uu.seUppsala universitets publikasjoner
Endre søk
Begrens søket
1 - 44 of 44
RefereraExporteraLink til resultatlisten
Permanent link
Referera
Referensformat
  • apa
  • ieee
  • modern-language-association
  • vancouver
  • Annet format
Fler format
Språk
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Annet språk
Fler språk
Utmatningsformat
  • html
  • text
  • asciidoc
  • rtf
Treff pr side
  • 5
  • 10
  • 20
  • 50
  • 100
  • 250
Sortering
  • Standard (Relevans)
  • Forfatter A-Ø
  • Forfatter Ø-A
  • Tittel A-Ø
  • Tittel Ø-A
  • Type publikasjon A-Ø
  • Type publikasjon Ø-A
  • Eldste først
  • Nyeste først
  • Skapad (Eldste først)
  • Skapad (Nyeste først)
  • Senast uppdaterad (Eldste først)
  • Senast uppdaterad (Nyeste først)
  • Disputationsdatum (tidligste først)
  • Disputationsdatum (siste først)
  • Standard (Relevans)
  • Forfatter A-Ø
  • Forfatter Ø-A
  • Tittel A-Ø
  • Tittel Ø-A
  • Type publikasjon A-Ø
  • Type publikasjon Ø-A
  • Eldste først
  • Nyeste først
  • Skapad (Eldste først)
  • Skapad (Nyeste først)
  • Senast uppdaterad (Eldste først)
  • Senast uppdaterad (Nyeste først)
  • Disputationsdatum (tidligste først)
  • Disputationsdatum (siste først)
Merk
Maxantalet träffar du kan exportera från sökgränssnittet är 250. Vid större uttag använd dig av utsökningar.
  • 1.
    Amselem, Elias
    et al.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Marklund, Emil
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Kipper, Kalle
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi.
    Johansson, Magnus
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Deindl, Sebastian
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Elf, Johan
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Real- Time Single Protein Tracking with Polarization Readout using a Confocal Microscope2017Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 112, nr 3, s. 295A-295AArtikkel i tidsskrift (Annet vitenskapelig)
  • 2.
    Ballet, Caroline
    et al.
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Correia, Mario S. P.
    Uppsala universitet, Science for Life Laboratory, SciLifeLab. Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi.
    Conway, Louis P.
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Locher, Theresa L.
    Uppsala universitet, Science for Life Laboratory, SciLifeLab. Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi.
    Lehmann, Laura C.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Garg, Neeraj
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Kemisk biologi för biomarkörer. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Vujasinovic, Miroslav
    Karolinska Univ Hosp, Dept Digest Dis, Stockholm, Sweden.
    Deindl, Sebastian
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Lohr, J. -Matthias
    Karolinska Inst, Dept Clin Sci Intervent & Technol CLINTEC, Stockholm, Sweden;Karolinska Univ Hosp, Dept Digest Dis, Stockholm, Sweden.
    Globisch, Daniel
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Kemisk biologi för biomarkörer. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    New enzymatic and mass spectrometric methodology for the selective investigation of gut microbiota-derived metabolites2018Inngår i: Chemical Science, ISSN 2041-6520, E-ISSN 2041-6539, Vol. 9, nr 29, s. 6233-6239Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Gut microbiota significantly impact human physiology through metabolic interaction. Selective investigation of the co-metabolism of bacteria and their human host is a challenging task and methods for their analysis are limited. One class of metabolites associated with this co-metabolism are O-sulfated compounds. Herein, we describe the development of a new enzymatic assay for the selective mass spectrometric investigation of this phase II modification class. Analysis of human urine and fecal samples resulted in the detection of 206 sulfated metabolites, which is three times more than reported in the Human Metabolome Database. We confirmed the chemical structure of 36 sulfated metabolites including unknown and commonly reported microbiota-derived sulfated metabolites using synthesized internal standards and mass spectrometric fragmentation experiments. Our findings demonstrate that enzymatic sample pre-treatment combined with state-of-the-art metabolomics analysis represents a new and efficient strategy for the discovery of unknown microbiota-derived metabolites in human samples. Our described approach can be adapted for the targeted investigation of other metabolite classes as well as the discovery of biomarkers for diseases affected by microbiota.

  • 3.
    Baltekin, Özden
    Uppsala universitet, Science for Life Laboratory, SciLifeLab. Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Astrego Diagnost AB, Uppsala, Sweden.
    KN Direct phenotypic antimicrobial susceptibility testing under 30 minutes: Dynamics of single cell response revealed in automated microscopy with microfluidics2018Inngår i: Journal of Veterinary Pharmacology and Therapeutics, ISSN 0140-7783, E-ISSN 1365-2885, Vol. 41, nr S1, s. 17-17Artikkel i tidsskrift (Annet vitenskapelig)
  • 4.
    Baltekin, Özden
    et al.
    Uppsala universitet, Science for Life Laboratory, SciLifeLab. Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Boucharin, Alexis
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Tano, Eva
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinska vetenskaper, Infektionsmedicin.
    Andersson, Dan I
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk biokemi och mikrobiologi.
    Elf, Johan
    Uppsala universitet, Science for Life Laboratory, SciLifeLab. Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Antibiotic susceptibility testing in less than 30 min using direct single-cell imaging2017Inngår i: Proceedings of the National Academy of Sciences of the United States of America, ISSN 0027-8424, E-ISSN 1091-6490, Vol. 114, nr 34, s. 9170-9175Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The emergence and spread of antibiotic-resistant bacteria are aggravated by incorrect prescription and use of antibiotics. A core problem is that there is no sufficiently fast diagnostic test to guide correct antibiotic prescription at the point of care. Here, we investigate if it is possible to develop a point-of-care susceptibility test for urinary tract infection, a disease that 100 million women suffer from annually and that exhibits widespread antibiotic resistance. We capture bacterial cells directly from samples with low bacterial counts (10(4) cfu/mL) using a custom-designed microfluidic chip and monitor their individual growth rates using microscopy. By averaging the growth rate response to an antibiotic over many individual cells, we can push the detection time to the biological response time of the bacteria. We find that it is possible to detect changes in growth rate in response to each of nine antibiotics that are used to treat urinary tract infections in minutes. In a test of 49 clinical uropathogenic Escherichia coli (UPEC) isolates, all were correctly classified as susceptible or resistant to ciprofloxacin in less than 10 min. The total time for antibiotic susceptibility testing, from loading of sample to diagnostic readout, is less than 30 min, which allows the development of a point-of-care test that can guide correct treatment of urinary tract infection.

  • 5.
    Balzarotti, Francisco
    et al.
    Max Planck Inst Biophys Chem, Dept NanoBiophoton, Gottingen, Germany..
    Eilers, Yvan
    Max Planck Inst Biophys Chem, Dept NanoBiophoton, Gottingen, Germany..
    Gwosch, Klaus C.
    Max Planck Inst Biophys Chem, Dept NanoBiophoton, Gottingen, Germany..
    Gynnå, Arvid H.
    Uppsala universitet, Science for Life Laboratory, SciLifeLab. Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Westphal, Volker
    Max Planck Inst Biophys Chem, Dept NanoBiophoton, Gottingen, Germany..
    Stefani, Fernando D.
    Consejo Nacl Invest Cient & Tecn, Ctr Invest Bionanociencias CIBION, Buenos Aires, DF, Argentina.;Univ Buenos Aires, Fac Ciencias Exactas & Nat, Dept Fis, Buenos Aires, DF, Argentina..
    Elf, Johan
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Hell, Stefan W.
    Max Planck Inst Biophys Chem, Dept NanoBiophoton, Gottingen, Germany.;Max Planck Inst Med Res, Dept Opt Nanoscopy, Heidelberg, Germany.;German Canc Res Ctr, Opt Nanoscopy Div, Heidelberg, Germany..
    Nanometer resolution imaging and tracking of fluorescent molecules with minimal photon fluxes2017Inngår i: Science, ISSN 0036-8075, E-ISSN 1095-9203, Vol. 355, nr 6325, s. 606-612Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    We introduce MINFLUX, a concept for localizing photon emitters in space. By probing the emitter with a local intensity minimum of excitation light, MINFLUX minimizes the fluorescence photons needed for high localization precision. In our experiments, 22 times fewer fluorescence photons are required as compared to popular centroid localization. In superresolutionmicroscopy, MINFLUXattained similar to 1-nanometer precision, resolving molecules only 6 nanometers apart. MINFLUX tracking of single fluorescent proteins increased the temporal resolution and the number of localizations per trace by a factor of 100, as demonstrated with diffusing 30S ribosomal subunits in living Escherichia coli. As conceptual limits have not been reached, we expect this localization modality to break new ground for observing the dynamics, distribution, and structure of macromolecules in living cells and beyond.

