requires the conserved six-subunit Elongator complex. Although Elongator has been implicated in several different cellular processes, accumulating evidence suggests that its primary, and possibly only, cellular function is to promote modification of tRNAs. In this review, we discuss the biosynthesis and function of modified wobble uridines in eukaryotic cytoplasmic tRNAs, focusing on the in vivo role of Elongator-dependent modifications in Saccharomyces cerevisiae. 

  • 412.
    Johansson, Mikael
    Umeå University, Faculty of Science and Technology, Department of Plant Physiology.
    The circadian clock in annuals and perennials: coordination of Growth with Environmental Rhythms2010Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Since the first signs of life on planet earth, organisms have had to adapt to the daily changes between light and dark, and high and low temperatures. This has led to the evolution of an endogenous time keeper, known as the circadian clock. This biological timing system helps the organism to synchronize developmental and metabolic events to the most favorable time of the day. Such a mechanism is of considerable value to plants, since they in contrast to animals cannot change location when the environment becomes unfavorable. Thus is the ability to predict coming events of central importance in a plants life. This thesis is a study of the molecular machinery behind the clockwork in the small weed plant Arabidopsis thaliana as well as its close relative perennial; the woody species Populus. We have characterized a novel component of the circadian clock, EARLY BIRD (EBI). EBI is involved in transcriptional and translational regulation, via interaction with the known post-translational clock regulator ZEITLUPE (ZTL). In Populus, we describe the role of the circadian clock and its components with respect to entry and exit of dormancy and show that gene expression of the Populus LATE ELONATED HYPOCOTYL (LHY) genes are crucial importance for freezing tolerance and thereby survival at high latitudes. Furthermore, the input to the Populus clock is mediated via the phytochrome A (phyA) photoreceptor.

  • 413.
    Johansson, Mikael
    et al.
    Umeå University, Faculty of Science and Technology, Department of Plant Physiology.
    Kozarewa, Iwanka
    Breakthrough Breast Cancer Research Centre, The Institute of Cancer Research, London, UK.
    Eriksson, Maria E.
    Umeå University, Faculty of Science and Technology, Department of Plant Physiology.
    ZEITLUPE promoter activity is modulated in an EARLY BIRD dependent manner in response to abscisic acidManuscript (preprint) (Other academic)
    Abstract [en]

    The newly described clock regulator EARLY BIRD (EBI) was shown to interact with ZEITLUPE (ZTL) and to influence the function of the core circadian oscillator. Studiesof mutants in the EBI locus have earlier been used to ascribe a role for EBI in responsesto abiotic stress.Here, we test the early bird-1 (ebi-1) clock mutant’s response to salt stress. Also, weprobe the effect on the expression and regulation of clock associated genes in response tothe known stress signaling modulator, abscisic acid (ABA). We show that ZTL promoteractivity is under circadian clock control and that these oscillations are stabilized to morerobust rhythms in response to ABA. The clock regulation of ZTL is found to beindependent of EBI and different from the regulation of CAB2 suggesting that they, atleast partially, are driven by separate oscillators. In addition, EBI appears to modulate thestabilization of these oscillations in response to ABA suggesting that it may act as amediator between oscillators and in the clock’s response to stress.

  • 414.
    Johansson, Mikael
    et al.
    Umeå University, Faculty of Science and Technology, Department of Plant Physiology.
    McWatters, Harriet G.
    Department of Plant Sciences, Oxford University, UK.
    Bakó, László
    Umeå University, Faculty of Science and Technology, Department of Plant Physiology.
    Takata, Naoki
    Umeå University, Faculty of Science and Technology, Department of Plant Physiology.
    Gyula, Péter
    Institute of Plant Biology, Biological Research Centre of the Hungarian Academy of Sciences, Temesvári, Hungary.
    Hall, Anthony
    School of Biological Sciences, Liverpool University, Liverpool, UK.
    Somers, David E.
    Department of Plant Cellular and Molecular Biology, Ohio State University, Columbus, Ohio, USA.
    Millar, Andrew J.
    Centre for Systems Biology at Edinburgh, Edinburgh University, Edinburgh, UK.
    Eriksson, Maria E.
    Umeå University, Faculty of Science and Technology, Department of Plant Physiology.
    Partners in time: EARLY BIRD reveals novel regulatory function of ZEITLUPE in the Arabidopsis clockManuscript (preprint) (Other academic)
    Abstract [en]

    The circadian clock of the model plant Arabidopsis thaliana is made up of acomplex series of interacting feedback loops whereby proteins regulate their ownexpression across day and night. early bird (ebi) is a circadian mutation that causesthe clock to speed up: ebi plants have short circadian periods, early phase of clockgene expression and are early flowering. We show that EBI associates with ZEITLUPE (ZTL), known to act in the plant clock as a post-translational mediator of protein degradation. However, EBI is not degraded by its interaction with ZTL. Instead, EBI acts in opposition to ZTL, modulating the expression of key circadiancomponents. The partnership of EBI with ZTL reveals a novel mechanism involved incontrolling the complex transcription-translation feedback loops of the clock. Thiswork highlights the importance of cross-talk between the ubiquitination pathway andtranscriptional control for regulation of the plant clock

  • 415. Jonasson, Jenni
    et al.
    Ström, Anna-Lena
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Hart, Patricia
    Brännström, Thomas
    Forsgren, Lars
    Umeå University, Faculty of Medicine, Department of Medical Biosciences, Pathology.
    Holmberg, Monica
    Expression of ataxin-7 in CNS and non-CNS tissue of normal and SCA7 individuals2002In: Acta Neuropathologica, ISSN 0001-6322, E-ISSN 1432-0533, Vol. 104, no 1, p. 29-37Article in journal (Refereed)
    Abstract [en]

    Spinocerebellar ataxia type 7 (SCA7) is a neurodegenerative disorder primarily affecting the cerebellum, brain stem and retina. The disease is caused by an expanded polyglutamine tract in the protein ataxin-7. In this study we analyzed the expression pattern of ataxin-7 in CNS and non-CNS tissue from three SCA7 patients and age-matched controls. SCA7 is a rare autosomal dominant disorder, limiting the number of patients available for analysis. We therefore compiled data on ataxin-7 expression from all SCA7 patients (n=5) and controls (n=7) published to date, and compared with the results obtained in this study. Expression of ataxin-7 was found in neurons throughout the CNS and was highly abundant in Purkinje cells of the cerebellum, in regions of the hippocampus and in cerebral cortex. Ataxin-7 expression was not restricted to regions of pathology, and there were no apparent regional differences in ataxin-7 expression patterns between patients and controls. The subcellular distribution of ataxin-7 was primarily nuclear in all brain regions studied. In cerebellar Purkinje cells, however, differences in subcellular distribution of ataxin-7 were observed between patients and controls of different ages. Here we provide an increased understanding of the distribution of ataxin-7, and the possible implication of subcellular localization of this protein on disease pathology is discussed.

  • 416.
    Jonna, Venkateswara Rao
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Class I Ribonucleotide Reductases: overall activity regulation, oligomerization, and drug targeting2017Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Ribonucleotide reductase (RNR) is a key enzyme in the de novo biosynthesis and homeostatic maintenance of all four DNA building blocks by being able to make deoxyribonucleotides from the corresponding ribonucleotides. It is important for the cell to control the production of a balanced supply of the dNTPs to minimize misincorporations in DNA. Because RNR is the rate-limiting enzyme in DNA synthesis, it is an important target for antimicrobial and antiproliferative molecules. The enzyme RNR has one of the most sophisticated allosteric regulations known in Nature with four allosteric effectors (ATP, dATP, dGTP, and dTTP) and two allosteric sites. One of the sites (s-site) controls the substrate specificity of the enzyme, whereas the other one (a-site) regulates the overall activity.  The a-site binds either dATP, which inhibits the enzyme or ATP that activates the enzyme. In eukaryotes, ATP activation is directly through the a-site and in E. coli it is a cross-talk effect between the a and s-sites. It is important to study and get more knowledge about the overall activity regulation of RNR, both because it has an important physiological function, but also because it may provide important clues to the design of antibacterial and antiproliferative drugs, which can target RNR.

    Previous studies of class I RNRs, the class found in nearly all eukaryotes and many prokaryotes have revealed that the overall activity regulation is dependent on the formation of oligomeric complexes. The class I RNR consists of two subunits, a large α subunit, and a small β subunit. The oligomeric complexes vary between different species with the mammalian and yeast enzymes cycle between structurally different active and inactive α6β2 complexes, and the E. coli enzyme cycles between active α2β2 and inactive α4β4 complexes. Because RNR equilibrates between many different oligomeric forms that are not resolved by most conventional methods, we have used a technique termed gas-phase electrophoretic macromolecule analysis (GEMMA). In the present studies, our focus is on characterizing both prokaryotic and mammalian class I RNRs. In one of our projects, we have studied the class I RNR from Pseudomonas aeruginosa and found that it represents a novel mechanism of overall activity allosteric regulation, which is different from the two known overall activity allosteric regulation found in E. coli and eukaryotic RNRs, respectively.  The structural differences between the bacterial and the eukaryote class I RNRs are interesting from a drug developmental viewpoint because they open up the possibility of finding inhibitors that selectively target the pathogens. The biochemical data that we have published in the above project was later supported by crystal structure and solution X-ray scattering data that we published together with Derek T. Logan`s research group.

    We have also studied the effect of a novel antiproliferative molecule, NSC73735, on the oligomerization of the human RNR large subunit. This collaborative research results showed that the molecule NSC73735 is the first reported non-nucleoside molecule which alters the oligomerization to inhibit human RNR and the molecule disrupts the cell cycle distribution in human leukemia cells.

