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  • 101.
    Björkholm, Patrik
    et al.
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Ernst, Andreas M.
    Hacke, Moritz
    Wieland, Felix
    Bruegger, Britta
    von Heijne, Gunnar
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Identification of novel sphingolipid-binding motifs in mammalian membrane proteins2014In: Biochimica et Biophysica Acta - Biomembranes, ISSN 0005-2736, E-ISSN 1879-2642, Vol. 1838, no 8, p. 2066-2070Article in journal (Refereed)
    Abstract [en]

    Specific interactions between transmembrane proteins and sphingolipids is a poorly understood phenomenon, and only a couple of instances have been identified. The best characterized example is the sphingolipid-binding motif VXXTLXXIY found in the transmembrane helix of the vesicular transport protein p24. Here, we have used a simple motif-probability algorithm (MOPRO) to identify proteins that contain putative sphingolipid-binding motifs in a dataset comprising proteomes from mammalian organisms. From these motif-containing candidate proteins, four with different numbers of transmembrane helices were selected for experimental study: i) major histocompatibility complex II Q alpha chain subtype (DQA1), ii) GPI-attachment protein 1 (GAA1), iii) tetraspanin-7 TSN7, and iv), metabotropic glutamate receptor 2 (GRM2). These candidates were subjected to photo-affinity labeling using radiolabeled sphingolipids, confirming all four candidate proteins as sphingolipid-binding proteins. The sphingolipid-binding motifs are enriched in the 7TM family of G-protein coupled receptors, predominantly in transmembrane helix 6. The ability of the motif-containing candidate proteins to bind sphingolipids with high specificity opens new perspectives on their respective regulation and function.

  • 102. Blau, Christian
    et al.
    Yvonnesdotter, Linnea
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Lindahl, Erik
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Gentle and fast all-atom model refinement to cryo-EM densities via a maximum likelihood approach2023In: PloS Computational Biology, ISSN 1553-734X, E-ISSN 1553-7358, Vol. 19, no 7, article id e1011255Article in journal (Refereed)
    Abstract [en]

    Better detectors and automated data collection have generated a flood of high-resolution cryo-EM maps, which in turn has renewed interest in improving methods for determining structure models corresponding to these maps. However, automatically fitting atoms to densities becomes difficult as their resolution increases and the refinement potential has a vast number of local minima. In practice, the problem becomes even more complex when one also wants to achieve a balance between a good fit of atom positions to the map, while also establishing good stereochemistry or allowing protein secondary structure to change during fitting. Here, we present a solution to this challenge using a maximum likelihood approach by formulating the problem as identifying the structure most likely to have produced the observed density map. This allows us to derive new types of smooth refinement potential-based on relative entropy-in combination with a novel adaptive force scaling algorithm to allow balancing of force-field and density-based potentials. In a low-noise scenario, as expected from modern cryo-EM data, the relative-entropy based refinement potential outperforms alternatives, and the adaptive force scaling appears to aid all existing refinement potentials. The method is available as a component in the GROMACS molecular simulation toolkit. Author summaryCryo-electron microscopy has gone through a revolution and now regularly produces data with 2 & ANGS; resolution. However, this data comes in the shape of density maps, and fitting atomic coordinates into these maps can be a labor-intensive and challenging problem. This is particularly valid when there are multiple conformations, flexible regions, or parts of the structure with lower resolution. In many cases it is also desirable to to understand how a molecule moves between such conformations. This can be addressed with molecular dynamics simulations using densities as target restraints, but the refinement potentials commonly used can distort protein structure or get stuck in local minima when the cryo-EM map has high resolution. This work derives new refinement potentials based on models of the cryo-EM scattering process that provide a gentle way to fit protein structures to densities in simulations, and we also suggest an automated heuristic way to balance the influence of the map and simulation force field.

  • 103. Bohutínská, Magdalena
    et al.
    Vlček, Jakub
    Yair, Sivan
    Laenen, Benjamin
    Stockholm University, Faculty of Science, Department of Ecology, Environment and Plant Sciences. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Konečná, Veronika
    Fracassetti, Marco
    Stockholm University, Science for Life Laboratory (SciLifeLab). Stockholm University, Faculty of Science, Department of Ecology, Environment and Plant Sciences.
    Slotte, Tanja
    Stockholm University, Faculty of Science, Department of Ecology, Environment and Plant Sciences. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Kolář, Filip
    Genomic basis of parallel adaptation varies with divergence in Arabidopsis and its relatives2021In: Proceedings of the National Academy of Sciences of the United States of America, ISSN 0027-8424, E-ISSN 1091-6490, Vol. 118, no 21, article id e2022713118Article in journal (Refereed)
    Abstract [en]

    Parallel adaptation provides valuable insight into the predictability of evolutionary change through replicated natural experiments. A steadily increasing number of studies have demonstrated genomic parallelism, yet the magnitude of this parallelism varies depending on whether populations, species, or genera are compared. This led us to hypothesize that the magnitude of genomic parallelism scales with genetic divergence between lineages, but whether this is the case and the underlying evolutionary processes remain unknown. Here, we resequenced seven parallel lineages of two Arabidopsis species, which repeatedly adapted to challenging alpine environments. By combining genome-wide divergence scans with model-based approaches, we detected a suite of 151 genes that show parallel signatures of positive selection associated with alpine colonization, involved in response to cold, high radiation, short season, herbivores, and pathogens. We complemented these parallel candidates with published gene lists from five additional alpine Brassicaceae and tested our hypothesis on a broad scale spanning ∼0.02 to 18 My of divergence. Indeed, we found quantitatively variable genomic parallelism whose extent significantly decreased with increasing divergence between the compared lineages. We further modeled parallel evolution over the Arabidopsis candidate genes and showed that a decreasing probability of repeated selection on the same standing or introgressed alleles drives the observed pattern of divergence-dependent parallelism. We therefore conclude that genetic divergence between populations, species, and genera, affecting the pool of shared variants, is an important factor in the predictability of genome evolution.