  • 6.
    Belikov, Sergey
    et al.
    Karolinska Inst, Dept Cell & Mol Biol, SE-17177 Stockholm, Sweden.;Stockholm Univ, Wenner Gren Inst, Dept Mol Biosci, SE-10691 Stockholm, Sweden..
    Berg, Otto G.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Wrange, Orjan
    Karolinska Inst, Dept Cell & Mol Biol, SE-17177 Stockholm, Sweden..
    Quantification of transcription factor-DNA binding affinity in a living cell2016Inngår i: Nucleic Acids Research, ISSN 0305-1048, E-ISSN 1362-4962, Vol. 44, nr 7, s. 3045-3058Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The apparent dissociation constant (K-d) for specific binding of glucocorticoid receptor (GR) and androgen receptor (AR) to DNA was determined in vivo in Xenopus oocytes. The total nuclear receptor concentration was quantified as specifically retained [H-3]-hormone in manually isolated oocyte nuclei. DNA was introduced by nuclear microinjection of single stranded phagemid DNA, chromatin is then formed during second strand synthesis. The fraction of DNA sites occupied by the expressed receptor was determined by dimethylsulphate in vivo footprinting and used for calculation of the receptor-DNA binding affinity. The forkhead transcription factor FoxA1 enhanced the DNA binding by GR with an apparent K-d of similar to 1 mu M and dramatically stimulated DNA binding by AR with an apparent K-d of similar to 0.13 mu M at a composite androgen responsive DNA element containing one FoxA1 binding site and one palindromic hormone receptor binding site known to bind one receptor homodimer. FoxA1 exerted a weak constitutive- and strongly cooperative DNA binding together with AR but had a less prominent effect with GR, the difference reflecting the licensing function of FoxA1 at this androgen responsive DNA element.

  • 7.
    Belliveau, Nathan M.
    et al.
    CALTECH, Div Biol & Biol Engn, Pasadena, USA.
    Barnes, Stephanie L.
    CALTECH, Div Biol & Biol Engn, Pasadena, USA.
    Ireland, William T.
    CALTECH, Dept Phys, Pasadena, USA.
    Jones, Daniel L.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Sweredoski, Michael J.
    CALTECH, Beckman Inst, Proteome Explorat Lab, Pasadena, USA.
    Moradian, Annie
    CALTECH, Beckman Inst, Proteome Explorat Lab, Pasadena, USA.
    Hess, Sonja
    CALTECH, Beckman Inst, Proteome Explorat Lab, Pasadena, CA 91125 USA;MedImmune, Antibody Discovery & Prot Engn, Gaithersburg, USA.
    Kinney, Justin B.
    Cold Spring Harbor Lab, Simons Ctr Quantitat Biol, POB 100, Cold Spring Harbor, USA.
    Phillips, Rob
    CALTECH, Div Biol & Biol Engn, Pasadena, USA;CALTECH, Dept Phys, Pasadena, CA 91125 USA;CALTECH, Dept Appl Phys, Pasadena, CA 91125 USA.
    Systematic approach for dissecting the molecular mechanisms of transcriptional regulation in bacteria2018Inngår i: Proceedings of the National Academy of Sciences of the United States of America, ISSN 0027-8424, E-ISSN 1091-6490, Vol. 115, nr 21, s. E4796-E4805Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Gene regulation is one of the most ubiquitous processes in biology. However, while the catalog of bacterial genomes continues to expand rapidly, we remain ignorant about how almost all of the genes in these genomes are regulated. At present, characterizing the molecular mechanisms by which individual regulatory sequences operate requires focused efforts using low-throughput methods. Here, we take a first step toward multipromoter dissection and show how a combination of massively parallel reporter assays, mass spectrometry, and information-theoretic modeling can be used to dissect multiple bacterial promoters in a systematic way. We show this approach on both well-studied and previously uncharacterized promoters in the enteric bacterium Escherichia coli. In all cases, we recover nucleotide-resolution models of promoter mechanism. For some promoters, including previously unannotated ones, the approach allowed us to further extract quantitative biophysical models describing input-output relationships. Given the generality of the approach presented here, it opens up the possibility of quantitatively dissecting the mechanisms of promoter function in E. coli and a wide range of other bacteria.

  • 8.
    Berg, Otto G.
    et al.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Mahmutovic, Anel
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Marklund, Emil
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Elf, Johan
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    The helical structure of DNA facilitates binding2016Inngår i: Journal of Physics A: Mathematical and Theoretical, ISSN 1751-8113, E-ISSN 1751-8121, Vol. 9, nr 36, artikkel-id 364002Artikkel i tidsskrift (Annet vitenskapelig)
    Abstract [en]

    The helical structure of DNA imposes constraints on the rate of diffusion-limited protein binding. Here we solve the reaction-diffusion equations for DNA-like geometries and extend with simulations when necessary. We find that the helical structure can make binding to the DNA more than twice as fast compared to a case where DNA would be reactive only along one side. We also find that this rate advantage remains when the contributions from steric constraints and rotational diffusion of the DNA-binding protein are included. Furthermore, we find that the association rate is insensitive to changes in the steric constraints on the DNA in the helix geometry, while it is much more dependent on the steric constraints on the DNA-binding protein. We conclude that the helical structure of DNA facilitates the nonspecific binding of transcription factors and structural DNA-binding proteins in general.

  • 9.
    Bowman, Gregory D.
    et al.
    Johns Hopkins Univ, Thomas C Jenkins Dept Biophys, Baltimore, MD 21218 USA.
    Deindl, Sebastian
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Remodeling the genome with DNA twists2019Inngår i: Science, ISSN 0036-8075, E-ISSN 1095-9203, Vol. 366, nr 6461, s. 35-36Artikkel i tidsskrift (Annet vitenskapelig)
  • 10.
    Doudna, Jennifer
    et al.
    Univ Calif Berkeley, Berkeley, CA 94720 USA..
    Bar-Ziv, Roy
    Weizmann Inst Sci, IL-76100 Rehovot, Israel..
    Elf, Johan
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Noireaux, Vincent
    Univ Minnesota, Minneapolis, MN 55455 USA..
    Berro, Julien
    Yale Univ, New Haven, CT 06520 USA..
    Saiz, Leonor
    Univ Calif Davis, Davis, CA 95616 USA..
    Vavylonis, Dimitrios
    Lehigh Univ, Bethlehem, PA 18015 USA..
    Faulon, Jean-Loup
    INRA, Jouy En Josas, France..
    Fordyce, Polly
    Stanford Univ, Stanford, CA 94305 USA..
    How Will Kinetics and Thermodynamics Inform Our Future Efforts to Understand and Build Biological Systems?2017Inngår i: CELL SYSTEMS, ISSN 2405-4712, Vol. 4, nr 2, s. 144-146Artikkel i tidsskrift (Fagfellevurdert)
  • 11.
    Elf, Johan
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Hypothesis: Homologous Recombination Depends on Parallel Search2016Inngår i: CELL SYSTEMS, ISSN 2405-4712, Vol. 3, nr 4, s. 325-327Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    It is not known how a cell manages to find a specific DNA sequence sufficiently fast to repair a broken chromosome through homologous recombination. I propose a solution based on freely diffusing molecules that are programmed with sequences corresponding to those flanking the break site. In such a process, the target search would be parallelized.

  • 12.
    Elf, Johan
    et al.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Barkefors, Irmeli
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk biokemi och mikrobiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Single-Molecule Kinetics in Living Cells2019Inngår i: Annual Review of Biochemistry, ISSN 0066-4154, E-ISSN 1545-4509, Vol. 88, s. 635-659Artikkel, forskningsoversikt (Fagfellevurdert)
    Abstract [en]

    In the past decades, advances in microscopy have made it possible to study the dynamics of individual biomolecules in vitro and resolve intramolecular kinetics that would otherwise be hidden in ensemble averages. More recently, single-molecule methods have been used to image, localize, and track individually labeled macromolecules in the cytoplasm of living cells, allowing investigations of intermolecular kinetics under physiologically relevant conditions. In this review, we illuminate the particular advantages of single-molecule techniques when studying kinetics in living cells and discuss solutions to specific challenges associated with these methods.