  • 417.
    Jonna, Venkateswara Rao
    et al.
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Crona, Mikael
    Rofougaran, Reza
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Lundin, Daniel
    Johansson, Samuel
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Brännström, Kristoffer
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Sjöberg, Britt-Marie
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Diversity in Overall Activity Regulation of Ribonucleotide Reductase2015In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 290, no 28, p. 17339-17348Article in journal (Refereed)
    Abstract [en]

    Ribonucleotide reductase (RNR) catalyzes the reduction of ribonucleotides to the corresponding deoxyribonucleotides, which are used as building blocks for DNA replication and repair. This process is tightly regulated via two allosteric sites, the specificity site (s-site) and the overall activity site (a-site). The a-site resides in an N-terminal ATP cone domain that binds dATP or ATP and functions as an on/off switch, whereas the composite s-site binds ATP, dATP, dTTP, or dGTP and determines which substrate to reduce. There are three classes of RNRs, and class I RNRs consist of different combinations of α and β subunits. In eukaryotic and Escherichia coli class I RNRs, dATP inhibits enzyme activity through the formation of inactive α6 and α4β4 complexes, respectively. Here we show that the Pseudomonas aeruginosa class I RNR has a duplicated ATP cone domain and represents a third mechanism of overall activity regulation. Each α polypeptide binds three dATP molecules, and the N-terminal ATP cone is critical for binding two of the dATPs because a truncated protein lacking this cone could only bind dATP to its s-site. ATP activates the enzyme solely by preventing dATP from binding. The dATP-induced inactive form is an α4 complex, which can interact with β2 to form a non-productive α4β2 complex. Other allosteric effectors induce a mixture of α2 and α4 forms, with the former being able to interact with β2 to form active α2β2 complexes. The unique features of the P. aeruginosa RNR are interesting both from evolutionary and drug discovery perspectives.

  • 418.
    Jonsson, Leif J.
    et al.
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Martin, Carlos
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Pretreatment of lignocellulose: Formation of inhibitory by-products and strategies for minimizing their effects2016In: Bioresource Technology, ISSN 0960-8524, E-ISSN 1873-2976, Vol. 199, p. 103-112Article, review/survey (Refereed)
    Abstract [en]

    Biochemical conversion of lignocellulosic feedstocks to advanced biofuels and other commodities through a sugar-platform process involves a pretreatment step enhancing the susceptibility of the cellulose to enzymatic hydrolysis. A side effect of pretreatment is formation of lignocellulose-derived by-products that inhibit microbial and enzymatic biocatalysts. This review provides an overview of the formation of inhibitory by-products from lignocellulosic feedstocks as a consequence of using different pretreatment methods and feedstocks as well as an overview of different strategies used to alleviate problems with inhibitors. As technologies for biorefining of lignocellulose become mature and are transferred from laboratory environments to industrial contexts, the importance of management of inhibition problems is envisaged to increase as issues that become increasingly relevant will include the possibility to use recalcitrant feedstocks, obtaining high product yields and high productivity, minimizing the charges of enzymes and microorganisms, and using high solids loadings to obtain high product titers.

  • 419.
    Jonsson, Leif
    et al.
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Wu, Guochao
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Xu, Zixiang
    Saccharomyces cerevisiae transcription factors involved in multidrug resistance2013In: Yeast, ISSN 0749-503X, E-ISSN 1097-0061, Vol. 30, no Supplement: 1, p. 158-158Article in journal (Other academic)
  • 420.
    Jortikka, Matti
    et al.
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Inkinen, Ritva
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Tammi, Markku
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Parkkinen, Jyrki
    Department of Pathology, University of Kuopio, Kuopio, Finland.
    Haapala, Jussi
    Department of Surgery, Kuopio University Hospital, Kuopio, Finland.
    Kiviranta, Ilkka
    Helminen, Heikki
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Lammi, Mikko
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Immobilisation causes longlasting matrix changes both in the immobilised and contralateral joint cartilage.1997In: Annals of the Rheumatic Diseases, ISSN 0003-4967, E-ISSN 1468-2060, Vol. 56, no 4, p. 255-261, article id 9165998Article in journal (Refereed)
    Abstract [en]

    OBJECTIVE: The capacity of articular cartilage matrix to recover during 50 weeks of remobilisation after an atrophy caused by 11 weeks of immobilisation of the knee (stifle) joint in 90 degrees flexion starting at the age of 29 weeks, was studied in young beagle dogs.

    METHODS: Proteoglycan concentration (uronic acid) and synthesis ([35S]sulphate incorporation) were determined in six and three knee joint surface locations, respectively. Proteoglycans extracted from the cartilages were characterised by chemical determinations, gel filtration, and western blotting for chondroitin sulphate epitope 3B3.

    RESULTS: The proteoglycan concentrations that were reduced in all sample sites immediately after the immobilisation, remained 14-28% lower than controls after 50 weeks of remobilisation in the patella, the summit of medial femoral condyle, and the superior femoropatellar surface. In the contralateral joint, there was a 49% increase of proteoglycans in the inferior femoropatellar surface after remobilisation, while a 34% decrease was simultaneously noticed on the summit of the medial femoral condyle. Total proteoglycan synthesis was not significantly changed after immobilisation or 50 weeks' remobilisation in the treated or contralateral joint, compared with age matched controls. The chondroitin 6- to 4- sulphate ratio was reduced by immobilisation both in the radioactively labelled and the total tissue proteoglycans. In the remobilised joint, this ratio was restored in femur, while in tibia it remained at a level lower than controls. Neither immobilisation nor remobilisation induced epitopes recognised by the monoclonal antibody 3B3 on native (undigested) proteoglycans.

    CONCLUSION: These results show that the depletion of proteoglycans observed after 11 weeks of immobilisation was not completely restored in certain surface sites after 50 weeks of remobilisation. The significant changes that developed in the contralateral joint during the remobilisation period give further support to the idea that a permanent alteration of matrix metabolism results even from a temporary modification of loading pattern in immature joints.

  • 421.
    Jortikka, Matti
    et al.
    epartment of Anatomy, University of Kuopio, Kuopio, Finland.
    Lammi, Mikko
    epartment of Anatomy, University of Kuopio, Kuopio, Finland.
    Parkkinen, Jyrki
    epartment of Anatomy, University of Kuopio, Kuopio, Finland.
    Lahtinen, Risto
    epartment of Anatomy, University of Kuopio, Kuopio, Finland.
    Tammi, Markku
    epartment of Anatomy, University of Kuopio, Kuopio, Finland.
    A high sensitivity dot-blot assay for proteoglycans by cuprolinic blue precipitation.1993In: Connective Tissue Research, ISSN 0300-8207, E-ISSN 1607-8438, Vol. 29, no 4, p. 263-272, article id 8269703Article in journal (Refereed)
    Abstract [en]

    A highly sensitive blot-assay was developed for glycosaminoglycans (GAGs) and proteoglycans (PGs) utilizing a precipitation reaction by a cationic dye Cuprolinic Blue. The precipitates were deposited into 1-2 mm2 spots on nitrocellulose membrane by using a 96-well filtration apparatus. The dried sheet was digitized by a flat bed scanner and the intensity of the dots was quantitated by an image analysis software. The working range for chondroitin sulfate was 10-300 ng. The response of various GAGs differed according to the number of anionic groups, both sulphate and carboxyl groups being able to bind the dye. The sensitivity of the assay was decreased by high concentrations of GuC, CsC and protein, but not by nonionic detergents, common buffers and 8 M urea. Contact exposure to autoradiography film enabled quantitation of 25-250 DPM, and 1-10 DPM, of 35SO4-radioactivity in precipitated PGs after overnight and 14 days' exposures, respectively.

  • 422.
    Jortikka, Matti
    et al.
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Parkkinen, Jyrki
    Department of Pathology, University of Kuopio, Kuopio, Finland.
    Inkinen, Rtiva
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Kärner, Jüri
    Department of Zoology, University of Tartu, Tartu, Estonia.
    Järveläinen, Hannu
    Department of Medicine, University of Turku, Finland; Medical Biochemistry, University of Turku, Finland.
    Nelimarkka, Lassi
    Medical Biochemistry, University of Turku, Finland.
    Tammi, Markku
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Lammi, Mikko
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    The role of microtubules in the regulation of proteoglycan synthesis in chondrocytes under hydrostatic pressure.2000In: Archives of Biochemistry and Biophysics, ISSN 0003-9861, E-ISSN 1096-0384, Vol. 374, no 2, p. 172-180, article id 10666295Article in journal (Refereed)
    Abstract [en]

    Chondrocytes of the articular cartilage sense mechanical factors associated with joint loading, such as hydrostatic pressure, and maintain the homeostasis of the extracellular matrix by regulating the metabolism of proteoglycans (PGs) and collagens. Intermittent hydrostatic pressure stimulates, while continuous high hydrostatic pressure inhibits, the biosynthesis of PGs. High continuous hydrostatic pressure also changes the structure of cytoskeleton and Golgi complex in cultured chondrocytes. Using microtubule (MT)-affecting drugs nocodazole and taxol as tools we examined whether MTs are involved in the regulation of PG synthesis in pressurized primary chondrocyte monolayer cultures. Disruption of the microtubular array by nocodazole inhibited [(35)S]sulfate incorporation by 39-48%, while MT stabilization by taxol caused maximally a 17% inhibition. Continuous hydrostatic pressure further decreased the synthesis by 34-42% in nocodazole-treated cultures. This suggests that high pressure exerts its inhibitory effect through mechanisms independent of MTs. On the other hand, nocodazole and taxol both prevented the stimulation of PG synthesis by cyclic 0. 5 Hz, 5 MPa hydrostatic pressure. The drugs did not affect the structural and functional properties of the PGs, and none of the treatments significantly affected cell viability, as indicated by the high level of PG synthesis 24-48 h after the release of drugs and/or high hydrostatic pressure. Our data on two-dimensional chondrocyte cultures indicate that inhibition of PG synthesis by continuous high hydrostatic pressure does not interfere with the MT-dependent vesicle traffic, while the stimulation of synthesis by cyclic pressure does not occur if the dynamic nature of MTs is disturbed by nocodazole. Similar phenomena may operate in cartilage matrix embedded chondrocytes.