  • 104. Boije, Henrik
    et al.
    Ring, Henrik
    Fard, Shahrzad Shirazi
    Grundberg, Ida
    Nilsson, Mats
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab). Uppsala University .
    Hallbook, Finn
    Alternative Splicing of the Chromodomain Protein Morf4l1 Pre-mRNA Has Implications on Cell Differentiation in the Developing Chicken Retina2013In: Journal of Molecular Neuroscience, ISSN 0895-8696, E-ISSN 1559-1166, Vol. 51, no 2, p. 615-628Article in journal (Refereed)
    Abstract [en]

    The proliferation, cell cycle exit and differentiation of progenitor cells are controlled by several different factors. The chromodomain protein mortality factor 4-like 1 (Morf4l1) has been ascribed a role in both proliferation and differentiation. Little attention has been given to the existence of alternative splice variants of the Morf4l1 mRNA, which encode two Morf41l isoforms: a short isoform (S-Morf4l1) with an intact chromodomain and a long isoform (L-Morf4l1) with an insertion in or in the vicinity of the chromodomain. The aim of this study was to investigate if this alternative splicing has a function during development. We analysed the temporal and spatial distribution of the two mRNAs and over-expressed both isoforms in the developing retina. The results showed that the S-Morf4l1 mRNA is developmentally regulated. Over-expression of S-Morf4l1 using a retrovirus vector produced a clear phenotype with an increase of early-born neurons: retinal ganglion cells, horizontal cells and cone photoreceptor cells. Over-expression of L-Morf4l1 did not produce any distinguishable phenotype. The over-expression of S-Morf4l1 but not L-Morf4l1 also increased apoptosis in the infected regions. Our results suggest that the two Morf4l1 isoforms have different functions during retinogenesis and that Morf4l1 functions are fine-tuned by developmentally regulated alternative splicing. The data also suggest that Morf4l1 contributes to the regulation of cell genesis in the retina.

  • 105. Bonagas, Nadilly
    et al.
    Gustafsson, Nina M. S.
    Henriksson, Martin
    Marttila, Petra
    Gustafsson, Robert
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
    Wiita, Elisée
    Borhade, Sanjay
    Green, Alanna C.
    Vallin, Karl S. A.
    Sarno, Antonio
    Svensson, Richard
    Göktürk, Camilla
    Pham, Therese
    Jemth, Ann-Sofie
    Loseva, Olga
    Cookson, Victoria
    Kiweler, Nicole
    Sandberg, Lars
    Stockholm University, Science for Life Laboratory (SciLifeLab). Stockholm University, Faculty of Science, Department of Organic Chemistry.
    Rasti, Azita
    Unterlass, Judith E.
    Haraldsson, Martin
    Andersson, Yasmin
    Scaletti, Emma R.
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Lund University, Sweden.
    Bengtsson, Christoffer
    Stockholm University, Science for Life Laboratory (SciLifeLab). Stockholm University, Faculty of Science, Department of Organic Chemistry.
    Paulin, Cynthia B. J.
    Sanjiv, Kumar
    Abdurakhmanov, Eldar
    Pudelko, Linda
    Kunz, Ben
    Desroses, Matthieu
    Iliev, Petar
    Färnegårdh, Katarina
    Stockholm University, Science for Life Laboratory (SciLifeLab). Stockholm University, Faculty of Science, Department of Organic Chemistry.
    Krämer, Andreas
    Garg, Neeraj
    Michel, Maurice
    Häggblad, Sara
    Stockholm University, Science for Life Laboratory (SciLifeLab). Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
    Jarvius, Malin
    Kalderén, Christina
    Bögedahl Jensen, Amanda
    Almlöf, Ingrid
    Karsten, Stella
    Zhang, Si Min
    Häggblad, Maria
    Stockholm University, Science for Life Laboratory (SciLifeLab). Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
    Eriksson, Anders
    Liu, Jianping
    Glinghammar, Björn
    Nekhotiaeva, Natalia
    Klingegård, Fredrik
    Stockholm University, Science for Life Laboratory (SciLifeLab). Stockholm University, Faculty of Science, Department of Organic Chemistry.
    Koolmeister, Tobias
    Martens, Ulf
    Stockholm University, Science for Life Laboratory (SciLifeLab). Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
    Llona-Minguez, Sabin
    Moulson, Ruth
    Nordström, Helena
    Parrow, Vendela
    Dahllund, Leif
    Sjöberg, Birger
    Vargas, Irene L.
    Vo, Duy Duc
    Wannberg, Johan
    Knapp, Stefan
    Krokan, Hans E.
    Arvidsson, Per
    Scobie, Martin
    Meiser, Johannes
    Stenmark, Pål
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Lund University, Sweden.
    Warpman Berglund, Ulrika
    Homan, Evert J.
    Helleday, Thomas
    Pharmacological targeting of MTHFD2 suppresses acute myeloid leukemia by inducing thymidine depletion and replication stress2022In: Nature cancer, ISSN 2662-1347, Vol. 3, no 2, p. 156-172Article in journal (Refereed)
    Abstract [en]

    The folate metabolism enzyme MTHFD2 (methylenetetrahydrofolate dehydrogenase/cyclohydrolase) is consistently overexpressed in cancer but its roles are not fully characterized, and current candidate inhibitors have limited potency for clinical development. In the present study, we demonstrate a role for MTHFD2 in DNA replication and genomic stability in cancer cells, and perform a drug screen to identify potent and selective nanomolar MTHFD2 inhibitors; protein cocrystal structures demonstrated binding to the active site of MTHFD2 and target engagement. MTHFD2 inhibitors reduced replication fork speed and induced replication stress followed by S-phase arrest and apoptosis of acute myeloid leukemia cells in vitro and in vivo, with a therapeutic window spanning four orders of magnitude compared with nontumorigenic cells. Mechanistically, MTHFD2 inhibitors prevented thymidine production leading to misincorporation of uracil into DNA and replication stress. Overall, these results demonstrate a functional link between MTHFD2-dependent cancer metabolism and replication stress that can be exploited therapeutically with this new class of inhibitors. Helleday and colleagues describe a nanomolar MTHFD2 inhibitor that causes replication stress and DNA damage accumulation in cancer cells via thymidine depletion, demonstrating a potential therapeutic strategy in AML tumors in vivo.

  • 106.
    Bonath, Franziska
    et al.
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Domingo-Prim, Judit
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    Tarbier, Marcel
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Friedländer, Marc R.
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Visa, Neus
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    Next-generation sequencing reveals two populations of damage-induced small RNAs at endogenous DNA double-strand breaks2018In: Nucleic Acids Research, ISSN 0305-1048, E-ISSN 1362-4962, Vol. 46, no 22, p. 11869-11882Article in journal (Refereed)
    Abstract [en]

    Recent studies suggest that transcription takes place at DNA double-strand breaks (DSBs), that transcripts at DSBs are processed by Drosha and Dicer into damage-induced small RNAs (diRNAs), and that diRNAs are required for DNA repair. However, diRNAs have been mostly detected in reporter constructs or repetitive sequences, and their existence at endogenous loci has been questioned by recent reports. Using the homing endonuclease I-PpoI, we have investigated diRNA production in genetically unperturbed human and mouse cells. I-PpoI is an ideal tool to clarify the requirements for diRNA production because it induces DSBs in different types of loci: the repetitive 28S locus, unique genes and intergenic loci. We show by extensive sequencing that the rDNA locus produces substantial levels of diRNAs, whereas unique genic and intergenic loci do not. Further characterization of diRNAs emerging from the 28S locus reveals the existence of two diRNA subtypes. Surprisingly, Drosha and its partner DGCR8 are dispensable for diRNA production and only one diRNAs subtype depends on Dicer processing. Furthermore, we provide evidence that diRNAs are incorporated into Argonaute. Our findings provide direct evidence for diRNA production at endogenous loci in mammalian cells and give insights into RNA processing at DSBs.