  • 13.
    Elias-Wolff, Federico
    et al.
    Stockholm Univ, Dept Biochem & Biophys, Stockholm, Sweden; Stockholm Univ, Dept Mat & Environm Chem, Stockholm, Sweden.
    Lindén, Martin
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Lyubartsev, Alexander P.
    Stockholm Univ, Dept Mat & Environm Chem, Stockholm, Sweden.
    Brandt, Erik G.
    Stockholm Univ, Dept Mat & Environm Chem, Stockholm, Sweden.
    Computing Curvature Sensitivity of Biomolecules in Membranes by Simulated Buckling2018Inngår i: Journal of Chemical Theory and Computation, ISSN 1549-9618, E-ISSN 1549-9626, Vol. 14, nr 3, s. 1643-1655Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Membrane curvature sensing, where the binding free energies of membrane-associated molecules depend on the local membrane curvature, is a key factor to modulate and maintain the shape and organization of cell membranes. However, the microscopic mechanisms are not well understood, partly due to absence of efficient simulation methods. Here, we describe a method to compute the curvature dependence of the binding free energy of a membrane associated probe molecule that interacts with a buckled membrane, which has been created by lateral compression of a flat bilayer patch. This buckling approach samples a wide range of curvatures in a single simulation, and anisotropic effects can be extracted from the orientation statistics. We develop an efficient and robust algorithm to extract the motion of the probe along the buckled membrane surface, and evaluate its numerical properties by extensive sampling of three coarse-grained model systems: local lipid density in a curved environment for single-component bilayers, curvature preferences of individual lipids in two-component membranes, and curvature sensing by a homotrimeric transmembrane protein. The method can be used to complement experimental data from curvature partition assays and provides additional insight into mesoscopic theories and molecular mechanisms for curvature sensing.

  • 14.
    Elias-Wolff, Federico
    et al.
    Stockholm Univ, Dept Biochem & Biophys, Stockholm, Sweden;Stockholm Univ, Dept Mat & Environm Chem, Stockholm, Sweden.
    Lindén, Martin
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Scania CV AB, Sodertalje, Sweden.
    Lyubartsev, Alexander P.
    Stockholm Univ, Dept Mat & Environm Chem, Stockholm, Sweden.
    Brandt, Erik G.
    Stockholm Univ, Dept Mat & Environm Chem, Stockholm, Sweden.
    Curvature sensing by cardiolipin in simulated buckled membranes2019Inngår i: Soft Matter, ISSN 1744-683X, E-ISSN 1744-6848, Vol. 15, nr 4, s. 792-802Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Cardiolipin is a non-bilayer phospholipid with a unique dimeric structure. It localizes to negative curvature regions in bacteria and is believed to stabilize respiratory chain complexes in the highly curved mitochondrial membrane. Cardiolipin's localization mechanism remains unresolved, because important aspects such as the structural basis and strength for lipid curvature preferences are difficult to determine, partly due to the lack of efficient simulation methods. Here, we report a computational approach to study curvature preferences of cardiolipin by simulated membrane buckling and quantitative modeling. We combine coarse-grained molecular dynamics with simulated buckling to determine the curvature preferences in three-component bilayer membranes with varying concentrations of cardiolipin, and extract curvature-dependent concentrations and lipid acyl chain order parameter profiles. Cardiolipin shows a strong preference for negative curvatures, with a highly asymmetric chain order parameter profile. The concentration profiles are consistent with an elastic model for lipid curvature sensing that relates lipid segregation to local curvature via the material constants of the bilayers. These computations constitute new steps to unravel the molecular mechanism by which cardiolipin senses curvature in lipid membranes, and the method can be generalized to other lipids and membrane components as well.

  • 15.
    Forcier, Talitha L.
    et al.
    Cold Spring Harbor Lab, Simons Ctr Quantitat Biol, POB 100, Cold Spring Harbor, NY 11724 USA.
    Ayaz, Andalus
    Cold Spring Harbor Lab, Simons Ctr Quantitat Biol, POB 100, Cold Spring Harbor, NY 11724 USA.
    Gill, Manraj S.
    Cold Spring Harbor Lab, Simons Ctr Quantitat Biol, POB 100, Cold Spring Harbor, NY 11724 USA;MIT, Dept Biol, Cambridge, MA 02139 USA.
    Jones, Daniel
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Cold Spring Harbor Lab, Simons Ctr Quantitat Biol, POB 100, Cold Spring Harbor, NY 11724 USA;CALTECH, Dept Appl Phys, Pasadena, CA 91125 USA.
    Phillips, Rob
    CALTECH, Dept Appl Phys, Pasadena, CA 91125 USA.
    Kinney, Justin B.
    Cold Spring Harbor Lab, Simons Ctr Quantitat Biol, POB 100, Cold Spring Harbor, NY 11724 USA.
    Measuring cis-regulatory energetics in living cells using allelic manifolds2018Inngår i: eLIFE, E-ISSN 2050-084X, Vol. 7, artikkel-id e40618Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Gene expression in all organisms is controlled by cooperative interactions between DNA-bound transcription factors (TFs), but quantitatively measuring TF-DNA and TF-TF interactions remains difficult. Here we introduce a strategy for precisely measuring the Gibbs free energy of such interactions in living cells. This strategy centers on the measurement and modeling of 'allelic manifolds', a multidimensional generalization of the classical genetics concept of allelic series. Allelic manifolds are measured using reporter assays performed on strategically designed cis-regulatory sequences. Quantitative biophysical models are then fit to the resulting data. We used this strategy to study regulation by two Escherichia coli TFs, CRP and sigma(70) RNA polymerase. Doing so, we consistently obtained energetic measurements precise to similar to 0.1 kcal/mol. We also obtained multiple results that deviate from the prior literature. Our strategy is compatible with massively parallel reporter assays in both prokaryotes and eukaryotes, and should therefore be highly scalable and broadly applicable.

    Editorial note: This article has been through an editorial process in which the authors decide how to respond to the issues raised during peer review. The Reviewing Editor's assessment is that minor issues remain unresolved (see decision letter).

  • 16.
    Ghosh, Anirban
    et al.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Mikrobiologi.
    Baltekin, Özden
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Wäneskog, Marcus
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Mikrobiologi.
    Elkhalifa, Dina
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi.
    Larsson, Disa
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Mikrobiologi.
    Elf, Johan
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Koskiniemi, Sanna
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Mikrobiologi.
    Contact-dependent growth inhibition induces high levels of antibiotic-tolerant persister cells in clonal bacterial populations2018Inngår i: EMBO Journal, ISSN 0261-4189, E-ISSN 1460-2075, Vol. 37, nr 9, artikkel-id UNSP e98026Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Bacterial populations can use bet-hedging strategies to cope with rapidly changing environments. One example is non-growing cells in clonal bacterial populations that are able to persist antibiotic treatment. Previous studies suggest that persisters arise in bacterial populations either stochastically through variation in levels of global signalling molecules between individual cells, or in response to various stresses. Here, we show that toxins used in contact-dependent growth inhibition (CDI) create persisters upon direct contact with cells lacking sufficient levels of CdiI immunity protein, which would otherwise bind to and neutralize toxin activity. CDI-mediated persisters form through a feedforward cycle where the toxic activity of the CdiA toxin increases cellular (p)ppGpp levels, which results in Lon-mediated degradation of the immunity protein and more free toxin. Thus, CDI systems mediate a population density-dependent bet-hedging strategy, where the fraction of non-growing cells is increased only when there are many cells of the same genotype. This may be one of the mechanisms of how CDI systems increase the fitness of their hosts.

  • 17.
    Jones, Daniel
    et al.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Elf, Johan
    Uppsala universitet, Science for Life Laboratory, SciLifeLab. Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Bursting onto the scene?: Exploring stochastic mRNA production in bacteria2018Inngår i: Current Opinion in Microbiology, ISSN 1369-5274, E-ISSN 1879-0364, Vol. 45, s. 124-130Artikkel, forskningsoversikt (Fagfellevurdert)
    Abstract [en]

    Recent large-scale measurements of gene expression variability (or noise) in E. coli have led to the unexpected conclusion that the variability is in large part dictated by and increasing with the mean level of expression. Here we review the evidence for this apparent universal trend in variability, as well as for the related idea that transcription is fundamentally bursty. We examine recently proposed mechanisms for burstiness and universality and argue that they do not explain important features of observed data. Finally, we discuss potential limitations and pitfalls in the interpretation of experimental measurements of cell-to-cell variability.