  • 423.
    Jäger, Gunilla
    et al.
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Chen, Peng
    Björk, Glenn R.
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Transfer RNA Bound to MnmH Protein Is Enriched with Geranylated tRNA - A Possible Intermediate in Its Selenation?2016In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 11, no 4, article id e0153488Article in journal (Refereed)
    Abstract [en]

    The wobble nucleoside 5-methylaminomethyl-2-thio-uridine (mnm(5)s(2)U) is present in bacterial tRNAs specific for Lys and Glu and 5-carboxymethylaminomethyl-2-thio-uridine (cmnm(5)s(2)U) in tRNA specific for Gln. The sulfur of (c) mnm(5)s(2)U may be exchanged by selenium (Se)-a reaction catalyzed by the selenophosphate-dependent tRNA 2-selenouridine synthase encoded by the mnmH (ybbB, selU, sufY) gene. The MnmH protein has a rhodanese domain containing one catalytic Cys (C97) and a P-loop domain containing a Walker A motif, which is a potential nucleotide binding site. We have earlier isolated a mutant of Salmonella enterica, serovar Typhimurium with an alteration in the rhodanese domain of the MnmH protein (G67E) mediating the formation of modified nucleosides having a geranyl (ge)-group (C10H17-fragment) attached to the s(2) group of mnm(5)s(2)U and of cmnm(5)s(2)U in tRNA. To further characterize the structural requirements to increase the geranylation activity, we here report the analysis of 39 independently isolated mutants catalyzing the formation of mnm(5)ges(2)U. All these mutants have amino acid substitutions in the rhodanese domain demonstrating that this domain is pivotal to increase the geranylation activity. The wild type form of MnmH(+) also possesses geranyltransferase activity in vitro although only a small amount of the geranyl derivatives of (c) mnm(5)s(2)U is detected in vivo. The selenation activity in vivo has an absolute requirement for the catalytic Cys97 in the rhodanese domain whereas the geranylation activity does not. Clearly, MnmH has two distinct enzymatic activities for which the rhodanese domain is pivotal. An intact Walker motif in the P-loop domain is required for the geranylation activity implying that it is the binding site for geranylpyrophosphate (GePP), which is the donor molecule in vitro in the geranyltransfer reaction. Purified MnmH from wild type and from the MnmH(G67E) mutant have bound tRNA, which is enriched with geranylated tRNA. This in conjunction with earlier published data, suggests that this bound geranylated tRNA may be an intermediate in the selenation of the tRNA.

  • 424.
    Jäger, Gunilla
    et al.
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Nilsson, Kristina
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Björk, Glenn R.
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    The Phenotype of Many Independently Isolated+1 Frameshift Suppressor Mutants Supports a Pivotal Role of the P-Site in Reading Frame Maintenance2013In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 8, no 4, p. e60246-Article in journal (Refereed)
    Abstract [en]

    The main features of translation are similar in all organisms on this planet and one important feature of it is the way the ribosome maintain the reading frame. We have earlier characterized several bacterial mutants defective in tRNA maturation and found that some of them correct a +1 frameshift mutation; i.e. such mutants possess an error in reading frame maintenance. Based on the analysis of the frameshifting phenotype of such mutants we proposed a pivotal role of the ribosomal grip of the peptidyl-tRNA to maintain the correct reading frame. To test the model in an unbiased way we first isolated many (467) independent mutants able to correct a +1 frameshift mutation and thereafter tested whether or not their frameshifting phenotypes were consistent with the model. These 467+1 frameshift suppressor mutants had alterations in 16 different loci of which 15 induced a defective tRNA by hypo- or hypermodifications or altering its primary sequence. All these alterations of tRNAs induce a frameshift error in the P-site to correct a +1 frameshift mutation consistent with the proposed model. Modifications next to and 39 of the anticodon (position 37), like 1-methylguanosine, are important for proper reading frame maintenance due to their interactions with components of the ribosomal P-site. Interestingly, two mutants had a defect in a locus (rpsI), which encodes ribosomal protein S9. The C-terminal of this protein contacts position 32-34 of the peptidyl-tRNA and is thus part of the P-site environment. The two rpsI mutants had a C-terminal truncated ribosomal protein S9 that destroys its interaction with the peptidyl-tRNA resulting in +1 shift in the reading frame. The isolation and characterization of the S9 mutants gave strong support of our model that the ribosomal grip of the peptidylt-RNA is pivotal for the reading frame maintenance.

  • 425.
    Kaarniranta, Kai
    et al.
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Elo, Mika
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Sironen, Reijo
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Karjalainen, Hannu
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Helminen, Heikki
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Lammi, Mikko
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Stress responses of mammalian cells to high hydrostatic pressure.2003In: Biorheology, ISSN 0006-355X, E-ISSN 1878-5034, Vol. 40, no 1-3, p. 87-92, article id 12454391Article in journal (Refereed)
    Abstract [en]

    High hydrostatic pressure causes stress response in many types of mammalian cells. We have previously shown that an accumulation of heat shock protein 70 (Hsp70) in a chondrocytic cell line occurred without an activation of the gene itself. Stabilization of the hsp70 mRNA was shown to be the reason for the Hsp70 stress response in the pressurized cells. Since accumulation of Hsp70 in pressurized cells indicated that high hydrostatic pressure induces a stress response without heat shock transcription factor activation, we decided to investigate the activation of two other stress-associated transcription factors, activator protein-1 (AP-1) and nuclear factor-kappaB (NF-kappaB). Induction of Hsp70 in immortalized and primary chondrocytes, murine Neuro-2a neuroblastoma and HeLa cervical carcinoma cell lines was investigated at both mRNA and protein levels. In immortalized chondrocytes and HeLa cells, hsp70 mRNA levels were clearly elevated after 6 hours of the onset of 30 MPa continuous hydrostatic pressure, while in primary chondrocytes and Neuro-2a cells (the cells known to be stress-sensitive) no induction was observed. Surprisingly, neither heat shock nor high hydrostatic pressure could induce the hsp70 mRNA in Neuro-2a cells, although an activation of heat shock transcription factor could be observed in heat-shocked cells. No activation of the AP-1 and NF-kappaB binding to their target DNA sequences could be shown in the immortalized chondrocytes.

  • 426.
    Kaarniranta, Kai
    et al.
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Elo, Mika
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Sironen, Reijo
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Lammi, Mikko
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Goldring, Mary
    Beth Israel Deaconess Medical Center and Harvard Medical School, Boston, MA, USA.
    Eriksson, John
    †Turku Centre for Biotechnology, University of Turku, Åbo Akademi University, Turku, Finland.
    Sistonen, Lea
    †Turku Centre for Biotechnology, University of Turku, Åbo Akademi University, Turku, Finland.
    Helminen, Heikki
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Hsp70 accumulation in chondrocytic cells exposed to high continuous hydrostatic pressure coincides with mRNA stabilization rather than transcriptional activation.1998In: Proceedings of the National Academy of Sciences of the United States of America, ISSN 0027-8424, E-ISSN 1091-6490, Vol. 95, no 5, p. 2319-2324, article id 9482883Article in journal (Refereed)
    Abstract [en]

    In response to various stress stimuli, heat shock genes are induced to express heat shock proteins (Hsps). Previous studies have revealed that expression of heat shock genes is regulated both at transcriptional and posttranscriptional level, and the rapid transcriptional induction of heat shock genes involves activation of the specific transcription factor, heat shock factor 1 (HSF1). Furthermore, the transcriptional induction can vary in intensity and kinetics in a signal- and cell-type-dependent manner. In this study, we demonstrate that mechanical loading in the form of hydrostatic pressure increases heat shock gene expression in human chondrocyte-like cells. The response to continuous high hydrostatic pressure was characterized by elevated mRNA and protein levels of Hsp70, without activation of HSF1 and transcriptional induction of hsp70 gene. The increased expression of Hsp70 was mediated through stabilization of hsp70 mRNA molecules. Interestingly, in contrast to static pressurization, cyclic hydrostatic loading did not result in the induction of heat shock genes. Our findings show that hsp70 gene expression is regulated posttranscriptionally without transcriptional induction in chondrocyte-like cells upon exposure to high continuous hydrostatic pressure. We suggest that the posttranscriptional regulation in the form of hsp70 mRNA stabilization provides an additional mode of heat shock gene regulation that is likely to be of significant importance in certain forms of stress.