  • 107.
    Bonath, Franziska
    et al.
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Domingo-Prim, Judit
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    Tarbier, Marcel
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Friedländer, Marc
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Visa, Neus
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    Next-generation sequencing reveals two populations of damage- induced small RNAs at endogenous DNA double-strand breaksIn: Nucleic Acids Research, ISSN 0305-1048, E-ISSN 1362-4962Article in journal (Refereed)
  • 108. Bondarenko, Vasyl
    et al.
    Chen, Qiang
    Singewald, Kevin
    Haloi, Nandan
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab). KTH Royal Institute of Technology, Sweden.
    Tillman, Tommy S.
    Howard, Rebecca J.
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab). KTH Royal Institute of Technology, Sweden.
    Lindahl, Erik
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab). KTH Royal Institute of Technology, Sweden.
    Xu, Yan
    Tang, Pei
    Structural Elucidation of Ivermectin Binding to α7nAChR and the Induced Channel Desensitization2023In: ACS Chemical Neuroscience, E-ISSN 1948-7193, Vol. 14, no 6, p. 1156-1165Article in journal (Refereed)
    Abstract [en]

    The α7 nicotinic acetylcholine receptor (α7nAChR) mediates signaling in the central nervous system and cholinergic anti-inflammatory pathways. Ivermectin is a positive allosteric modulator of a full-length α7nAChR and an agonist of the α7nAChR construct containing transmembrane (TMD) and intracellular (ICD) domains, but structural insights of the binding have not previously been determined. Here, combining nuclear magnetic resonance as a primary experimental tool with Rosetta comparative modeling and molecular dynamics simulations, we have revealed details of ivermectin binding to the α7nAChR TMD + ICD and corresponding structural changes in an ivermectin-induced desensitized state. Ivermectin binding was stabilized predominantly by hydrophobic interactions from interfacial residues between adjacent subunits near the extracellular end of the TMD, where the inter-subunit gap was substantially expanded in comparison to the apo structure. The ion-permeation pathway showed a profile distinctly different from the resting-state profile but similar to profiles of desensitized α7nAChR. The ICD also exhibited structural changes, including reorientation of the MX and h3 helices relative to the channel axis. The resulting structures of the α7nAChR TMD + ICD in complex with ivermectin provide opportunities for discovering new modulators of therapeutic potential and exploring the structural basis of cytoplasmic signaling under different α7nAChR functional states. 

  • 109. Bondarenko, Vasyl
    et al.
    Wells, Marta M.
    Chen, Qiang
    Tillman, Tommy S.
    Singewald, Kevin
    Lawless, Matthew J.
    Caporoso, Joel
    Brandon, Nicole
    Coleman, Jonathan A.
    Saxena, Sunil
    Lindahl, Erik
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab). KTH Royal Institute of Technology, Sweden.
    Xu, Yan
    Tang, Pei
    Structures of highly flexible intracellular domain of human α7 nicotinic acetylcholine receptor2022In: Nature Communications, E-ISSN 2041-1723, Vol. 13, article id 793Article in journal (Refereed)
    Abstract [en]

    The intracellular domain (ICD) of Cys-loop receptors mediates diverse functions. To date, no structure of a full-length ICD is available due to challenges stemming from its dynamic nature. Here, combining nuclear magnetic resonance (NMR) and electron spin resonance experiments with Rosetta computations, we determine full-length ICD structures of the human α7 nicotinic acetylcholine receptor in a resting state. We show that ~57% of the ICD residues are in highly flexible regions, primarily in a large loop (loop L) with the most mobile segment spanning ~50 Å from the central channel axis. Loop L is anchored onto the MA helix and virtually forms two smaller loops, thereby increasing its stability. Previously known motifs for cytoplasmic binding, regulation, and signaling are found in both the helices and disordered flexible regions, supporting the essential role of the ICD conformational plasticity in orchestrating a broad range of biological processes. 

  • 110.
    Bonnefille, Bénilde
    et al.
    Stockholm University, Faculty of Science, Department of Environmental Science. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Karlsson, Oskar
    Stockholm University, Science for Life Laboratory (SciLifeLab). Stockholm University, Faculty of Science, Department of Environmental Science.
    Rian, May Britt
    Stockholm University, Faculty of Science, Department of Environmental Science. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Raqib, Rubhana
    Parvez, Faruque
    Papazian, Stefano
    Stockholm University, Science for Life Laboratory (SciLifeLab). Stockholm University, Faculty of Science, Department of Environmental Science.
    Islam, M. Sirajul
    Martin, Jonathan W.
    Stockholm University, Faculty of Science, Department of Environmental Science. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Nontarget Analysis of Polluted Surface Waters in Bangladesh Using Open Science Workflows2023In: Environmental Science and Technology, ISSN 0013-936X, E-ISSN 1520-5851, Vol. 57, no 17, p. 6808-6824Article in journal (Refereed)
    Abstract [en]

    Nontarget mass spectrometry has great potential to reveal patterns of water contamination globally through community science, but few studies are conducted in low-income countries, nor with open-source workflows, and few datasets are FAIR (Findable, Accessible, Interoperable, Reusable). Water was collected from urban and rural rivers around Dhaka, Bangladesh, and analyzed by liquid chromatography high-resolution mass spectrometry in four ionization modes (electrospray ionization +/-, atmospheric pressure chemical ionization +/-) with data -independent MS2 acquisition. The acquisition strategy was complementary: 19,427 and 7365 features were unique to ESI and APCI, respectively. The complexity of water pollution was revealed by >26,000 unique molecular features resolved by MS-DIAL, among which >20,000 correlated with urban sources in Dhaka. A major wastewater treatment plant was not a dominant pollution source, consistent with major contributions from uncontrolled urban drainage, a result that encourages development of further wastewater infrastructures. Matching of deconvoluted MS2 spectra to public libraries resulted in 62 confident annotations (i.e., Level 1-2a) and allowed semiquantification of 42 analytes including pharmaceuticals, pesticides, and personal care products. In silico structure prediction for the top 100 unknown molecular features associated with an urban source allowed 15 additional chemicals of anthropogenic origin to be annotated (i.e., Level 3). The authentic MS2 spectra were uploaded to MassBank Europe, mass spectral data were openly shared on the MassIVE repository, a tool (i.e., MASST) that could be used for community science environmental surveillance was demonstrated, and current limitations were discussed.