  • 18.
    Jones, Daniel
    et al.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Leroy, Prune
    Uppsala universitet, Science for Life Laboratory, SciLifeLab. Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Unoson, Cecilia
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Mikrobiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Fange, David
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Curic, Vladimir
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Lawson, Michael J.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Elf, Johan
    Uppsala universitet, Science for Life Laboratory, SciLifeLab. Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Uppsala Univ, Dept Cell & Mol Biol, Sci Life Lab, Uppsala, Sweden..
    Kinetics of dCas9 target search in Escherichia coli2017Inngår i: Science, ISSN 0036-8075, E-ISSN 1095-9203, Vol. 357, nr 6358, s. 1420-1423Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    How fast can a cell locate a specific chromosomal DNA sequence specified by a single-stranded oligonucleotide? To address this question, we investigate the intracellular search processes of the Cas9 protein, which can be programmed by a guide RNA to bind essentially any DNA sequence. This targeting flexibility requires Cas9 to unwind the DNA double helix to test for correct base pairing to the guide RNA. Here we study the search mechanisms of the catalytically inactive Cas9 (dCas9) in living Escherichia coli by combining single-molecule fluorescence microscopy and bulk restriction-protection assays. We find that it takes a single fluorescently labeled dCas9 6 hours to find the correct target sequence, which implies that each potential target is bound for less than 30 milliseconds. Once bound, dCas9 remains associated until replication. To achieve fast targeting, both Cas9 and its guide RNA have to be present at high concentrations.

  • 19.
    Jones, Daniel
    et al.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Unoson, Cecilia
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi.
    Leroy, Prune
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi.
    Curic, Vladimir
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi.
    Elf, Johan
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Kinetics of dCas9 Target Search in Escherichia Coli2017Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 112, nr 3, s. 314A-314AArtikkel i tidsskrift (Annet vitenskapelig)
  • 20.
    Kipper, Kalle
    et al.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Eremina, Nadja
    Uppsala universitet, Science for Life Laboratory, SciLifeLab. Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi.
    Marklund, Emil
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Tubasum, Sumera
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Mao, Guanzhong
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylärbiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Lehmann, Laura Christina
    Uppsala universitet, Science for Life Laboratory, SciLifeLab. Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Elf, Johan
    Uppsala universitet, Science for Life Laboratory, SciLifeLab. Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Deindl, Sebastian
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Structure-guided approach to site-specific fluorophore labeling of the lac repressor LacI2018Inngår i: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 13, nr 6, artikkel-id e0198416Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The lactose operon repressor protein LacI has long served as a paradigm of the bacterial transcription factors. However, the mechanisms whereby LacI rapidly locates its cognate binding site on the bacterial chromosome are still elusive. Single-molecule fluorescence imaging approaches are well suited for the study of these mechanisms but rely on a functionally compatible fluorescence labeling of LacI. Particularly attractive for protein fluorescence labeling are synthetic fluorophores due to their small size and favorable photophysical characteristics. Synthetic fluorophores are often conjugated to natively occurring cysteine residues using maleimide chemistry. For a site-specific and functionally compatible labeling with maleimide fluorophores, the target protein often needs to be redesigned to remove unwanted native cysteines and to introduce cysteines at locations better suited for fluorophore attachment. Biochemical screens can then be employed to probe for the functional activity of the redesigned protein both before and after dye labeling. Here, we report a mutagenesis- based redesign of LacI to enable a functionally compatible labeling with maleimide fluorophores. To provide an easily accessible labeling site in LacI, we introduced a single cysteine residue at position 28 in the DNA-binding headpiece of LacI and replaced two native cysteines with alanines where derivatization with bulky substituents is known to compromise the protein's activity. We find that the redesigned LacI retains a robust activity in vitro and in vivo, provided that the third native cysteine at position 281 is retained in LacI. In a total internal reflection microscopy assay, we observed individual Cy3-labeled LacI molecules bound to immobilized DNA harboring the cognate O-1 operator sequence, indicating that the dye-labeled LacI is functionally active. We have thus been able to generate a functional fluorescently labeled LacI that can be used to unravel mechanistic details of LacI target search at the single molecule level.

  • 21.
    Kipper, Kalle
    et al.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Lundius, Ebba G.
    Uppsala universitet, Science for Life Laboratory, SciLifeLab. Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi.
    Ćurić, Vladimir
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Nikic, Ivana
    European Mol Biol Lab, Cell Biol & Biophys Unit, Struct & Computat Biol Unit, D-69117 Heidelberg, Germany..
    Wiessler, Manfred
    Deutsch Krebsforschungszentrum, Biol Chem, D-69120 Heidelberg, Germany..
    Lemke, Edward A.
    European Mol Biol Lab, Cell Biol & Biophys Unit, Struct & Computat Biol Unit, D-69117 Heidelberg, Germany..
    Elf, Johan
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Application of Noncanonical Amino Acids for Protein Labeling in a Genomically Recoded Escherichia coli2017Inngår i: ACS Photonics, E-ISSN 2330-4022, Vol. 6, nr 2, s. 233-255Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Small synthetic fluorophores are in many ways superior to fluorescent proteins as labels for imaging. A major challenge is to use them for a protein-specific labeling in living cells. Here, we report on our use of noncanonical amino acids that are genetically encoded via the pyrrolysyl-tRNA/pyrrolysyl-RNA synthetase pair at artificially introduced TAG codons in a recoded E. coli strain. The strain is lacking endogenous TAG codons and the TAG-specific release factor RF1. The amino acids contain bioorthogonal groups that can be clicked to externally supplied dyes, thus enabling protein-specific labeling in live cells. We find that the noncanonical amino acid incorporation into the target protein is robust for diverse amino acids and that the usefulness of the recoded E. coli strain mainly derives from the absence of release factor RF1. However, the membrane permeable dyes display high nonspecific binding in intracellular environment and the electroporation of hydrophilic nonmembrane permeable dyes severely impairs growth of the recoded strain. In contrast, proteins exposed on the outer membrane of E. coli can be labeled with hydrophilic dyes with a high specificity as demonstrated by labeling of the osmoporin OmpC. Here, labeling can be made sufficiently specific to enable single molecule studies as exemplified by OmpC single particle tracking.

  • 22.
    Lawson, Michael J.
    et al.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Camsund, Daniel
    Uppsala universitet, Science for Life Laboratory, SciLifeLab. Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Larsson, Jimmy
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Baltekin, Özden
    Uppsala universitet, Science for Life Laboratory, SciLifeLab. Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Fange, David
    Uppsala universitet, Science for Life Laboratory, SciLifeLab. Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Elf, Johan
    Uppsala universitet, Science for Life Laboratory, SciLifeLab. Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    In situ genotyping of a pooled strain library after characterizing complex phenotypes2017Inngår i: Molecular Systems Biology, ISSN 1744-4292, E-ISSN 1744-4292, Vol. 13, nr 10, artikkel-id 947Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    In this work, we present a proof-of-principle experiment that extends advanced live cell microscopy to the scale of pool-generated strain libraries. We achieve this by identifying the genotypes for individual cells in situ after a detailed characterization of the phenotype. The principle is demonstrated by single-molecule fluorescence time-lapse imaging of Escherichia coli strains harboring barcoded plasmids that express a sgRNA which suppresses different genes in the E.coli genome through dCas9 interference. In general, the method solves the problem of characterizing complex dynamic phenotypes for diverse genetic libraries of cell strains. For example, it allows screens of how changes in regulatory or coding sequences impact the temporal expression, location, or function of a gene product, or how the altered expression of a set of genes impacts the intracellular dynamics of a labeled reporter.