  • 427.
    Kaarniranta, Kai
    et al.
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Holmberg, Carina
    Turku Centre for Biotechnology, University of Turku, Åbo Akademi University, Turku, Finland; Department of Biochemistry and Pharmacy, Åbo Akademi University, Turku, Finland.
    Helminen, Heikki
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Eriksson, John
    Turku Centre for Biotechnology, University of Turku, Åbo Akademi University, Turku, Finland; Department of Biology, University of Turku, Turku, Finland.
    Sistonen, Lea
    Turku Centre for Biotechnology, University of Turku, Åbo Akademi University, Turku, Finland.
    Lammi, Mikko
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Protein synthesis is required for stabilization of hsp70 mRNA upon exposure to both hydrostatic pressurization and elevated temperature.2000In: FEBS Letters, ISSN 0014-5793, E-ISSN 1873-3468, Vol. 475, no 3, p. 283-286, article id 10869572Article in journal (Refereed)
    Abstract [en]

    We have recently described that in chondrocytic cells high hydrostatic pressure (HP) causes a heat shock response via mRNA stabilization without a transcriptional activation of the hsp70 gene. In this study, we investigated whether this exceptional regulatory mechanism occurs more generally in different types of cells. Indeed, hsp70 mRNA and protein accumulated in HeLa, HaCat and MG-63 cells under 30 MPa HP, without DNA-binding of heat shock transcription factor 1 (HSF1) to the heat shock element of the hsp70 gene or formation of nuclear HSF1 granules, revealing a lack of transcriptional activation. Moreover, we observed that protein synthesis is needed for mRNA stabilization. Thus, high HP offers a model to study the mechanisms of hsp70 mRNA stabilization without HSF1-mediated induction of the heat shock gene response.

  • 428.
    Kaarniranta, Kai
    et al.
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Holmberg, Carina
    Turku Centre for Biotechnology, University of Turku, Åbo Akademi University, Turku, Finland; Department of Biochemistry and Pharmacy, Åbo Akademi University, Turku, Finland.
    Lammi, Mikko
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Eriksson, John
    Turku Centre for Biotechnology, University of Turku, Åbo Akademi University, Turku, Finland.
    Sistonen, Lea
    Turku Centre for Biotechnology, University of Turku, Åbo Akademi University, Turku, Finland; Department of Biology, Åbo Akademi University, Turku, Finland.
    Helminen, Heikki
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Primary chondrocytes resist hydrostatic pressure-induced stress while primary synovial cells and fibroblasts show modified Hsp70 response.2001In: Osteoarthritis and Cartilage, ISSN 1063-4584, E-ISSN 1522-9653, Vol. 9, no 1, p. 7-13, article id 11178942Article in journal (Refereed)
    Abstract [en]

    OBJECTIVE: During joint loading, chondrocytes in the articular cartilage are subjected to gradients of high compressive hydrostatic pressure (HP). In response to diverse chemical or physical stresses, heat shock genes are induced to express heat shock proteins (Hsps). This study sought to examine the role of Hsps in baroresistance in primary bovine chondrocytes and synovial cells, as well as in primary human fibroblasts.

    METHODS: Northern blotting was used to analyze the steady-state levels of hsp70 mRNA in the primary cells exposed to HP or heat stress. Hsp70 protein accumulation was analyzed by Western blotting, and the DNA-binding activity was examined by gel mobility shift assay.

    RESULTS: Primary bovine chondrocytes which have been adapted to live under pressurized conditions showed negligible Hsp70 response upon HP loading, whereas primary bovine synovial cells and human fibroblasts accumulated hsp70 mRNA and protein when subjected to HP. The response was initiated without activation of the heat shock transcription factor 1. Interestingly, pre-conditioning of the barosensitive fibroblasts with HP or heat shock reduced the Hsp70 response, indicating induction of baroresistance.

    CONCLUSION: This study suggests that Hsp70 can play an important role in the early stages of adaptation of cells to HP. Thus, the Hsp70 gene expression upon HP loading may serve as one indicator of the chondrocytic phenotype of the cells. This can be of use in the treatment of cartilage lesions.

  • 429.
    Kaarniranta, Kai
    et al.
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Oksala, Niku
    Department of Surgery, Kuopio University Hospital, Kuopio, Finland.
    Karjalainen, Hannu
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Suuronen, Tiina
    Department of Neuroscience and Neurology, University of Kuopio, Kuopio, Finland.
    Sistonen, Lea
    Turku Centre for Biotechnology, University of Turku, Åbo Akademi University, Turku, Finland.
    Helminen, Heikki
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Salminen, Antero
    Department of Neurology, Kuopio University Hospital, Kuopio, Finland; Department of Neuroscience and Neurology, University of Kuopio, Kuopio, Finland.
    Lammi, Mikko
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Neuronal cells show regulatory differences in the hsp70 gene response.2002In: Brain Research. Molecular Brain Research, ISSN 0169-328X, E-ISSN 1872-6941, Vol. 101, no 1-2, p. 136-140, article id 12007842Article in journal (Refereed)
    Abstract [en]

    The synthesis of heat shock proteins (Hsps), encoded by heat shock genes, is increased in response to various stress stimuli. Hsps function as molecular chaperones, they dissociate cytotoxic stress-induced protein aggregates within cells and ensure improved survival. Induction of heat shock genes is mainly regulated at the transcriptional level. The stress responsive transcription factor, heat shock factor 1 (HSF1), is involved in the transcriptional induction of the heat shock genes. Our objective was to examine how hsp70 genes are regulated in different transformed and primary neurons upon exposure to elevated temperature. Our findings reveal that the Hsp70 response is regulated at the translational level in Neuro-2a neuroblastoma cells, while the IMR-32 neuroblastoma cells respond to stress by the classical HSF1-driven transcriptional regulatory mechanism. Primary rat hippocampal neurons show a lack of HSF1 and induction of the hsp70 gene. These observations suggest that neuronal cells display different hsp70 gene expression patterns which range from undetected response to transcriptional and posttranscriptional regulation during heat stress.

  • 430.
    Kaarniranta, Kai
    et al.
    Department of Ophthalmology, University of Kuopio, Kuopio, Finland; Department of Ophthalmology, Kuopio University Hospital, Kuopio, Finland.
    Ryhänen, Tuomas
    Department of Ophthalmology, University of Kuopio, Kuopio, Finland.
    Sironen, Reijo
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Suuronen, Tiina
    Department of Neuroscience and Neurology, University of Kuopio, Kuopio, Finland.
    Elo, Mika
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Karjalainen, Hannu
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Lammi, Mikko
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Teräsvirta, Markku
    Department of Ophthalmology, Kuopio University Hospital, Kuopio, Finland.
    Uusitalo, Hannu
    Department of Neuroscience and Neurology, University of Kuopio, Kuopio, Finland; Department of Neurology, Kuopio University Hospital, Kuopio, Finland.
    Salminen, Antero
    Department of Neuroscience and Neurology, University of Kuopio, Kuopio, Finland; Department of Neurology, Kuopio University Hospital, Kuopio, Finland.
    Geldanamycin activates Hsp70 response and attenuates okadaic acid-induced cytotoxicity in human retinal pigment epithelial cells.2005In: Brain Research. Molecular Brain Research, ISSN 0169-328X, E-ISSN 1872-6941, Vol. 137, no 1-2, p. 126-131, article id 15950770Article in journal (Refereed)
    Abstract [en]

    Reversible protein phosphorylation regulates the biological activities of many human proteins involved in crucial cellular processes, e.g., protein-protein interactions, cell signaling, gene transcription, cell growth, and death. A malfunction of cellular homeostasis in retinal pigment epithelial (RPE) cells is involved in the age-related retinal degeneration. In this study, we examined cytotoxicity in human RPE cells subjected to the protein phosphatase inhibitor, okadaic acid (OA). Moreover, the influence of Hsp90 inhibitor geldanamycin (GA), a benzoquinone ansamycin, in cytoprotection was assessed. Hsp70 protein levels were analyzed by Western blot. Cellular viability was determined by LDH and MTT assays. To study apoptotic cell death, caspase-3 enzyme activity was measured by assaying the cleavage of a fluorescent peptide substrate and Hoechst dye was used to visualize nuclear morphology. OA treatment caused morphological changes and induced cytotoxicity by caspase-3-independent manner in the RPE cells. No evidence of nuclear fragmentation was observed in response to OA. Interestingly, GA treatment accumulated Hsp70 protein and attenuated OA-induced cytotoxicity. This study suggests that Hsp70 and Hsp90 are closely related to cytoprotection of RPE cells in response to protein phosphatase inhibition.

  • 431.
    Kaati, Gunnar
    et al.
    Umeå University, Faculty of Medicine, Department of Public Health and Clinical Medicine. Department of Biosciences and Nutrition, Karolinska Institutet, Stockholm, Sweden.
    Bygren, Lars Olov
    Umeå University, Faculty of Medicine, Department of Community Medicine and Rehabilitation. Department of Biosciences and Nutrition, Karolinska Institutet, Stockholm, Sweden.
    Pembrey, Marcus
    Sjöstrom, Michael
    Transgenerational response to nutrition, early life circumstances and longevity2007In: European Journal of Human Genetics, ISSN 1018-4813, E-ISSN 1476-5438, Vol. 15, no 7, p. 784-790Article in journal (Refereed)
    Abstract [en]

    Nutrition might induce, at some loci, epigenetic or other changes that could be transmitted to the next generation impacting on health. The slow growth period (SGP) before the prepubertal peak in growth velocity has emerged as a sensitive period where different food availability is followed by different transgenerational response (TGR). The aim of this study is to investigate to what extent the probands own childhood circumstances are in fact the determinants of the findings. In the analysis, data from three random samples, comprising 271 probands and their 1626 parents and grandparents, left after exclusions because of missing data, were utilized. The availability of food during any given year was classified based on regional statistics. The ancestors' SGP was set at the ages of 8-12 years and the availability of food during these years classified as good, intermediate or poor. The probands' childhood circumstances were defined by the father's ownership of land, the number of siblings and order in the sibship, the death of parents and the parents' level of literacy. An earlier finding of a sex-specific influence from the ancestors' nutrition during the SGP, going from the paternal grandmother to the female proband and from the paternal grandfather to the male proband, was confirmed. In addition, a response from father to son emerged when childhood social circumstances of the son were accounted for. Early social circumstances influenced longevity for the male proband. TGRs to ancestors' nutrition prevailed as the main influence on longevity.