  • 111. Borroto-Escuela, Dasiel O.
    et al.
    Rodriguez, David
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Romero-Fernandez, Wilber
    Kapla, Jon
    Jaiteh, Mariama
    Ranganathan, Anirudh
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Lazarova, Tzvetana
    Fuxe, Kjell
    Carlsson, Jens
    Mapping the Interface of a GPCR Dimer: A Structural Model of the A(2A) Adenosine and D-2 Dopamine Receptor Heteromer2018In: Frontiers in Pharmacology, E-ISSN 1663-9812, Vol. 9, article id 829Article in journal (Refereed)
    Abstract [en]

    The A(2A) adenosine (A(2A)R) and D-2 dopamine (D2R) receptors form oligomers in the cell membrane and allosteric interactions across the A(2A)R-D2R heteromer represent a target for development of drugs against central nervous system disorders. However, understanding of the molecular determinants of A(2A)R-D2R heteromerization and the allosteric antagonistic interactions between the receptor protomers is still limited. In this work, a structural model of the A(2A)R-D2R heterodimer was generated using a combined experimental and computational approach. Regions involved in the heteromer interface were modeled based on the effects of peptides derived from the transmembrane (TM) helices on A(2A)R-D2R receptor-receptor interactions in bioluminescence resonance energy transfer (BRET) and proximity ligation assays. Peptides corresponding to TM-IV and TM-V of the A(2A)R blocked heterodimer interactions and disrupted the allosteric effect of A(2A)R activation on D2R agonist binding. Protein-protein docking was used to construct a model of the A(2A)R-D2R heterodimer with a TM-IV/V interface, which was refined using molecular dynamics simulations. Mutations in the predicted interface reduced A(2A)R-D2R interactions in BRET experiments and altered the allosteric modulation. The heterodimer model provided insights into the structural basis of allosteric modulation and the technique developed to characterize the A(2A)R-D2R interface can be extended to study the many other G protein-coupled receptors that engage in heteroreceptor complexes.

  • 112. Borroto-Escuela, Dasiel O.
    et al.
    Wydra, Karolina
    Li, Xiang
    Rodriguez, David
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab). Uppsala University BMC, Sweden.
    Carlsson, Jens
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab). Uppsala University BMC, Sweden.
    Jastrzebska, Joanna
    Filip, Malgorzata
    Fuxe, Kjell
    Disruption of A2AR-D2R Heteroreceptor Complexes After A2AR Transmembrane 5 Peptide Administration Enhances Cocaine Self-Administration in Rats2018In: Molecular Neurobiology, ISSN 0893-7648, E-ISSN 1559-1182, Vol. 55, no 8, p. 7038-7048Article in journal (Refereed)
    Abstract [en]

    Antagonistic allosteric A2AR-D2R receptor-receptor interactions in heteroreceptor complexes counteract cocaine self-administration and cocaine seeking in rats as seen in biochemical and behavioral experiments. It was shown that the human A2AR transmembrane five (TM5) was part of the interface of the human A2AR-D2R receptor heteromer. In the current paper, the rat A2AR synthetic TM5 (synthTM5) peptide disrupts the A2AR-D2R heteroreceptor complex in HEK293 cells as shown by the bioluminescence resonance energy transfer method. Rat A2AR synthTM5 peptide, microinjected into the nucleus accumbens, produced a complete counteraction of the inhibitory effects of the A2AR agonist CGS21680 on cocaine self-administration. It was linked to a disappearance of the accumbal A2AR-D2R heteroreceptor complexes and the A2AR agonist induced inhibition of D2R recognition using proximity ligation assay and biochemical binding techniques. However, possible effects of the A2AR synthTM5 peptide on accumbal A2AR-D3R and A2AR-D4R heteroreceptor complexes remain to be excluded. Evidence is provided that accumbal A2AR-D2R-like heteroreceptor complexes with their antagonistic receptor-receptor interactions can be major targets for treatment of cocaine use disorder.

  • 113. Brack, P.
    et al.
    Crowther, P.
    Soiland-Reyes, S.
    Owen, S.
    Lowe, D.
    R. Williams, A.
    Groom, Q.
    Dillen, M.
    Coppens, F.
    Gruning, B.
    Eguinoa, I.
    Ewels, Philip
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Goble, C.
    Ten simple rules for making a software tool workflow-ready2022In: PloS Computational Biology, ISSN 1553-734X, E-ISSN 1553-7358, Vol. 18, no 3, article id e1009823Article in journal (Other academic)
  • 114. Bradley, Frideborg
    et al.
    Boger, Mathias Franzén
    Kaldhusdal, Vilde
    Åhlberg, Alexandra
    Edfeldt, Gabriella
    Lajoie, Julie
    Bergström, Sofia
    Omollo, Kenneth
    Damdimopoulos, Anastasios
    Czarnewski, Paulo
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Månberg, Anna
    Oyugi, Julius
    Kimani, Joshua
    Nilsson, Peter
    Fowke, Keith
    Tjernlund, Annelie
    Broliden, Kristina
    Multi-omics analysis of the cervical epithelial integrity of women using depot medroxyprogesterone acetate2022In: PLoS Pathogens, ISSN 1553-7366, E-ISSN 1553-7374, Vol. 18, no 5, article id e1010494Article in journal (Refereed)
    Abstract [en]

    Depot medroxyprogesterone acetate (DMPA) is an injectable hormonal contraceptive used by millions of women worldwide. However, experimental studies have associated DMPA use with genital epithelial barrier disruption and mucosal influx of human immunodeficiency virus (HIV) target cells. We explored the underlying molecular mechanisms of these findings. Ectocervical biopsies and cervicovaginal lavage (CVL) specimens were collected from HIV-seronegative Kenyan sex workers using DMPA (n = 32) or regularly cycling controls (n = 64). Tissue samples were assessed by RNA-sequencing and quantitative imaging analysis, whereas protein levels were measured in CVL samples. The results suggested a DMPA-associated upregulation of genes involved in immune regulation, including genes associated with cytokine-mediated signaling and neutrophil-mediated immunity. A transcription factor analysis further revealed DMPA-associated upregulation of RELA and NFKB1 which are involved in several immune activation pathways. Several genes significantly downregulated in the DMPA versus the control group were involved in epithelial structure and function, including genes encoding keratins, small proline-rich proteins, and cell-cell adhesion proteins. Pathway analyses indicated DMPA use was associated with immune activation and suppression of epithelium development, including keratinization and cornification processes. The cervicovaginal microbiome composition (Lactobacillus dominant and non-Lactobacillus dominant) had no overall interactional impact on the DMPA associated tissue gene expression. Imaging analysis verified that DMPA use was associated with an impaired epithelial layer as illustrated by staining for the selected epithelial junction proteins E-cadherin, desmoglein-1 and claudin-1. Additional staining for CD4+ cells revealed a more superficial location of these cells in the ectocervical epithelium of DMPA users versus controls. Altered protein levels of SERPINB1 and ITIH2 were further observed in the DMPA group. Identification of specific impaired epithelial barrier structures at the gene expression level, which were verified at the functional level by tissue imaging analysis, illustrates mechanisms by which DMPA adversely may affect the integrity of the genital mucosa.