  • 23.
    Lehmann, Laura C.
    et al.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Hewitt, Graeme
    Francis Crick Inst, 1 Midland Rd, London NW1 1AT, England..
    Aibara, Shintaro
    Stockholm Univ, Dept Biochem & Biophys, Sci Life Lab, S-17165 Solna, Sweden..
    Leitner, Alexander
    Swiss Fed Inst Technol, Inst Mol Syst Biol, Dept Biol, CH-8093 Zurich, Switzerland..
    Marklund, Emil
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Maslen, Sarah L.
    MRC Lab Mol Biol, Francis Crick Ave,Cambridge Biomed Campus, Cambridge CB2 0QH, England..
    Maturi, Varun
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwiginstitutet för cancerforskning. Uppsala universitet, Science for Life Laboratory, SciLifeLab. Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk biokemi och mikrobiologi.
    Chen, Yang
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    van der Spoel, David
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Beräkningsbiologi och bioinformatik.
    Skehel, J. Mark
    MRC Lab Mol Biol, Francis Crick Ave,Cambridge Biomed Campus, Cambridge CB2 0QH, England..
    Moustakas, Aristidis
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwiginstitutet för cancerforskning. Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk biokemi och mikrobiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Boulton, Simon J.
    Francis Crick Inst, 1 Midland Rd, London NW1 1AT, England..
    Deindl, Sebastian
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Mechanistic Insights into Autoinhibition of the Oncogenic Chromatin Remodeler ALC12017Inngår i: Molecular Cell, ISSN 1097-2765, E-ISSN 1097-4164, Vol. 68, nr 5, s. 847-859.e7Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Human ALC1 is an oncogene-encoded chromatin-remodeling enzyme required for DNA repair that possesses a poly(ADP-ribose) (PAR)-binding macro domain. Its engagement with PARylated PARP1 activates ALC1 at sites of DNA damage, but the underlying-mechanism remains unclear. Here, we establish a dual role for the macro domain in autoinhibition of ALC1 ATPase activity and coupling to nucleosome mobilization. In the absence of DNA damage, an inactive conformation of the ATPase is maintained by juxtaposition of the macro domain against predominantly the C-terminal ATPase lobe through conserved electrostatic interactions. Mutations within this interface displace the macro domain, constitutively activate the ALC1 ATPase independent of PARylated PARP1, and alter the dynamics of ALC1 recruitment at DNA damage sites. Upon DNA damage, binding of PARylated PARP1 by the macro domain induces a conformational change that relieves autoinhibitory interactions with the ATPase motor, which selectively activates ALC1 remodeling upon recruitment to sites of DNA damage.

  • 24.
    Levendosky, Robert F.
    et al.
    Johns Hopkins Univ, TC Jenkins Dept Biophys, Baltimore, MD 21218 USA..
    Sabantsev, Anton
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Deindl, Sebastian
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Bowman, Gregory D.
    Johns Hopkins Univ, TC Jenkins Dept Biophys, Baltimore, MD 21218 USA..
    The Chd1 chromatin remodeler shifts hexasomes unidirectionally2016Inngår i: eLIFE, E-ISSN 2050-084X, Vol. 5, artikkel-id e21356Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Despite their canonical two-fold symmetry, nucleosomes in biological contexts are often asymmetric: functionalized with post-translational modifications (PTMs), substituted with histone variants, and even lacking H2A/H2B dimers. Here we show that the Widom 601 nucleosome positioning sequence can produce hexasomes in a specific orientation on DNA, providing a useful tool for interrogating chromatin enzymes and allowing for the generation of nucleosomes with precisely defined asymmetry. Using this methodology, we demonstrate that the Chd1 chromatin remodeler from Saccharomyces cerevisiae requires H2A/H2B on the entry side for sliding, and thus, unlike the back-and-forth sliding observed for nucleosomes, Chd1 shifts hexasomes unidirectionally. Chd1 takes part in chromatin reorganization surrounding transcribing RNA polymerase II (Pol II), and using asymmetric nucleosomes we show that ubiquitin-conjugated H2B on the entry side stimulates nucleosome sliding by Chd1. We speculate that biased nucleosome and hexasome sliding due to asymmetry contributes to the packing of arrays observed in vivo.

  • 25.
    Liljeruhm, Josefine
    et al.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylärbiologi.
    Funk, Saskia K.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi.
    Tietscher, Sandra
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi.
    Edlund, Anders D.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi. Uppsala Univ, iGEM Uppsala, Uppsala, Sweden.
    Jamal, Sabri
    Uppsala Univ, iGEM Uppsala, Uppsala, Sweden.
    Yuen, Pikkei
    Uppsala Univ, iGEM Uppsala, Uppsala, Sweden.
    Dyrhage, Karl
    Uppsala Univ, iGEM Uppsala, Uppsala, Sweden.
    Gynnå, Arvid H.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Uppsala Univ, iGEM Uppsala, Uppsala, Sweden.
    Ivermark, Katarina
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för biologisk grundutbildning.
    Lövgren, Jessica
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för biologisk grundutbildning.
    Törnblom, Viktor
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för biologisk grundutbildning.
    Virtanen, Anders
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Mikrobiologi.
    Lundin, Erik
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk biokemi och mikrobiologi. Uppsala Univ, iGEM Uppsala, Uppsala, Sweden.
    Wistand-Yuen, Erik
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk biokemi och mikrobiologi.
    Forster, Anthony C.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylärbiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Engineering a palette of eukaryotic chromoproteins for bacterial synthetic biology2018Inngår i: Journal of Biological Engineering, ISSN 1754-1611, E-ISSN 1754-1611, Vol. 12, artikkel-id 8Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Background: Coral reefs are colored by eukaryotic chromoproteins (CPs) that are homologous to green fluorescent protein. CPs differ from fluorescent proteins (FPs) by intensely absorbing visible light to give strong colors in ambient light. This endows CPs with certain advantages over FPs, such as instrument-free detection uncomplicated by ultra-violet light damage or background fluorescence, efficient Forster resonance energy transfer (FRET) quenching, and photoacoustic imaging. Thus, CPs have found utility as genetic markers and in teaching, and are attractive for potential cell biosensor applications in the field. Most near-term applications of CPs require expression in a different domain of life: bacteria. However, it is unclear which of the eukaryotic CP genes might be suitable and how best to assay them.

    Results: Here, taking advantage of codon optimization programs in 12 cases, we engineered 14 CP sequences (meffRed, eforRed, asPink, spisPink, scOrange, fwYellow, amilGFP, amajLime, cjBlue, mefiBlue, aeBlue, amilCP, tsPurple and gfasPurple) into a palette of Escherichia coil BioBrick plasmids. BioBricks comply with synthetic biology's most widely used, simplified, cloning standard. Differences in color intensities, maturation times and fitness costs of expression were compared under the same conditions, and visible readout of gene expression was quantitated. A surprisingly large variation in cellular fitness costs was found, resulting in loss of color in some overnight liquid cultures of certain high-copy-plasmid-borne CPs, and cautioning the use of multiple CPs as markers in competition assays. We solved these two problems by integrating pairs of these genes into the chromosome and by engineering versions of the same CP with very different colors.

    Conclusion: Availability of 14 engineered CP genes compared in E coil, together with chromosomal mutants suitable for competition assays, should simplify and expand CP study and applications. There was no single plasmid-borne CP that combined all of the most desirable features of intense color, fast maturation and low fitness cost, so this study should help direct future engineering efforts.

  • 26.
    Lind, Peter A.
    et al.
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk biokemi och mikrobiologi. Umea Univ, Dept Mol Biol, S-90187 Umea, Sweden..
    Arvidsson, Lars
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk biokemi och mikrobiologi.
    Berg, Otto
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Andersson, Dan I.
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk biokemi och mikrobiologi.
    Variation in Mutational Robustness between Different Proteins and the Predictability of Fitness Effects2017Inngår i: Molecular biology and evolution, ISSN 0737-4038, E-ISSN 1537-1719, Vol. 34, nr 2, s. 408-418Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Random mutations in genes from disparate protein classes may have different distributions of fitness effects (DFEs) depending on different structural, functional, and evolutionary constraints. We measured the fitness effects of 156 single mutations in the genes encoding AraC (transcription factor), AraD (enzyme), and AraE (transporter) used for bacterial growth on L-arabinose. Despite their different molecular functions these genes all had bimodal DFEs with most mutations either being neutral or strongly deleterious, providing a general expectation for the DFE. This contrasts with the unimodal DFEs previously obtained for ribosomal protein genes where most mutations were slightly deleterious. Based on theoretical considerations, we suggest that the 33-fold higher average mutational robustness of ribosomal proteins is due to stronger selection for reduced costs of translational and transcriptional errors. Whereas the large majority of synonymous mutations were deleterious for ribosomal proteins genes, no fitness effects could be detected for the AraCDE genes. Four mutations in AraC and AraE increased fitness, suggesting that slightly advantageous mutations make up a significant fraction of the DFE, but that they often escape detection due to the limited sensitivity of commonly used fitness assays. We show that the fitness effects of amino acid substitutions can be predicted based on evolutionary conservation, but those weakly deleterious mutations are less reliably detected. This suggests that large-effect mutations and the fraction of highly deleterious mutations can be computationally predicted, but that experiments are required to characterize the DFE close to neutrality, where many mutations ultimately fixed in a population will occur.