  • 432.
    Kahn, Tatiana
    et al.
    Institute of Gene Biology, RAS, Russia.
    Savitsky, Mikhail
    Institute of Gene Biology, RAS, Russia.
    Georgiev, Pavel
    Institute of Gene Biology, RAS, Russia.
    Attachment of HeT-A sequences to chromosomal termini in Drosophila melanogaster may occur by different mechanisms2000In: Molecular and Cellular Biology, ISSN 0270-7306, E-ISSN 1098-5549, Vol. 20, no 20, p. 7634-7642Article in journal (Refereed)
    Abstract [en]

    Drosophila telomeres contain arrays of the retrotransposonlike elements HeT-A and TART. Their transposition to broken chromosomal termini has been implicated in chromosome healing and telomere elongation. The HeT-A element is attached by its 3' end, which contains the promoter. To monitor the behavior of HeT-A elements, we used the yellow gene with terminal deficiencies consisting of breaks in the yellow promoter region that result in the y-null phenotype. Attachment of the HeT-A element provides the promoterless yellow gene with a promoter that activates yellow expression in bristles. The frequency of HeT-A transpositions to the yellow terminal deficiency depends on the genotype of the line and varies from 2 x 10(-3) to less than 2 x 10(-5). Loss of the attached HeT-A due to incomplete replication at the telomere leads to inactivation of yellow expression, which is restored by attachment of a new HeT-A element upstream of yellow. New HeT-A additions occur at a frequency of about 1.2 x 10(-3). Short DNA attachments are generated by gene conversion using the homologous telomeric sequences as templates. Longer DNA attachments are generated either by conventional transposition of an HeT-A element to the chromosomal terminus or by recombination between the 3' terminus of telomeric HeT-A elements and the receding end of HeT-A attached to the yellow gene.

  • 433.
    Kahn, Tatyana G.
    et al.
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Dorafshan, Eshagh
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Schultheis, Dorothea
    Zare, Aman
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Stenberg, Per
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology). Division of CBRN Defense and Security, Swedish Defense Research Agency, FOI, Umea, 906 21, Sweden.
    Reim, Ingolf
    Pirrotta, Vincenzo
    Schwartz, Yuri B.
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Interdependence of PRC1 and PRC2 for recruitment to Polycomb Response Elements2016In: Nucleic Acids Research, ISSN 0305-1048, E-ISSN 1362-4962, Vol. 44, no 21, p. 10132-10149Article in journal (Refereed)
    Abstract [en]

    Polycomb Group (PcG) proteins are epigenetic repressors essential for control of development and cell differentiation. They form multiple complexes of which PRC1 and PRC2 are evolutionary conserved and obligatory for repression. The targeting of PRC1 and PRC2 is poorly understood and was proposed to be hierarchical and involve tri-methylation of histone H3 (H3K27me3) and/or monoubiquitylation of histone H2A (H2AK118ub). Here, we present a strict test of this hypothesis using the Drosophila model. We discover that neither H3K27me3 nor H2AK118ub is required for targeting PRC complexes to Polycomb Response Elements (PREs). We find that PRC1 can bind PREs in the absence of PRC2 but at many PREs PRC2 requires PRC1 to be targeted. We show that one role of H3K27me3 is to allow PcG complexes anchored at PREs to interact with surrounding chromatin. In contrast, the bulk of H2AK118ub is unrelated to PcG repression. These findings radically change our view of how PcG repression is targeted and suggest that PRC1 and PRC2 can communicate independently of histone modifications.

  • 434.
    Kahra, Dana
    et al.
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Mondol, Tanumoy
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Niemiec, Moritz S
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Wittung-Stafshede, Pernilla
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Human Copper Chaperone Atox1 Translocates to the Nucleus but does not Bind DNA In Vitro2015In: Protein peptide letters, ISSN 0929-8665, E-ISSN 1875-5305, Vol. 22, no 6, p. 532-538Article in journal (Refereed)
    Abstract [en]

    After Ctr1-mediated cell uptake, copper (Cu) is transported by the cytoplasmic Cu chaperone Atox1 to P1B type ATPases ATP7A and ATP7B in the Golgi network, for incorporation into Cudependent enzymes. Atox1 is a small 68-residue protein that binds Cu in a conserved CXXC motif; it delivers Cu to target domains in ATP7A/B via direct protein-protein interactions. Specific transcription factors regulating expression of the human Cu transport proteins have not been reported although Atox1 was recently suggested to have dual functionality such that it, in addition to its cytoplasmic chaperone function, acts as a transcription factor in the nucleus. To examine this hypothesis, here we investigated the localization of Atox1 in HeLa cells using fluorescence imaging in combination with in vitro binding experiments to fluorescently labeled DNA duplexes harboring the proposed promotor sequence. We found that whereas Atox1 is present in the nucleus in HeLa cells, it does not bind to DNA in vitro. It appears that Atox1 mediates transcriptional regulation via additional (unknown) proteins.

  • 435. Kaiser, Nadine
    et al.
    Corkery, Dale
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Wu, Yao-Wen
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Laraia, Luca
    Waldmann, Herbert
    Modulation of autophagy by the novel mitochondrial complex I inhibitor Authipyrin2019In: Bioorganic & Medicinal Chemistry, ISSN 0968-0896, E-ISSN 1464-3391, Vol. 27, no 12, p. 2444-2448Article in journal (Refereed)
    Abstract [en]

    Autophagy ensures cellular homeostasis by the degradation of long-lived proteins, damaged organelles and pathogens. This catabolic process provides essential cellular building blocks upon nutrient deprivation. Cellular metabolism, especially mitochondrial respiration, has a significant influence on autophagic flux, and complex I function is required for maximal autophagy. In Parkinson’s disease mitochondrial function is frequently impaired and autophagic flux is altered. Thus, dysfunctional organelles and protein aggregates accumulate and cause cellular damage. In order to investigate the interdependency between mitochondrial function and autophagy, novel tool compounds are required. Herein, we report the discovery of a structurally novel autophagy inhibitor (Authipyrin) using a high content screening approach. Target identification and validation led to the discovery that Authipyrin targets mitochondrial complex I directly, leading to the potent inhibition of mitochondrial respiration as well as autophagy.

    The full text will be freely available from 2021-03-31 14:28
  • 436.
    Kallioniemi, Antti
    et al.
    Department of Physics, University of Kuopio, Kuopio, Finland.
    Jurvelin, Jukka
    Department of Physics, University of Kuopio, Kuopio, Finland; Department of Clinical Physiology and Nuclear Medicine, Kuopio University Hospital, Kuopio, Finland.
    Nieminen, Miika
    Department of Diagnostic Radiology, Oulu University Hospital, Oulu, Finland .
    Lammi, Mikko
    Department of Anatomy, Institute of Biomedicine, University of Kuopio, Kuopio, Finland .
    Töyräs, Juha
    Department of Clinical Neurophysiology, Kuopio University Hospital, Kuopio, Finland.
    Contrast agent enhanced pQCT of articular cartilage.2007In: Physics in Medicine and Biology, ISSN 0031-9155, E-ISSN 1361-6560, Vol. 52, no 4, p. 1209-1219, article id 17264381Article in journal (Refereed)
    Abstract [en]

    The delayed gadolinium enhanced MRI of cartilage (dGEMRIC) technique is the only non-invasive means to estimate proteoglycan (PG) content in articular cartilage. In dGEMRIC, the anionic paramagnetic contrast agent gadopentetate distributes in inverse relation to negatively charged PGs, leading to a linear relation between T1,Gd and spatial PG content in tissue. In the present study, for the first time, contrast agent enhanced peripheral quantitative computed tomography (pQCT) was applied, analogously to dGEMRIC, for the quantitative detection of spatial PG content in cartilage. The suitability of two anionic radiographic contrast agents, gadopentetate and ioxaglate, to detect enzymatically induced PG depletion in articular cartilage was investigated. First, the interrelationships of x-ray absorption, as measured with pQCT, and the contrast agent solution concentration were investigated. Optimal contrast agent concentrations for the following experiments were selected. Second, diffusion rates for both contrast agents were investigated in intact (n=3) and trypsin-degraded (n=3) bovine patellar cartilage. The contrast agent concentration of the cartilaginous layer was measured prior to and 2-27 h after immersion. Optimal immersion time for the further experiments was selected. Third, the suitability of gadopentetate and ioxaglate enhanced pQCT to detect the enzymatically induced specific PG depletion was investigated by determining the contrast agent concentrations and uronic acid and water contents in digested and intact osteochondral samples (n=16). After trypsin-induced PG loss (-70%, p<0.05) the penetration of gadopentetate and ioxaglate increased (p<0.05) by 34% and 48%, respectively. Gadopentetate and ioxaglate concentrations both showed strong correlation (r=-0.95, r=-0.94, p<0.01, respectively) with the uronic acid content. To conclude, contrast agent enhanced pQCT provides a technique to quantify PG content in normal and experimentally degraded articular cartilage in vitro. As high resolution imaging of e.g. the knee joint is possible with pQCT, the present technique may be further developed for in vivo quantification of PG depletion in osteoarthritic cartilage. However, careful in vitro and in vivo characterization of diffusion mechanics and optimal contrast agent concentrations are needed before diagnostic applications are feasible.