  • 115. Branca, Rui M. M.
    et al.
    Orre, Lukas M.
    Johansson, Henrik J.
    Granholm, Viktor
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Huss, Mikael
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Perez-Bercoff, Åsa
    Forshed, Jenny
    Käll, Lukas
    Lehtio, Janne
    HiRIEF LC-MSMS enables deep proteome coverage and unbiased proteogenomics2014In: Nature Methods, ISSN 1548-7091, E-ISSN 1548-7105, Vol. 11, no 1, p. 59-+Article in journal (Refereed)
    Abstract [en]

    We present a liquid chromatography-mass spectrometry (LC-MSMS)-based method permitting unbiased (gene prediction-independent) genome-wide discovery of protein-coding loci in higher eukaryotes. Using high-resolution isoelectric focusing (HiRIEF) at the peptide level in the 3.7-5.0 pH range and accurate peptide isoelectric point (pI) prediction, we probed the six-reading-frame translation of the human and mouse genomes and identified 98 and 52 previously undiscovered protein-coding loci, respectively. The method also enabled deep proteome coverage, identifying 13,078 human and 10,637 mouse proteins.

  • 116. Brandão, Luiz Eduardo Mateus
    et al.
    Espes, Daniel
    Orzechowski Westholm, Jakub
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Martikainen, Teemu
    Westerlund, Nestori
    Lampola, Lauri
    Popa, Alexandru
    Vogel, Heike
    Schürmann, Annette
    Dickson, Suzanne L.
    Benedict, Christian
    Cedernaes, Jonathan
    Acute sleep loss alters circulating fibroblast growth factor 21 levels in humans: A randomised crossover trial2022In: Journal of Sleep Research, ISSN 0962-1105, E-ISSN 1365-2869, Vol. 31, no 2, article id e13472Article in journal (Refereed)
    Abstract [en]

    The hormone fibroblast growth factor 21 (FGF21) modulates tissue metabolism and circulates at higher levels in metabolic conditions associated with chronic sleep–wake disruption, such as type 2 diabetes and obesity. In the present study, we investigated whether acute sleep loss impacts circulating levels of FGF21 and tissue-specific production, and response pathways linked to FGF21. A total of 15 healthy normal-weight young men participated in a randomised crossover study with two conditions, sleep loss versus an 8.5-hr sleep window. The evening before each intervention, fasting blood was collected. Fasting, post-intervention morning skeletal muscle and adipose tissue samples underwent quantitative polymerase chain reaction and DNA methylation analyses, and serum FGF21 levels were measured before and after an oral glucose tolerance test. Serum levels of FGF21 were higher after sleep loss compared with sleep, both under fasting conditions and following glucose intake (~27%–30%, p = 0.023). Fasting circulating levels of fibroblast activation protein, a protein which can degrade circulating FGF21, were not altered by sleep loss, whereas DNA methylation in the FGF21 promoter region increased only in adipose tissue. However, even though specifically the muscle exhibited transcriptional changes indicating adverse alterations to redox and metabolic homeostasis, no tissue-based changes were observed in expression of FGF21, its receptors, or selected signalling targets, in response to sleep loss. In summary, we found that acute sleep loss resulted in increased circulating levels of FGF21 in healthy young men, which may occur independent of a tissue-based stress response in metabolic peripheral tissues. Further studies may decipher whether changes in FGF21 signalling after sleep loss modulate metabolic outcomes associated with sleep or circadian disruption.

  • 117. Brate, Jon
    et al.
    Neumann, Ralf S.
    Fromm, Bastian
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute. Stockholm University, Science for Life Laboratory (SciLifeLab). Oslo University Hospital, Norway.
    Haraldsen, Arthur A. B.
    Tarver, James E.
    Suga, Hiroshi
    Donoghue, Philip C. J.
    Peterson, Kevin J.
    Ruiz-Trillo, Inaki
    Grini, Paul E.
    Shalchian-Tabrizi, Kamran
    Unicellular Origin of the Animal MicroRNA Machinery2018In: Current Biology, ISSN 0960-9822, E-ISSN 1879-0445, Vol. 28, no 20, p. 3288-+Article in journal (Refereed)
    Abstract [en]

    The emergence of multicellular animals was associated with an increase in phenotypic complexity and with the acquisition of spatial cell differentiation and embryonic development. Paradoxically, this phenotypic transition was not paralleled by major changes in the underlying developmental toolkit and regulatory networks. In fact, most of these systems are ancient, established already in the unicellular ancestors of animals [1-5]. In contrast, the Microprocessor protein machinery, which is essential for microRNA (miRNA) biogenesis in animals, as well as the miRNA genes themselves produced by this Microprocessor, have not been identified outside of the animal kingdom [6]. Hence, the Microprocessor, with the key proteins Pasha and Drosha, is regarded as an animal innovation [7-9]. Here, we challenge this evolutionary scenario by investigating unicellular sister lineages of animals through genomic and transcriptomic analyses. We identify in Ichthyosporea both Drosha and Pasha (DGCR8 in vertebrates), indicating that the Microprocessor complex evolved long before the last common ancestor of animals, consistent with a pre-metazoan origin of most of the animal developmental gene elements. Through small RNA sequencing, we also discovered expressed bona fide miRNA genes in several species of the ichthyosporeans harboring the Microprocessor. A deep, pre-metazoan origin of the Microprocessor and miRNAs comply with a view that the origin of multicellular animals was not directly linked to the innovation of these key regulatory components.