  • 27.
    Lindén, Martin
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    An early peak in ion channel research2018Inngår i: Nature Physics, ISSN 1745-2473, E-ISSN 1745-2481, Vol. 14, nr 2, s. 105-105Artikkel i tidsskrift (Annet vitenskapelig)
  • 28.
    Lindén, Martin
    et al.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Curic, Vladimir
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Amselem, Elias
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Elf, Johan
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Pointwise error estimates in localization microscopy2017Inngår i: Nature Communications, ISSN 2041-1723, E-ISSN 2041-1723, Vol. 8, artikkel-id 15115Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Pointwise localization of individual fluorophores is a critical step in super-resolution localization microscopy and single particle tracking. Although the methods are limited by the localization errors of individual fluorophores, the pointwise localization precision has so far been estimated using theoretical best case approximations that disregard, for example, motion blur, defocus effects and variations in fluorescence intensity. Here, we show that pointwise localization precision can be accurately estimated directly from imaging data using the Bayesian posterior density constrained by simple microscope properties. We further demonstrate that the estimated localization precision can be used to improve downstream quantitative analysis, such as estimation of diffusion constants and detection of changes in molecular motion patterns. Finally, the quality of actual point localizations in live cell super-resolution microscopy can be improved beyond the information theoretic lower bound for localization errors in individual images, by modelling the movement of fluorophores and accounting for their pointwise localization uncertainty.

  • 29.
    Lindén, Martin
    et al.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Elf, Johan
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Variational Algorithms for Analyzing Noisy Multistate Diffusion Trajectories2018Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 115, nr 2, s. 276-282Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Single-particle tracking offers a noninvasive high-resolution probe of biomolecular reactions inside living cells. However, efficient data analysis methods that correctly account for various noise sources are needed to realize the full quantitative potential of the method. We report algorithms for hidden Markov-based analysis of single-particle tracking data, which incorporate most sources of experimental noise, including heterogeneous localization errors and missing positions. Compared to previous implementations, the algorithms offer significant speedups, support for a wider range of inference methods, and a simple user interface. This will enable more advanced and exploratory quantitative analysis of single-particle tracking data.

  • 30.
    Lindén, Martin
    et al.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Ćurić, Vladimir
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Boucharin, Alexis
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Fange, David
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Elf, Johan
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Simulated single molecule microscopy with SMeagol2016Inngår i: Bioinformatics, ISSN 1367-4803, E-ISSN 1367-4811, Vol. 32, nr 15, s. 2394-2395Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Summary: SMeagol is a software tool to simulate highly realistic microscopy data based on spatial systems biology models, in order to facilitate development, validation and optimization of advanced analysis methods for live cell single molecule microscopy data.

    Availability and implementation: SMeagol runs on Matlab R2014 and later, and uses compiled binaries in C for reaction–diffusion simulations. Documentation, source code and binaries for Mac OS, Windows and Ubuntu Linux can be downloaded from http://smeagol.sourceforge.net.

  • 31.
    Marklund, Emil
    et al.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Amselem, Elias
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Kipper, Kalle
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi.
    Johansson, Magnus
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Deindl, Sebastian
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Elf, Johan
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Measuring the Orientation of Single Proteins Interacting with DNA using Fluorescence Polarization Microscopy2017Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 112, nr 3, s. 169A-169AArtikkel i tidsskrift (Annet vitenskapelig)
  • 32.
    Martyna, Agnieszka
    et al.
    Univ Kent, Sch Biosci, Canterbury CT2 7NJ, Kent, England..
    Gomez-Llobregat, Jordi
    Stockholm Univ, Dept Biochem & Biophys, Ctr Biomembrane Res, SE-10691 Stockholm, Sweden..
    Lindén, Martin
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Stockholm Univ, Dept Biochem & Biophys, Ctr Biomembrane Res, SE-10691 Stockholm, Sweden..
    Rossman, Jeremy S.
    Univ Kent, Sch Biosci, Canterbury CT2 7NJ, Kent, England..
    Curvature Sensing by a Viral Scission Protein2016Inngår i: Biochemistry, ISSN 0006-2960, E-ISSN 1520-4995, Vol. 55, nr 25, s. 3493-3496Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Membrane scission is the final step in all budding processes wherein a membrane neck is sufficiently constricted so as to allow for fission and the release of the budded particle. For influenza viruses, membrane scission is mediated by an amphipathic helix (AH) domain in the viral M2 protein. While it is known that the M2AH alters membrane curvature, it is not known how the protein is localized to the center neck of budding virions where it would be able to cause membrane scission. Here, we use molecular dynamics simulations on buckled lipid bilayers to show that the M2AH senses membrane curvature and preferentially localizes to regions of high membrane curvature, comparable to that seen at the center neck of budding influenza viruses. These results were then validated using in vitro binding assays to show that the M2AH senses membrane curvature by detecting lipid packing defects in the membrane. Our results show that the M2AH senses membrane curvature and suggest that the AH domain may localize the protein at the viral neck where it can then mediate membrane scission and the release of budding viruses.

  • 33.
    Pandzic, Tatjana
    et al.
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för immunologi, genetik och patologi, Experimentell och klinisk onkologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Rendo, Verónica
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för immunologi, genetik och patologi, Experimentell och klinisk onkologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Lim, Jinyeong
    Department of Cancer Biomedical Science, National Cancer Center Graduate School of Cancer Science and Policy, Goyangsi, Republic of Korea.
    Larsson, Chatarina
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för immunologi, genetik och patologi, Experimentell och klinisk onkologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Larsson, Jimmy
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Stoimenov, Ivaylo
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för immunologi, genetik och patologi, Experimentell och klinisk onkologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Kundu, Snehangshu
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för immunologi, genetik och patologi, Experimentell och klinisk onkologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Ali, Muhammad Akhtar
    Uppsala universitet, Science for Life Laboratory, SciLifeLab. Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för immunologi, genetik och patologi, Experimentell och klinisk onkologi.
    Hellström, Mats
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för immunologi, genetik och patologi, Experimentell och klinisk onkologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    He, Liqun
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för immunologi, genetik och patologi, Vaskulärbiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Lindroth, Anders M.
    Department of Cancer Biomedical Science, National Cancer Center Graduate School of Cancer Science and Policy, Goyangsi, Republic of Korea.
    Sjöblom, Tobias
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för immunologi, genetik och patologi, Experimentell och klinisk onkologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Somatic PRDM2 c.4467delA mutations in colorectal cancers control histone methylation and tumor growth2017Inngår i: OncoTarget, ISSN 1949-2553, E-ISSN 1949-2553, Vol. 8, nr 58, s. 98646-98659Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The chromatin modifier PRDM2/RIZ1 is inactivated by mutation in several forms of cancer and is a putative tumor suppressor gene. Frameshift mutations in the C-terminal region of PRDM2, affecting (A)8 or (A)9 repeats within exon 8, are found in one third of colorectal cancers with microsatellite instability, but the contribution of these mutations to colorectal tumorigenesis is unknown. To model somatic mutations in microsatellite unstable tumors, we devised a general approach to perform genome editing while stabilizing the mutated nucleotide repeat. We then engineered isogenic cell systems where the PRDM2 c.4467delA mutation in human HCT116 colorectal cancer cells was corrected to wild-type by genome editing. Restored PRDM2 increased global histone 3 lysine 9 dimethylation and reduced migration, anchorage-independent growth and tumor growth in vivo. Gene set enrichment analysis revealed regulation of several hallmark cancer pathways, particularly of epithelial-to-mesenchymal transition (EMT), with VIM being the most significantly regulated gene. These observations provide direct evidence that PRDM2 c.4467delA is a driver mutation in colorectal cancer and confirms PRDM2 as a cancer gene, pointing to regulation of EMT as a central aspect of its tumor suppressive action.