  • 437. Kaneko, H
    et al.
    Mehrotra, M
    Alander, C
    Lerner, Ulf
    Umeå University, Faculty of Medicine, Department of Odontology, Oral Cell Biology.
    Pilbeam, C
    Raisz, L
    Effects of prostaglandin E-2 and lipopolysaccharide on osteoclastogenesis in RAW 264.7 cells2007In: Prostaglandins, Leukotrienes and Essential Fatty Acids, ISSN 0952-3278, E-ISSN 1532-2823, Vol. 77, no 3-4, p. 181-186Article in journal (Refereed)
    Abstract [en]

    Introduction: Prostaglandins (PGs) can act on both hematopoietic and osteoblastic lineages to enhance osteoclast formation.

    Methods: We examined PGE(2) stimulated osteoclastogenesis in RAW 264.7 cells and the role of endogenous PGE2 in lipopolysaccharide (LPS) stimulated osteoclastogenesis.

    Results: RANKL (1-100ng/ml) increased formation of osteoclasts, defined as tartrate resistant acid phosphatase multinucleated cells, with peak effects at 30 ng/ml. Addition of PGE2 (0-01-1.0 mu M) to RANKL (30 ng/ml) dose dependently increased osteoclast number 30-150%. Use of NS-398 (0.1 mu M) or indomethacin (Indo, 1.0 mu M) to block endogenous PG synthesis had little effect on the response to RANKL alone but significantly decreased the response to PGE2. Addition of LPS (100 ng/ml) to RANKL increased osteoclast number 50%, and this response was significantly decreased by NS-398 and Indo. RANKL and PGE2 produced small, additive increases in COX-2 mRNA levels, while LPS produced a larger increase. PG release into the medium was not increased by RANKL and PGE2 but markedly increased by LPS.

    Conclusion: We conclude that RANKL stimulated osteoclastogenesis can be enhanced by PGE2 and LPS though direct effects on the hematopoietic cell lineage and that these effects may be mediated in part by induction of COX-2 and enhanced intracellular PG production.

  • 438. Karakostis, Konstantinos
    et al.
    Gnanasundram, Sivakumar Vadivel
    Lopez, Ignacio
    Thermou, Aikaterini
    Wang, Lixiao
    Umeå University, Faculty of Medicine, Department of Medical Biosciences.
    Nylander, Karin
    Olivares-Illana, Vanesa
    Fåhraeus, Robin
    Umeå University, Faculty of Medicine, Department of Medical Biosciences. 1 E´quipe Labellise´e Ligue Contre le Cancer, Universite´ Paris 7, 27 Rue Juliette Dodu, Paris, France; RECAMO, Masaryk Memorial Cancer Institute, Zluty kopec 7, Brno, Czech Republic.
    A single synonymous mutation determines the phosphorylation and stability of the nascent protein2019In: Journal of Molecular Cell Biology, ISSN 1674-2788, E-ISSN 1759-4685, Vol. 11, no 3, p. 187-199Article in journal (Refereed)
    Abstract [en]

    p53 is an intrinsically disordered protein with a large number of post-translational modifications and interacting partners. The hierarchical order and subcellular location of these events are still poorly understood. The activation of p53 during the DNA damage response (DDR) requires a switch in the activity of the E3 ubiquitin ligase MDM2 from a negative to a positive regulator of p53. This is mediated by the ATM kinase that regulates the binding of MDM2 to the p53 mRNA facilitating an increase in p53 synthesis. Here we show that the binding of MDM2 to the p53 mRNA brings ATM to the p53 polysome where it phosphorylates the nascent p53 at serine 15 and prevents MDM2-mediated degradation of p53. A single synonymous mutation in p53 codon 22 (L22L) prevents the phosphorylation of the nascent p53 protein and the stabilization of p53 following genotoxic stress. The ATM trafficking from the nucleus to the p53 polysome is mediated by MDM2, which requires its interaction with the ribosomal proteins RPL5 and RPL11. These results show how the ATM kinase phosphorylates the p53 protein while it is being synthesized and offer a novel mechanism whereby a single synonymous mutation controls the stability and activity of the encoded protein.

  • 439. Karakostis, Konstantinos
    et al.
    Ponnuswamy, Anand
    Fusee, Leila T. S.
    Bailly, Xavier
    Laguerre, Laurent
    Worall, Erin
    Vojtesek, Borek
    Nylander, Karin
    Umeå University, Faculty of Medicine, Department of Medical Biosciences.
    Fåhraeus, Robin
    Umeå University, Faculty of Medicine, Department of Medical Biosciences. E´quipe Labellise´e Ligue Contre le Cancer, Universite´ Paris 7, INSERM UMR 1162, Paris, France; Regional Centre for Applied Molecular Oncology, RECAMO and Masaryk Memorial Cancer Institute, Brno, Czech Republic.
    p53 mRNA and p53 Protein Structures Have Evolved Independently to Interact with MDM22016In: Molecular biology and evolution, ISSN 0737-4038, E-ISSN 1537-1719, Vol. 33, no 5, p. 1280-1292Article in journal (Refereed)
    Abstract [en]

    The p53 tumor suppressor and its key regulator MDM2 play essential roles in development, ageing, cancer, and cellular stress responses in mammals. Following DNA damage, MDM2 interacts with p53 mRNA in an ATM kinase-dependent fashion and stimulates p53 synthesis, whereas under normal conditions, MDM2 targets the p53 protein for degradation. The peptide-and RNA motifs that interact with MDM2 are encoded by the same conserved BOX-I sequence, but how these interactions have evolved is unknown. Here, we show that a temperature-sensitive structure in the invertebrate Ciona intestinalis (Ci) p53 mRNA controls its interaction with MDM2. We also show that a nonconserved flanking region of Ci-BOX-I domain prevents the p53-MDM2 protein-protein interaction. These results indicate that the temperature-regulated p53 mRNA-MDM2 interaction evolved to become kinase regulated in the mammalian DNA damage response. The data also suggest that the negative regulation of p53 by MDM2 via protein-protein interaction evolved in vertebrates following changes in the BOX-I flanking sequence.

  • 440. Karhula, Sakari
    et al.
    Finnilä, Mikko
    Lammi, Mikko
    Umeå University, Faculty of Medicine, Department of Integrative Medical Biology (IMB). School of Public Health, Health Science Center of Xi'an Jiaotong University; Key Laboratory of Trace Elements and Endemic Diseases, National Health and Family Planning Commission, P. R. China.
    Ylärinne, Janne
    Umeå University, Faculty of Medicine, Department of Integrative Medical Biology (IMB).
    Kauppinen, Sami
    Rieppo, Lassi
    Pritzker, Kenneth P H
    Nieminen, Heikki
    Saarakkala, Simo
    Effects of articular cartilage constituents on phosphotungstic acid enhanced micro-computed tomography2017In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 12, no 1, article id e0171075Article in journal (Refereed)
    Abstract [en]

    Contrast-enhanced micro-computed tomography (CEμCT) with phosphotungstic acid (PTA) has shown potential for detecting collagen distribution of articular cartilage. However, the selectivity of the PTA staining to articular cartilage constituents remains to be elucidated. The aim of this study was to investigate the dependence of PTA for the collagen content in bovine articular cartilage. Adjacent bovine articular cartilage samples were treated with chondroitinase ABC and collagenase to degrade the proteoglycan and the collagen constituents in articular cartilage, respectively. Enzymatically degraded samples were compared to the untreated samples using CEμCT and reference methods, such as Fourier-transform infrared imaging. Decrease in the X-ray attenuation of PTA in articular cartilage and collagen content was observed in cartilage depth of 0-13% and deeper in tissue after collagen degradation. Increase in the X-ray attenuation of PTA was observed in the cartilage depth of 13-39% after proteoglycan degradation. The X-ray attenuation of PTA-labelled articular cartilage in CEμCT is associated mainly with collagen content but the proteoglycans have a minor effect on the X-ray attenuation of the PTA-labelled articular cartilage. In conclusion, the PTA labeling provides a feasible CEμCT method for 3D characterization of articular cartilage.

  • 441.
    Karjalainen, Hannu
    et al.
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Sironen, Reijo
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Elo, Mika
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Kaarniranta, Kai
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Takigawa, Masaharu
    bDepartment of Biochemistry and Molecular Dentistry, Okayama University Graduate School of Medicine and Dentistry, Japan.
    Helminen, Heikki
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Lammi, Mikko
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Gene expression profiles in chondrosarcoma cells subjected to cyclic stretching and hydrostatic pressure. A cDNA array study.2003In: Biorheology, ISSN 0006-355X, E-ISSN 1878-5034, Vol. 40, no 1-3, p. 93-100, article id 12454392Article in journal (Refereed)
    Abstract [en]

    Mechanical forces have a profound effect on cartilage tissue and chondrocyte metabolism. Strenuous loading inhibits the cellular metabolism, while optimal level of loading at correct frequency raises an anabolic response in chondrocytes. In this study, we used Atlas Human Cancer cDNA array to investigate mRNA expression profiles in human chondrosarcoma cells stretched 8% for 6 hours at a frequency of 0.5 Hz. In addition, cultures were exposed to continuous and cyclic (0.5 Hz) 5 MPa hydrostatic pressure. Cyclic stretch had a more profound effect on the gene expression profiles than 5 MPa hydrostatic pressure. Several genes involved with the regulation of cell cycle were increased in stretched cells, as well as mRNAs for PDGF-B, glucose-1-phosphate uridylyltransferase, Tiam1, cdc37 homolog, Gem, integrin alpha6, and matrix metalloproteinase-3. Among down-regulated genes were plakoglobin, TGF-alpha, retinoic acid receptor-alpha and Wnt8b. A smaller number of changes was detected after pressure treatments. Plakoglobin was increased under cyclic and continuous 5 MPa hydrostatic pressure, while mitogen-activated protein kinase-9, proliferating cell nuclear antigen, Rad6, CD9 antigen, integrins alphaE and beta8, and vimentin were decreased. Cyclic and continuous pressurization induces a number of specific changes. In conclusion, a different set of genes were affected by three different types of mechanical stimuli applied on chondrosarcoma cells.