  • 118.
    Brindefalk, Björn
    et al.
    Stockholm University, Faculty of Science, Department of Ecology, Environment and Plant Sciences. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Ekman, Martin
    Stockholm University, Faculty of Science, Department of Ecology, Environment and Plant Sciences. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Ininbergs, Karolina
    Stockholm University, Faculty of Science, Department of Ecology, Environment and Plant Sciences. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Dupont, Christopher L.
    Yooseph, Shibu
    Pinhassi, Jarone
    Bergman, Birgitta
    Stockholm University, Faculty of Science, Department of Ecology, Environment and Plant Sciences. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Distribution and expression of microbial rhodopsins in the Baltic Sea and adjacent waters2016In: Environmental Microbiology, ISSN 1462-2912, E-ISSN 1462-2920, Vol. 18, no 12, p. 4442-4455Article in journal (Refereed)
    Abstract [en]

    Rhodopsins are light-driven ion-pumping membrane proteins found in many organisms and are proposed to be of global importance for oceanic microbial energy generation. Several studies have focused on marine environments, with less exploration of rhodopsins in brackish waters. We investigated microbial rhodopsins in the Baltic Sea using size-fractionated metagenomic and metatranscriptomic datasets collected along a salinity gradient spanning from similar to 0 to 35 PSU. The normalised genomic abundance of rhodopsins in Bacteria, as well as rhodopsin gene expression, was highest in the smallest size fraction (0.1-0.8 mu m), relative to the medium (0.8-3.0 mu m) and large (> 3.0 mu m) size fractions. The abundance of rhodopsins in the two smaller size fractions displayed a positive correlation with salinity. Proteobacteria and Bacteroidetes rhodopsins were the most abundant while Actinobacteria rhodopsins, or actinorhodopsins, were common at lower salinities. Phylogenetic analysis indicated that rhodopsins have adapted independently to the marine-brackish transition on multiple occasions, giving rise to green light-adapted variants from ancestral blue light-adapted ones. A notable diversity of viral-like rhodopsins was also detected in the dataset and potentially linked with eukaryotic phytoplankton blooms. Finally, a new clade of likely proton-pumping rhodopsin with non-canonical amino acids in the spectral tuning and proton accepting site was identified.

  • 119. Briones, Rodolfo
    et al.
    Blau, Christian
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Kutzner, Carsten
    de Groot, Bert L.
    Aponte-Santamaria, Camilo
    GROma rho s: A GROMACS-Based Toolset to Analyze Density Maps Derived from Molecular Dynamics Simulations2019In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 116, no 1, p. 4-11Article in journal (Refereed)
    Abstract [en]

    We introduce a computational toolset, named GROma rho s, to obtain and compare time-averaged density maps from molecular dynamics simulations. GROma rho s efficiently computes density maps by fast multi-Gaussian spreading of atomic densities onto a three-dimensional grid. It complements existing map-based tools by enabling spatial inspection of atomic average localization during the simulations. Most importantly, it allows the comparison between computed and reference maps (e.g., experimental) through calculation of difference maps and local and time-resolved global correlation. These comparison operations proved useful to quantitatively contrast perturbed and control simulation data sets and to examine how much biomolecular systems resemble both synthetic and experimental density maps. This was especially advantageous for multimolecule systems in which standard comparisons like RMSDs are difficult to compute. In addition, GROma rho s incorporates absolute and relative spatial free-energy estimates to provide an energetic picture of atomistic localization. This is an open-source GROMACS-based toolset, thus allowing for static or dynamic selection of atoms or even coarse-grained beads for the density calculation. Furthermore, masking of regions was implemented to speed up calculations and to facilitate the comparison with experimental maps. Beyond map comparison, GROma rho s provides a straightforward method to detect solvent cavities and average charge distribution in biomolecular systems. We employed all these functionalities to inspect the localization of lipid and water molecules in aquaporin systems, the binding of cholesterol to the G protein coupled chemokine receptor type 4, and the identification of permeation pathways through the dermicidin antimicrobial channel. Based on these examples, we anticipate a high applicability of GROma rho s for the analysis of molecular dynamics simulations and their comparison with experimentally determined densities.

  • 120. Brocke, Ekaterina
    et al.
    Bhalla, Upinder S.
    Djurfeldt, Mikael
    Hellgren Kotaleski, Jeanette
    Stockholm University, Science for Life Laboratory (SciLifeLab). Stockholm University, Faculty of Science, Numerical Analysis and Computer Science (NADA). KTH Royal Institute of Technology, Sweden; Karolinska Institute, Sweden.
    Hanke, Michael
    Efficient Integration of Coupled Electrical-Chemical Systems in Multiscale Neuronal Simulations2016In: Frontiers in Computational Neuroscience, E-ISSN 1662-5188, Vol. 10, article id 97Article in journal (Refereed)
    Abstract [en]

    Multiscale modeling and simulations in neuroscience is gaining scientific attention due to its growing importance and unexplored capabilities. For instance, it can help to acquire better understanding of biological phenomena that have important features at multiple scales of time and space. This includes synaptic plasticity, memory formation and modulation, homeostasis. There are several ways to organize multiscale simulations depending on the scientific problem and the system to be modeled. One of the possibilities is to simulate different components of a multiscale system simultaneously and exchange data when required. The latter may become a challenging task for several reasons. First, the components of a multiscale system usually span different spatial and temporal scales, such that rigorous analysis of possible coupling solutions is required. Then, the components can be defined by different mathematical formalisms. For certain classes of problems a number of coupling mechanisms have been proposed and successfully used. However, a strict mathematical theory is missing in many cases. Recent work in the field has not so far investigated artifacts that may arise during coupled integration of different approximation methods. Moreover, in neuroscience, the coupling of widely used numerical fixed step size solvers may lead to unexpected inefficiency. In this paper we address the question of possible numerical artifacts that can arise during the integration of a coupled system. We develop an efficient strategy to couple the components comprising a multiscale test problem in neuroscience. We introduce an efficient coupling method based on the second-order backward differentiation formula (BDF2) numerical approximation. The method uses an adaptive step size integration with an error estimation proposed by Skelboe (2000). The method shows a significant advantage over conventional fixed step size solvers used in neuroscience for similar problems. We explore different coupling strategies that define the organization of computations between system components. We study the importance of an appropriate approximation of exchanged variables during the simulation. The analysis shows a substantial impact of these aspects on the solution accuracy in the application to our multiscale neuroscientific test problem. We believe that the ideas presented in the paper may essentially contribute to the development of a robust and efficient framework for multiscale brain modeling and simulations in neuroscience.