  • 34.
    Romilly, Cedric
    et al.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Mikrobiologi.
    Deindl, Sebastian
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Wagner, Gerhart E. H.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Mikrobiologi.
    The ribosomal protein S1-dependent standby site in tisB mRNA consists of a single-stranded region and a 5 ' structure element2019Inngår i: Proceedings of the National Academy of Sciences of the United States of America, ISSN 0027-8424, E-ISSN 1091-6490, Vol. 116, nr 32, s. 15901-15906Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    In bacteria, stable RNA structures that sequester ribosome-binding sites (RBS) impair translation initiation, and thus protein output. In some cases, ribosome standby can overcome inhibition by structure: 30S subunits bind sequence-nonspecifically to a single-stranded region and, on breathing of the inhibitory structure, relocate to the RBS for initiation. Standby can occur over long distances, as in the active, +42 tisB mRNA, encoding a toxin. This mRNA is translationally silenced by an antitoxin sRNA, IstR-1, that base pairs to the standby site. In tisB and other cases, a direct interaction between 30S subunits and a standby site has remained elusive. Based on fluorescence anisotropy experiments, ribosome toeprinting results, in vitro translation assays, and cross-linking-immunoprecipitation (CLIP) in vitro, carried out on standby-proficient and standby-deficient tisB mRNAs, we provide a thorough characterization of the tisB standby site. 30S subunits and ribosomal protein S1 alone display high-affinity binding to standby-competent fluorescein-labeled +42 mRNA, but not to mRNAs that lack functional standby sites. Ribosomal protein S1 is essential for standby, as 30 Delta S1 subunits do not support standby-dependent toeprints and TisB translation in vitro. S1 alone- and 30S-CLIP followed by RNA-seq mapping shows that the functional tisB standby site consists of the expected single-stranded region, but surprisingly, also a 5'-end stem-loop structure. Removal of the latter by 5'-truncations, or disruption of the stem, abolishes 30S binding and standby activity. Based on the CLIP-read mapping, the long-distance standby effect in +42 tisB mRNA (similar to 100 nt) is tentatively explained by S1-dependent directional unfolding toward the downstream RBS.

  • 35.
    Sabantsev, Anton
    et al.
    Uppsala universitet, Science for Life Laboratory, SciLifeLab. Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Levendosky, Robert F.
    Johns Hopkins Univ, TC Jenkins Dept Biophys, Baltimore, MD 21218 USA.
    Zhuang, Xiaowei
    Harvard Univ, Howard Hughes Med Inst, Cambridge, MA 02138 USA;Harvard Univ, Dept Chem & Chem Biol, Cambridge, MA 02138 USA;Harvard Univ, Dept Phys, Cambridge, MA 02138 USA.
    Bowman, Gregory D.
    Johns Hopkins Univ, TC Jenkins Dept Biophys, Baltimore, MD 21218 USA.
    Deindl, Sebastian
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Direct observation of coordinated DNA movements on the nucleosome during chromatin remodelling2019Inngår i: Nature Communications, ISSN 2041-1723, E-ISSN 2041-1723, Vol. 10, artikkel-id 1720Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    ATP-dependent chromatin remodelling enzymes (remodellers) regulate DNA accessibility in eukaryotic genomes. Many remodellers reposition (slide) nucleosomes, however, how DNA is propagated around the histone octamer during this process is unclear. Here we examine the real-time coordination of remodeller-induced DNA movements on both sides of the nucleosome using three-colour single-molecule FRET. During sliding by Chd1 and SNF2h remodellers, DNA is shifted discontinuously, with movement of entry-side DNA preceding that of exit-side DNA. The temporal delay between these movements implies a single ratelimiting step dependent on ATP binding and transient absorption or buffering of at least one base pair. High-resolution cross-linking experiments show that sliding can be achieved by buffering as few as 3 bp between entry and exit sides of the nucleosome. We propose that DNA buffering ensures nucleosome stability during ATP-dependent remodelling, and provides a means for communication between remodellers acting on opposite sides of the nucleosome.

  • 36.
    Saupe, Falk
    et al.
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk biokemi och mikrobiologi.
    Reichel, Matthias
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk biokemi och mikrobiologi.
    Huijbers, Elisabeth J. M.
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk biokemi och mikrobiologi.
    Femel, Julia
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk biokemi och mikrobiologi.
    Markgren, Per-Olof
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Kemiska sektionen, Institutionen för kemi - BMC.
    Andersson, C. Evalena
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Deindl, Sebastian
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Danielson, U. Helena
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Kemiska sektionen, Institutionen för kemi - BMC, Biokemi.
    Hellman, Lars T.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Olsson, Anna-Karin
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk biokemi och mikrobiologi. Uppsala Univ, Dept Med Biochem & Microbiol, Husargatan 3,Box 582, SE-75123 Uppsala, Sweden..
    Development of a novel therapeutic vaccine carrier that sustains high antibody titers against several targets simultaneously2017Inngår i: The FASEB Journal, ISSN 0892-6638, E-ISSN 1530-6860, Vol. 31, nr 3, s. 1204-1214Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    With the aim to improve the efficacy of therapeutic vaccines that target self-antigens, we have developed a novel fusion protein vaccine on the basis of the C-terminal multimerizing end of the variable lymphocyte receptor B (VLRB), the Ig equivalent in jawless fishes. Recombinant vaccines were produced in Escherichia coli by fusing the VLRB sequence to 4 different cancer-associated target molecules. The anti-self-immune response generated in mice that were vaccinated with VLRB vaccines was compared with the response in mice that received vaccines that contained bacterial thioredoxin (TRX), previously identified as an efficient carrier. The anti-self-Abswere analyzed with respect to titers, binding properties, and duration of response. VLRB-vaccinatedmice displayed a 2-to 10-fold increase in anti-self-Ab titers and a substantial decrease in Abs against the foreign part of the fusion protein compared with the response in TRX-vaccinated mice (P < 0.01). VLRB-generated Ab response had duration similar to the corresponding TRX-generatedAbs, but displayed a higher diversity in binding characteristics. Of importance, VLRB vaccines could sustain an immune response against several targets simultaneously. VLRB vaccines fulfill several key criteria for an efficient therapeutic vaccine that targets self-antigens as a result of its small size, its multimerizing capacity, and nonexposed foreign sequences in the fusion protein.- Saupe, F., Reichel, M., Huijbers, E. J. M., Femel, J., Markgren, P.- O., Andersson, C. E., Deindl, S., Danielson, U. H., Hellman, L. T., Olsson, A.- K. Development of a novel therapeutic vaccine carrier that sustains high antibody titers against several targets simultaneously.