  • 442. Karlberg, Tobias
    et al.
    Hornyak, Peter
    Pinto, Ana Filipa
    Milanova, Stefina
    Ebrahimi, Mahsa
    Lindberg, Mikael J.
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Püllen, Nikolai
    Nordström, Axel
    Löverli, Elinor
    Caraballo, Remi
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Wong, Emily V.
    Näreoja, Katja
    Thorsell, Ann-Gerd
    Elofsson, Mikael
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    De la Cruz, Enrique M.
    Björkegren, Camilla
    Schüler, Herwig
    14-3-3 proteins activate Pseudomonas exotoxins-S and -T by chaperoning a hydrophobic surface2018In: Nature Communications, ISSN 2041-1723, E-ISSN 2041-1723, Vol. 9, article id 3785Article in journal (Refereed)
    Abstract [en]

    Pseudomonas are a common cause of hospital-acquired infections that may be lethal. ADP-ribosyltransferase activities of Pseudomonas exotoxin-S and -T depend on 14-3-3 proteins inside the host cell. By binding in the 14-3-3 phosphopeptide binding groove, an amphipathic C-terminal helix of ExoS and ExoT has been thought to be crucial for their activation. However, crystal structures of the 14-3-3 beta: ExoS and -ExoT complexes presented here reveal an extensive hydrophobic interface that is sufficient for complex formation and toxin activation. We show that C-terminally truncated ExoS ADP-ribosyltransferase domain lacking the amphipathic binding motif is active when co-expressed with 14-3-3. Moreover, swapping the amphipathic C-terminus with a fragment from Vibrio Vis toxin creates a 14-3-3 independent toxin that ADP-ribosylates known ExoS targets. Finally, we show that 14-3-3 stabilizes ExoS against thermal aggregation. Together, this indicates that 14-3-3 proteins activate exotoxin ADP-ribosyltransferase domains by chaperoning their hydrophobic surfaces independently of the amphipathic C-terminal segment.

  • 443. Karlberg, Tobias
    et al.
    Klepsch, Mirjam
    Thorsell, Ann-Gerd
    Andersson, C. David
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Linusson, Anna
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Schuler, Herwig
    Structural Basis for Lack of ADP-ribosyltransferase Activity in Poly(ADP-ribose) Polymerase-13/Zinc Finger Antiviral Protein2015In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 290, no 12, p. 7336-7344Article in journal (Refereed)
    Abstract [en]

    Background: PARP13 contains a divergent PARP homology ADP-ribosyltransferase domain of unknown function. Results: The consensus NAD(+) pocket of PARP13 is occluded by interacting protein side chains. Conclusion: PARP13 lacks the structural requirements for NAD(+) binding. Significance: Evolutionary conservation of enzymatic inactivity suggests a need for a rigid domain structure.

    The mammalian poly(ADP-ribose) polymerase (PARP) family includes ADP-ribosyltransferases with diphtheria toxin homology (ARTD). Most members have mono-ADP-ribosyltransferase activity. PARP13/ARTD13, also called zinc finger antiviral protein, has roles in viral immunity and microRNA-mediated stress responses. PARP13 features a divergent PARP homology domain missing a PARP consensus sequence motif; the domain has enigmatic functions and apparently lacks catalytic activity. We used x-ray crystallography, molecular dynamics simulations, and biochemical analyses to investigate the structural requirements for ADP-ribosyltransferase activity in human PARP13 and two of its functional partners in stress granules: PARP12/ARTD12, and PARP15/BAL3/ARTD7. The crystal structure of the PARP homology domain of PARP13 shows obstruction of the canonical active site, precluding NAD(+) binding. Molecular dynamics simulations indicate that this closed cleft conformation is maintained in solution. Introducing consensus side chains in PARP13 did not result in 3-aminobenzamide binding, but in further closure of the site. Three-dimensional alignment of the PARP homology domains of PARP13, PARP12, and PARP15 illustrates placement of PARP13 residues that deviate from the PARP family consensus. Introducing either one of two of these side chains into the corresponding positions in PARP15 abolished PARP15 ADP-ribosyltransferase activity. Taken together, our results show that PARP13 lacks the structural requirements for ADP-ribosyltransferase activity.

  • 444.
    Karlsborn, Tony
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Physiological consequences of Elongator complex inactivation in Eukaryotes2016Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Mutations found in genes encoding human Elongator complex subunits have been linked to neurodevelopmental disorders such as familial dysautonomia (FD), rolandic epilepsy and amyotrophic lateral sclerosis. In addition, loss-of-function mutations in genes encoding Elongator complex subunits cause defects in neurodevelopment and reduced neuronal function in both mice and nematodes. The Elongator complex is a conserved protein complex comprising six subunits (Elp1p-Elp6p) found in eukaryotes. The primary function of this complex in yeast is formation of the 5-methoxycarbonylmethyl (mcm5) and 5-carbamoylmethyl (ncm5) side chains found on wobble uridines (U34) in tRNAs. The aim of this thesis is to investigate the physiological consequences of Elongator complex inactivation in humans and in the yeast Saccharomyces cerevisiae.

    Inactivation of the Elongator complex causes widespread defects in a multitude of different cellular processes in S. cerevisiae. Thus, we investigated metabolic alterations resulting from Elongator complex inactivation. We show that deletion of the S. cerevisiae ELP3 gene leads to widespread metabolic alterations. Moreover, all global metabolic alterations observed in the elp3Δ strain are not restored in the presence of elevated levels of hypomodified tRNAs that normally have the modified nucleoside mcm5s2U. Collectively, we show that modified wobble nucleosides in tRNAs are required for metabolic homeostasis.

    Elongator mutants display sensitivity to DNA damage agents, but the underlying mechanism explaining this sensitivity remains elusive. We demonstrate that deletion of the S. cerevisiae ELP3 gene results in post-transcriptional reduction of Ixr1p levels. Further, we show that the reduced Ixr1p levels prevent adequate Rnr1p levels upon treatment with DNA damage agents. These findings suggest that reduced Ixr1p levels could in part explain why Elongator mutants are sensitive to DNA damage agents.

    Depletion of Elongator complex subunits results in loss of wobble uridine modifications in plants, nematodes, mice and yeast. Therefore, we investigated whether patients with the neurodegenerative disease familial dysautonomia (FD), who have lower levels of the ELP1 protein, display reduced amounts of modified wobble uridine nucleosides. We show that tRNA isolated from brain tissue and fibroblast cell lines derived from FD patients have 64–71% of the mcm5s2U nucleoside levels observed in total tRNA from non-FD brain tissue and non-FD fibroblasts. Overall, these results suggest that the cause for the neurodegenerative nature of FD could be translation impairment caused by reduced levels of modified wobble uridine nucleosides in tRNAs. Thus, our results give new insight on the importance of modified wobble uridine nucleosides for neurodevelopment.

  • 445.
    Karlsborn, Tony
    et al.
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Mahmud, A K M Firoj
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Tükenmez, Hasan
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Byström, Anders S.
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Loss of ncm5 and mcm5 wobble uridine side chains results in an altered metabolic profile2016In: Metabolomics, ISSN 1573-3882, E-ISSN 1573-3890, Vol. 12, no 12, article id 177Article in journal (Refereed)
    Abstract [en]

    Introduction: The Elongator complex, comprising six subunits (Elp1p-Elp6p), is required for formation of 5-carbamoylmethyl (ncm(5)) and 5-methoxycarbonylmethyl (mcm(5)) side chains on wobble uridines in 11 out of 42 tRNA species in Saccharomyces cerevisiae. Loss of these side chains reduces the efficiency of tRNA decoding during translation, resulting in pleiotropic phenotypes. Overexpression of hypomodified tRNA(s2UUU)(Lys); tRNA(s2UUG)(Gln) and tRNA(s2UUC)(Glu), which in wild-type strains are modified with mcm(5)s(2)U, partially suppress phenotypes of an elp3 Delta strain. Objectives: To identify metabolic alterations in an elp3 Delta strain and elucidate whether these metabolic alterations are suppressed by overexpression of hypomodified tRNA(s2UUU)(Lys); tRNA(s2UUG)(Gln) and tRNA(s2UUC)(Glu). Method: Metabolic profiles were obtained using untargeted GC-TOF-MS of a temperature-sensitive elp3 Delta strain carrying either an empty low-copy vector, an empty high-copy vector, a low-copy vector harboring the wild-type ELP3 gene, or a high-copy vector overexpressing tRNA(s2UUU)(Lys); tRNA(s2UUG)(Gln) and tRNA(s2UUC)(Glu). The temperature sensitive elp3 Delta strain derivatives were cultivated at permissive (30 degrees C) or semi-permissive (34 degrees C) growth conditions. Results: Culturing an elp3 Delta strain at 30 or 34 degrees C resulted in altered metabolism of 36 and 46 %, respectively, of all metabolites detected when compared to an elp3D strain carrying the wild-type ELP3 gene. Overexpression of hypomodified tRNA(s2UUU)(Lys); tRNA(s2UUG)(Gln) and tRNA(s2UUC)(Glu) suppressed a subset of the metabolic alterations observed in the elp3 Delta strain. Conclusion: Our results suggest that the presence of ncm(5)- and mcm(5)-side chains on wobble uridines in tRNA are important for metabolic homeostasis.