  • 121. Brodin, Johanna
    et al.
    Mild, Mattias
    Hedskog, Charlotte
    Sherwood, Ellen
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Leitner, Thomas
    Andersson, Bjoern
    Albert, Jan
    PCR-Induced Transitions Are the Major Source of Error in Cleaned Ultra-Deep Pyrosequencing Data2013In: PLOS ONE, E-ISSN 1932-6203, Vol. 8, no 7, article id e70388Article in journal (Refereed)
    Abstract [en]

    Background: Ultra-deep pyrosequencing (UDPS) is used to identify rare sequence variants. The sequence depth is influenced by several factors including the error frequency of PCR and UDPS. This study investigated the characteristics and source of errors in raw and cleaned UDPS data. Results: UDPS of a 167-nucleotide fragment of the HIV-1 SG3Denv plasmid was performed on the Roche/454 platform. The plasmid was diluted to one copy, PCR amplified and subjected to bidirectional UDPS on three occasions. The dataset consisted of 47,693 UDPS reads. Raw UDPS data had an average error frequency of 0.30% per nucleotide site. Most errors were insertions and deletions in homopolymeric regions. We used a cleaning strategy that removed almost all indel errors, but had little effect on substitution errors, which reduced the error frequency to 0.056% per nucleotide. In cleaned data the error frequency was similar in homopolymeric and non-homopolymeric regions, but varied considerably across sites. These site-specific error frequencies were moderately, but still significantly, correlated between runs (r = 0.15-0.65) and between forward and reverse sequencing directions within runs (r = 0.33-0.65). Furthermore, transition errors were 48-times more common than transversion errors (0.052% vs. 0.001%; p<0.0001). Collectively the results indicate that a considerable proportion of the sequencing errors that remained after data cleaning were generated during the PCR that preceded UDPS. Conclusions: A majority of the sequencing errors that remained after data cleaning were introduced by PCR prior to sequencing, which means that they will be independent of platform used for next-generation sequencing. The transition vs. transversion error bias in cleaned UDPS data will influence the detection limits of rare mutations and sequence variants.

  • 122.
    Broman, Elias
    et al.
    Stockholm University, Faculty of Science, Department of Ecology, Environment and Plant Sciences. Stockholm University, Faculty of Science, Stockholm University Baltic Sea Centre.
    Izabel-Shen, Dandan
    Stockholm University, Faculty of Science, Department of Ecology, Environment and Plant Sciences.
    Rodríguez-Gijón, Alejandro
    Stockholm University, Faculty of Science, Department of Ecology, Environment and Plant Sciences. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Bonaglia, Stefano
    Garcia, Sarahi L.
    Stockholm University, Faculty of Science, Department of Ecology, Environment and Plant Sciences. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Nascimento, Francisco J. A.
    Stockholm University, Faculty of Science, Department of Ecology, Environment and Plant Sciences. Stockholm University, Faculty of Science, Stockholm University Baltic Sea Centre.
    Microbial functional genes are driven by gradients in sediment stoichiometry, oxygen, and salinity across the Baltic benthic ecosystem2022In: Microbiome, E-ISSN 2049-2618, Vol. 10, no 126, p. 1-17Article in journal (Refereed)
    Abstract [en]

    Background: Microorganisms in the seafoor use a wide range of metabolic processes, which are coupled to thepresence of functional genes within their genomes. Aquatic environments are heterogenous and often characterizedby natural physiochemical gradients that structure these microbial communities potentially changing the diversityof functional genes and its associated metabolic processes. In this study, we investigated spatial variability and howenvironmental variables structure the diversity and composition of benthic functional genes and metabolic pathwaysacross various fundamental environmental gradients. We analyzed metagenomic data from sediment samples, meas‑ured related abiotic data (e.g., salinity, oxygen and carbon content), covering 59 stations spanning 1,145 km across theBaltic Sea.

    Results: The composition of genes and microbial communities were mainly structured by salinity plus oxygen, andthe carbon to nitrogen (C:N) ratio for specifc metabolic pathways related to nutrient transport and carbon metabo‑lism. Multivariate analyses indicated that the compositional change in functional genes was more prominent acrossenvironmental gradients compared to changes in microbial taxonomy even at genus level, and indicate functionaldiversity adaptation to local environments. Oxygen defcient areas (i.e., dead zones) were more diferent in gene com‑position when compared to oxic sediments.

    Conclusions: This study highlights how benthic functional genes are structured over spatial distances and by envi‑ronmental gradients and resource availability, and suggests that changes in, e.g., oxygenation, salinity, and carbonplus nitrogen content will infuence functional metabolic pathways in benthic habitats

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  • 123.
    Bromstrup, Torben
    et al.
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Howard, Rebecca J.
    Trudell, James R.
    Harris, R. Adron
    Lindahl, Erik
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Inhibition versus Potentiation of Ligand-Gated Ion Channels Can Be Altered by a Single Mutation that Moves Ligands between Intra- and Intersubunit Sites2013In: Structure, ISSN 0969-2126, E-ISSN 1878-4186, Vol. 21, no 8, p. 1307-1316Article in journal (Refereed)
    Abstract [en]

    Pentameric ligand-gated ion channels (pLGICs) are similar in structure but either inhibited or potentiated by alcohols and anesthetics. This dual modulation has previously not been understood, but the determination of X-ray structures of prokaryotic GLIC provides an ideal model system. Here, we show that a single-site mutation at the F14' site in the GLIC transmembrane domain turns desflurane and chloroform from inhibitors to potentiators, and that this is explained by competing allosteric sites. The F14'A mutation opens an intersubunit site lined by N239 (15'), 1240 (16'), and Y263. Free energy calculations confirm this site is the preferred binding location for desflurane and chloroform in GLIC F14'A. In contrast, both anesthetics prefer an intrasubunit site in wild-type GLIC. Modulation is therefore the net effect of competitive binding between the intersubunit potentiating site and an intrasubunit inhibitory site. This provides direct evidence for a dual-site model of allosteric regulation of pLGICs.

  • 124. Brown, Alan
    et al.
    Rathore, Sorbhi
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Kimanius, Dari
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Aibara, Shintaro
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Bai, Xiao-chen
    Rorbach, Joanna
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab). Karolinska Institutet, Sweden.
    Amunts, Alexey
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab). MRC Laboratory of Molecular Biology, UK.
    Ramakrishnan, V.
    Structures of the human mitochondrial ribosome in native states of assembly2017In: Nature Structural & Molecular Biology, ISSN 1545-9993, E-ISSN 1545-9985, Vol. 24, no 10, p. 866-869Article in journal (Refereed)
    Abstract [en]

    Mammalian mitochondrial ribosomes (mitoribosomes) have less rRNA content and 36 additional proteins compared with the evolutionarily related bacterial ribosome. These differences make the assembly of mitoribosomes more complex than the assembly of bacterial ribosomes, but the molecular details of mitoribosomal biogenesis remain elusive. Here, we report the structures of two late-stage assembly intermediates of the human mitoribosomal large subunit (mt-LSU) isolated from a native pool within a human cell line and solved by cryo-EM to similar to 3-angstrom resolution. Comparison of the structures reveals insights into the timing of rRNA folding and protein incorporation during the final steps of ribosomal maturation and the evolutionary adaptations that are required to preserve biogenesis after the structural diversification of mitoribosomes. Furthermore, the structures redefine the ribosome silencing factor (RsfS) family as multifunctional biogenesis factors and identify two new assembly factors (L0R8F8 and mt-ACP) not previously implicated in mitoribosomal biogenesis.