  • 37.
    Travin, Dmitrii Y.
    et al.
    Lomonosov Moscow State Univ, Dept Bioengn & Bioinformat, Moscow, Russia; Skolkovo Inst Sci & Technol, Ctr Data Intens Biomed & Biotechnol, Skolkovo, Russia.
    Metelev, Mikhail
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Skolkovo Inst Sci & Technol, Ctr Data Intens Biomed & Biotechnol, Skolkovo, Russia; Russian Acad Sci, Inst Gene Biol, Moscow, Russia.
    Serebryakova, Marina
    Skolkovo Inst Sci & Technol, Ctr Data Intens Biomed & Biotechnol, Skolkovo, Russia; Lomonosov Moscow State Univ, Dept Chem, Moscow, Russia; Lomonosov Moscow State Univ, AN Belozersky Inst Physicochem Biol, Moscow, Russia.
    Komarova, Ekaterina S.
    Lomonosov Moscow State Univ, Dept Bioengn & Bioinformat, Moscow, Russia; Skolkovo Inst Sci & Technol, Ctr Data Intens Biomed & Biotechnol, Skolkovo, Russia.
    Osterman, Ilya A.
    Lomonosov Moscow State Univ, Dept Chem, Moscow, Russia; Lomonosov Moscow State Univ, AN Belozersky Inst Physicochem Biol, Moscow, Russia; Skolkovo Inst Sci & Technol, Ctr Translat Biomed, Skolkovo, Russia.
    Ghilarov, Dmitry
    Skolkovo Inst Sci & Technol, Ctr Data Intens Biomed & Biotechnol, Skolkovo, Russia; Russian Acad Sci, Inst Gene Biol, Moscow, Russia; Jagiellonian Univ, Malopolska Ctr Biotechnol, Krakow, Poland.
    Severinov, Konstantin
    Skolkovo Inst Sci & Technol, Ctr Data Intens Biomed & Biotechnol, Skolkovo, Russia; Russian Acad Sci, Inst Gene Biol, Moscow, Russia; Rutgers State Univ, Waksman Inst Microbiol, Piscataway, NJ USA.
    Biosynthesis of Translation Inhibitor Klebsazolicin Proceeds through Heterocyclization and N-Terminal Amidine Formation Catalyzed by a Single YcaO Enzyme2018Inngår i: Journal of the American Chemical Society, ISSN 0002-7863, E-ISSN 1520-5126, Vol. 140, nr 16, s. 5625-5633Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Klebsazolicin (KLB) is a recently discovered Klebsiella pneumonia peptide antibiotic targeting the exit tunnel of bacterial ribosome. KLB contains an N-terminal amidine ring and four azole heterocycles installed into a ribosomally synthesized precursor by dedicated maturation machinery. Using an in vitro system for KLB production, we show that the YcaO-domain KlpD maturation enzyme is a bifunctional cyclodehydratase required for the formation of both the core heterocycles and the N-terminal amidine ring. We further demonstrate that the amidine ring is formed concomitantly with proteolytic cleavage of azole-containing pro-KLB by a cellular protease TldD/E. Members of the YcaO family are diverse enzymes known to activate peptide carbonyls during natural product biosynthesis leading to the formation of azoline, macroamidine, and thioamide moieties. The ability of KlpD to simultaneously perform two distinct types of modifications is unprecedented for known YcaO proteins. The versatility of KlpD opens up possibilities for rational introduction of modifications into various peptide backbones.

  • 38.
    Volkov, Ivan
    et al.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Aguirre, Javier
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Lindén, Martin
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Elf, Johan
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Johansson, Magnus
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    In Vivo Measurements of Protein Synthesis Kinetics using Single-Molecule Tracking of E.Coli tRNAS2016Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 110, nr 3, s. 351A-351AArtikkel i tidsskrift (Annet vitenskapelig)
  • 39.
    Volkov, Ivan
    et al.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Johansson, Magnus
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Single-Molecule Tracking Approaches to Protein Synthesis Kinetics in Living Cells2019Inngår i: Biochemistry, ISSN 0006-2960, E-ISSN 1520-4995, Vol. 58, nr 1, s. 7-14Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Decades of traditional biochemistry, structural approaches, and, more recently, single-molecule-based in vitro techniques have provided us with an astonishingly detailed understanding of the molecular mechanism of ribosome-catalyzed protein synthesis. However, in order to understand these details in the context of cell physiology and population biology, new techniques to probe the dynamics of molecular processes inside the cell are needed. Recent years' development in super-resolved fluorescence microscopy has revolutionized imaging of intracellular processes, and we now have the possibility to directly peek into the microcosm of biomolecules in their native environment. In this Perspective, we discuss how these methods are currently being applied and further developed to study the kinetics of protein synthesis directly inside living cells.

  • 40.
    Volkov, Ivan
    et al.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Lindén, Martin
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Aguirre, Javier
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Ieong, Ka-Weng
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylärbiologi.
    Metelev, Mikhail
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Elf, Johan
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Johansson, Magnus
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    tRNA tracking for direct measurements of protein synthesis kinetics in live cells2018Inngår i: Nature Chemical Biology, ISSN 1552-4450, E-ISSN 1552-4469, Vol. 14, nr 6, s. 618-626Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Our ability to directly relate results from test-tube biochemical experiments to the kinetics in living cells is very limited. Here we present experimental and analytical tools to directly study the kinetics of fast biochemical reactions in live cells. Dye-labeled molecules are electroporated into bacterial cells and tracked using super-resolved single-molecule microscopy.Trajectories are analyzed by machine-learning algorithms to directly monitor transitions between bound and free states. In particular, we measure the dwell time of tRNAs on ribosomes, and hence achieve direct measurements of translation rates inside living cells at codon resolution. We find elongation rates with tRNA(Phe) that are in perfect agreement with previous indirect estimates, and once fMet-tRNA(fMet) has bound to the 30S ribosomal subunit, initiation of translation is surprisingly fast and does not limit the overall rate of protein synthesis. The experimental and analytical tools for direct kinetics measurements in live cells have applications far beyond bacterial protein synthesis.

  • 41.
    Volkov, Ivan
    et al.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Seefeldt, A. Carolin
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Johansson, Magnus
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Tracking of single tRNAs for translation kinetics measurements in chloramphenicol treated bacteria2019Inngår i: Methods, ISSN 1046-2023, E-ISSN 1095-9130, Vol. 162-163, s. 23-30Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Chloramphenicol is a broad-spectrum antibiotic targeting the protein synthesis machinery by binding to the bacterial ribosome. Chloramphenicol has been considered a classic general inhibitor of translation, blocking the accommodation of aa-tRNA into the A site of the large ribosomal subunit. However, recent studies suggest that this proposed mechanism is a simplification and that the effect of chloramphenicol on mRNA translation is much more dynamic. By tracking single dye-labelled elongator and initiator tRNAs in Escherichia coli cells treated with chloramphenicol, we observe the direct effect of chloramphenicol on translation kinetics. We find clear indications of slow but significant mRNA translation on drug bound ribosomes.

  • 42.
    Walldén, Mats
    et al.
    Uppsala universitet, Science for Life Laboratory, SciLifeLab. Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk biokemi och mikrobiologi.
    Fange, David
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Lundius, Ebba Gregorsson
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Beräknings- och systembiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Baltekin, Özden
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    Elf, Johan
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
    The Synchronization of Replication and Division Cycles in Individual E. coli Cells2016Inngår i: Cell, ISSN 0092-8674, E-ISSN 1097-4172, Vol. 166, nr 3, s. 729-739Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Isogenic E. coli cells growing in a constant environment display significant variability in growth rates, division sizes, and generation times. The guiding principle appears to be that each cell, during one generation, adds a size increment that is uncorrelated to its birth size. Here, we investigate the mechanisms underlying this "adder'' behavior by mapping the chromosome replication cycle to the division cycle of individual cells using fluorescence microscopy. We have found that initiation of chromosome replication is triggered at a fixed volume per chromosome independent of a cell's birth volume and growth rate. Each initiation event is coupled to a division event after a growth-rate-dependent time. We formalize our findings in a model showing that cell-to-cell variation in division timing and cell size is mainly driven by variations in growth rate. The model also explains why fast-growing cells display adder behavior and correctly predict deviations from the adder behavior at slow growth.

  • 43.
    Wistrand-Yuen, Erik
    et al.
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk biokemi och mikrobiologi.
    Knopp, Michael
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk biokemi och mikrobiologi.
    Hjort, Karin
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk biokemi och mikrobiologi.
    Koskiniemi, Sanna
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Mikrobiologi.
    Berg, Otto G.
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Molekylär systembiologi.
    Andersson, Dan I.
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk biokemi och mikrobiologi.
    Evolution of high-level resistance during low-level antibiotic exposure2018Inngår i: Nature Communications, ISSN 2041-1723, E-ISSN 2041-1723, Vol. 9, nr 1, artikkel-id 1599Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    It has become increasingly clear that low levels of antibiotics present in many environments can select for resistant bacteria, yet the evolutionary pathways for resistance development during exposure to low amounts of antibiotics remain poorly defined. Here we show that Salmonella enterica exposed to sub-MIC levels of streptomycin evolved high-level resistance via novel mechanisms that are different from those observed during lethal selections. During lethal selection only rpsL mutations are found, whereas at sub-MIC selection resistance is generated by several small-effect resistance mutations that combined confer high-level resistance via three different mechanisms: (i) alteration of the ribosomal RNA target (gidB mutations), (ii) reduction in aminoglycoside uptake (cyoB, nuoG, and trkH mutations), and (iii) induction of the aminoglycoside-modifying enzyme AadA (znuA mutations). These results demonstrate how the strength of the selective pressure influences evolutionary trajectories and that even weak selective pressures can cause evolution of high-level resistance.