  • 446.
    Karlsborn, Tony
    et al.
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Tukenmez, Hasan
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Mahmud, A. K. M. Firoj
    Xu, Fu
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Xu, Hao
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Byström, Anders S
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Elongator, a conserved complex required for wobble uridine modifications in Eukaryotes2014In: RNA Biology, ISSN 1547-6286, E-ISSN 1555-8584, Vol. 11, no 12, p. 1519-1528Article in journal (Refereed)
    Abstract [en]

    Elongator is a 6 subunit protein complex highly conserved in eukaryotes. The role of this complex has been controversial as the pleiotropic phenotypes of Elongator mutants have implicated the complex in several cellular processes. However, in yeast there is convincing evidence that the primary and probably only role of this complex is in formation of the 5-methoxycarbonylmethyl (mcm(5)) and 5-carbamoylmethyl (ncm(5)) side chains on uridines at wobble position in tRNA. In this review we summarize the cellular processes that have been linked to the Elongator complex and discuss its role in tRNA modification and regulation of translation. We also describe additional gene products essential for formation of ncm(5) and mcm(5) side chains at U-34 and their influence on Elongator activity.

  • 447. Karlsson, E.
    et al.
    Larkeryd, A.
    Sjödin, Andreas
    Umeå University, Faculty of Science and Technology, Department of Chemistry. Swedish Defence Research Agency, Umeå, Sweden.
    Forsman, M.
    Stenberg, Per
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine). Swedish Defence Research Agency, Umeå, Sweden; Department of Chemistry, Computational Life Science Cluster (CLiC), Umeå University, Umeå, Sweden.
    Scaffolding of a bacterial genome using MinION nanopore sequencing2015In: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 5, article id 11996Article in journal (Refereed)
    Abstract [en]

    Second generation sequencing has revolutionized genomic studies. However, most genomes contain repeated DNA elements that are longer than the read lengths achievable with typical sequencers, so the genomic order of several generated contigs cannot be easily resolved. A new generation of sequencers offering substantially longer reads is emerging, notably the Pacific Biosciences (PacBio) RS II system and the MinION system, released in early 2014 by Oxford Nanopore Technologies through an early access program. The latter has highly advantageous portability and sequences samples by measuring changes in ionic current when single-stranded DNA molecules are translocated through nanopores. We show that the MinION system produces long reads with high mapability that can be used for scaffolding bacterial genomes, despite currently producing substantially higher error rates than PacBio reads. With further development we anticipate that MinION will be useful not only for assembling genomes, but also for rapid detection of organisms, potentially in the field.

  • 448.
    Karlsson, Jessica
    et al.
    Umeå University, Faculty of Medicine, Department of Pharmacology and Clinical Neuroscience, Pharmacology.
    Morgillo, Carmine M.
    Deplano, Alessandro
    Smaldone, Giovanni
    Pedone, Emilia
    Javier Luque, F.
    Svensson, Mona
    Umeå University, Faculty of Medicine, Department of Pharmacology and Clinical Neuroscience, Pharmacology.
    Novellino, Ettore
    Congiu, Cenzo
    Onnis, Valentina
    Catalanotti, Bruno
    Fowler, Christopher J.
    Umeå University, Faculty of Medicine, Department of Pharmacology and Clinical Neuroscience, Pharmacology.
    Interaction of the N-(3-Methylpyridin-2-yl) amide Derivatives of Flurbiprofen and Ibuprofen with FAAH: Enantiomeric Selectivity and Binding Mode2015In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 10, no 11, article id e0142711Article in journal (Refereed)
    Abstract [en]

    Background Combined fatty acid amide hydrolase (FAAH) and cyclooxygenase (COX) inhibition is a promising approach for pain-relief. The Flu-AM1 and Ibu-AM5 derivatives of flurbiprofen and ibuprofen retain similar COX-inhibitory properties and are more potent inhibitors of FAAH than the parent compounds. However, little is known as to the nature of their interaction with FAAH, or to the importance of their chirality. This has been explored here. Methodology/Principal Findings FAAH inhibitory activity was measured in rat brain homogenates and in lysates expressing either wild-type or FAAH(T488A)-mutated enzyme. Molecular modelling was undertaken using both docking and molecular dynamics. The (R)-and (S)-enantiomers of Flu-AM1 inhibited rat FAAH with similar potencies (IC50 values of 0.74 and 0.99 mu M, respectively), whereas the (S)-enantiomer of Ibu-AM5 (IC50 0.59 mu M) was more potent than the (R)-enantiomer (IC50 5.7 mu M). Multiple inhibition experiments indicated that both (R)-Flu-AM1 and (S)-Ibu-AM5 inhibited FAAH in a manner mutually exclusive to carprofen. Computational studies indicated that the binding site for the Flu-AM1 and Ibu-AM5 enantiomers was located between the acyl chain binding channel and the membrane access channel, in a site overlapping the carprofen binding site, and showed a binding mode in line with that proposed for carprofen and other non-covalent ligands. The potency of (R)-Flu-AM1 was lower towards lysates expressing FAAH mutated at the proposed carprofen binding area than in lysates expressing wild-type FAAH. Conclusions/Significance The study provides kinetic and structural evidence that the enantiomers of Flu-AM1 and Ibu-AM5 bind in the substrate channel of FAAH. This information will be useful in aiding the design of novel dual-action FAAH: COX inhibitors.

  • 449.
    Karppinen, Jaro
    et al.
    Department of Physical Medicine and Rehabilitation, University of Oulu, Oulu, Finland.
    Inkinen, Ritva
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Kääpä, Eeva
    Department of Physical Medicine and Rehabilitation, University of Oulu, Oulu, Finland.
    Lammi, Mikko
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Tammi, Markku
    Department of Anatomy, University of Kuopio, Kuopio, Finland.
    Holm, Sten
    Department of Orthopaedics, Sahlgren Hospital, University of Göteborg, Göteborg, Sweden.
    Vanharanta, Hannu
    Department of Physical Medicine and Rehabilitation, University of Oulu, Oulu, Finland.
    Effects of tiaprofenic acid and indomethacin on proteoglycans in the degenerating porcine intervertebral disc.1995In: Spine, ISSN 0362-2436, E-ISSN 1528-1159, Vol. 20, no 10, p. 1170-1177, article id 7638661Article in journal (Refereed)
    Abstract [en]

    STUDY DESIGN: Eighteen pigs were stabbed with a scalpel in the anterior part of the anulus fibrosus of a lumbar disc. After surgery, the pigs received either tiaprofenic acid or indomethacin daily, and a third group did not receive any medication.

    OBJECTIVES: Nonsteroidal anti-inflammatory agents are widely used in the treatment of low back patients, but their long-term effects on the matrix molecules in the degenerate disc are unknown.

    SUMMARY OF BACKGROUND DATA: Several in vitro and in vivo studies on articular cartilage have suggested that tiaprofenic acid may not have adverse effects on matrix metabolism, whereas indomethacin probably does.

    METHODS: Uronic acid, DNA, and water contents were determined from five different locations in each injured disc. Transport and incorporation of sulfate were examined by in vivo radioactive tracer analysis, and proteoglycan structures were analyzed by gel electrophoresis.

    RESULTS: Morphologically, there were no differences between the treatments. Tiaprofenic acid maintained a higher uronic acid content in the nucleus pulposus and outer anulus compared with that of the nonmedicated animals. Tiaprofenic acid decreased the incorporation of sulfate in the injured area and the water content at most sites. Indomethacin had no adverse effects compared with the nonmedicated group, and it increased water content in the posterior anulus fibrosus.

    CONCLUSIONS: Long-term administration of tiaprofenic acid and indomethacin did not have harmful effects on matrix metabolism after disc injury. On the contrary, tiaprofenic acid may slightly protect proteoglycans in the degenerating disc.

  • 450.
    Keefover-Ring, Kenneth
    Umeå University, Faculty of Science and Technology, Department of Plant Physiology. Umeå University, Faculty of Science and Technology, Umeå Plant Science Centre (UPSC).
    Making scents of defense: Do fecal shields and herbivore-caused volatiles match host plant chemical profiles?2013In: Chemoecology, ISSN 0937-7409, E-ISSN 1423-0445, Vol. 23, no 1, p. 1-11Article in journal (Refereed)
    Abstract [en]

    Many plant families have aromatic species that produce volatile compounds which they release when damaged, particularly after suffering herbivory. Monarda fistulosa (Lamiaceae) makes and stores volatile essential oils in peltate glandular trichomes on leaf and floral surfaces. This study examined the larvae of a specialist tortoise beetle, Physonota unipunctata, which feed on two M. fistulosa chemotypes and incorporate host compounds into fecal shields, structures related to defense. Comparisons of shield and host leaf chemistry showed differences between chemotypes and structures (leaves vs. shields). Thymol chemotype leaves and shields contained more of all compounds that differed than did carvacrol chemotypes, except for carvacrol. Shields had lower levels of most of the more volatile chemicals than leaves, but more than twice the amounts of the phenolic monoterpenes thymol and carvacrol and greater totals. Additional experiments measured the volatiles emitted from M. fistulosa in the absence and presence of P. unipunctata larvae and compared the flower and foliage chemistry of plants from these experiments. Flowers contained lower or equal amounts of most compounds and half the total amount, compared to leaves. Plants subjected to herbivory emitted higher levels of most volatiles and 12 times the total amount, versus controls with no larvae, including proportionally more of the low boiling point chemicals. Thus, chemical profiles of shields and volatile emissions are influenced by the amounts and volatilities of compounds present in the host plant. The implications of these results are explored for the chemical ecology of both the plant and the insect. 

6789101112 401 - 450 of 969
CiteExportLink to result list
Permanent link
Cite
Citation style
  • apa
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Other style
More styles
Language
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Other locale
More languages
Output format
  • html
  • text
  • asciidoc
  • rtf