  • 125. Bruhn-Olszewska, Bożena
    et al.
    Davies, Hanna
    Sarkisyan, Daniil
    Juhas, Ulana
    Rychlicka-Buniowska, Edyta
    Wójcik, Magdalena
    Horbacz, Monika
    Jąkalski, Marcin
    Olszewski, Paweł
    Orzechowski Westholm, Jakub
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Smialowska, Agata
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Wierzba, Karol
    Naluai, Åsa Torinsson
    Jern, Niklas
    Andersson, Lars-Magnus
    Järhult, Josef D.
    Filipowicz, Natalia
    Janson, Eva Tiensuu
    Rubertsson, Sten
    Lipcsey, Miklós
    Gisslén, Magnus
    Hultström, Michael
    Frithiof, Robert
    Dumanski, Jan P.
    Loss of Y in leukocytes as a risk factor for critical COVID-19 in men2022In: Genome Medicine, ISSN 1756-994X, E-ISSN 1756-994X, Vol. 14, no 1, article id 139Article in journal (Refereed)
    Abstract [en]

    Background: The COVID-19 pandemic, which has a prominent social and economic impact worldwide, shows a largely unexplained male bias for the severity and mortality of the disease. Loss of chromosome Y (LOY) is a risk factor candidate in COVID-19 due to its prior association with many chronic age-related diseases, and its impact on immune gene transcription.

    Methods: Publicly available scRNA-seq data of PBMC samples derived from male patients critically ill with COVID-19 were reanalyzed, and LOY status was added to the annotated cells. We further studied LOY in whole blood for 211 COVID-19 patients treated at intensive care units (ICU) from the first and second waves of the pandemic. Of these, 139 patients were subject to cell sorting for LOY analysis in granulocytes, low-density neutrophils (LDNs), monocytes, and PBMCs.

    Results: Reanalysis of available scRNA-seq data revealed LDNs and monocytes as the cell types most affected by LOY. Subsequently, DNA analysis indicated that 46%, 32%, and 29% of critically ill patients showed LOY above 5% cut-off in LDNs, granulocytes, and monocytes, respectively. Hence, the myeloid lineage that is crucial for the development of severe COVID-19 phenotype is affected by LOY. Moreover, LOY correlated with increasing WHO score (median difference 1.59%, 95% HDI 0.46% to 2.71%, p=0.025), death during ICU treatment (median difference 1.46%, 95% HDI 0.47% to 2.43%, p=0.0036), and history of vessel disease (median difference 2.16%, 95% HDI 0.74% to 3.7%, p=0.004), among other variables. In 16 recovered patients, sampled during ICU stay and 93–143 days later, LOY decreased significantly in whole blood and PBMCs. Furthermore, the number of LDNs at the recovery stage decreased dramatically (median difference 76.4 per 10,000 cell sorting events, 95% HDI 55.5 to 104, p=6e−11).

    Conclusions: We present a link between LOY and an acute, life-threatening infectious disease. Furthermore, this study highlights LOY as the most prominent clonal mutation affecting the myeloid cell lineage during emergency myelopoiesis. The correlation between LOY level and COVID-19 severity might suggest that this mutation affects the functions of monocytes and neutrophils, which could have consequences for male innate immunity.

  • 126.
    Bryant, Patrick
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Deep learning for protein complex structure prediction2023In: Current opinion in structural biology, ISSN 0959-440X, E-ISSN 1879-033X, Vol. 79, article id 102529Article in journal (Refereed)
    Abstract [en]

    Recent developments in the structure prediction of protein complexes have resulted in accuracies rivalling experimental methods in many cases. The high accuracy is mainly observed in dimeric complexes and other problems such as protein disorder and predicting the structure of host-pathogen in-teractions remain. This review highlights the foundation for current accurate structure prediction of protein complexes and possible ways to address the remaining limitations.

  • 127.
    Bryant, Patrick
    et al.
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Elofsson, Arne
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Decomposing Structural Response Due to Sequence Changes in Protein Domains with Machine Learning2020In: Journal of Molecular Biology, ISSN 0022-2836, E-ISSN 1089-8638, Vol. 432, no 16, p. 4435-4446Article in journal (Refereed)
    Abstract [en]

    How protein domain structure changes in response to mutations is not well understood. Some mutations change the structure drastically, while most only result in small changes. To gain an understanding of this, we decompose the relationship between changes in domain sequence and structure using machine learning. We select pairs of evolutionarily related domains with a broad range of evolutionary distances. In contrast to earlier studies, we do not find a strictly linear relationship between sequence and structural changes. We train a random forest regressor that predicts the structural similarity between pairs with an average accuracy of 0.029 IDDT ( local Distance Difference Test) score, and a correlation coefficient of 0.92. Decomposing the feature importance shows that the domain length, or analogously, size is the most important feature. Our model enables assessing deviations in relative structural response, and thus prediction of evolutionary trajectories, in protein domains across evolution.

  • 128.
    Bryant, Patrick
    et al.
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Elofsson, Arne
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Estimating the impact of mobility patterns on COVID-19 infection rates in 11 European countries2020In: PeerJ, E-ISSN 2167-8359, Vol. 8, article id e9879Article in journal (Refereed)
    Abstract [en]

    Background: As governments across Europe have issued non-pharmaceutical interventions (NPIs) such as social distancing and school closing, the mobility patterns in these countries have changed. Most states have implemented similar NPIs at similar time points. However, it is likely different countries and populations respond differently to the NPIs and that these differences cause mobility patterns and thereby the epidemic development to change.

    Methods: We build a Bayesian model that estimates the number of deaths on a given day dependent on changes in the basic reproductive number, R-0, due to differences in mobility patterns. We utilise mobility data from Google mobility reports using five different categories: retail and recreation, grocery and pharmacy, transit stations, workplace and residential. The importance of each mobility category for predicting changes in R-0 is estimated through the model.

    Findings: The changes in mobility have a considerable overlap with the introduction of governmental NPIs, highlighting the importance of government action for population behavioural change. The shift in mobility in all categories shows high correlations with the death rates 1 month later. Reduction of movement within the grocery and pharmacy sector is estimated to account for most of the decrease in R-0.

    Interpretation: Our model predicts 3-week epidemic forecasts, using real-time observations of changes in mobility patterns, which can provide governments with direct feedback on the effects of their NPIs. The model predicts the changes in a majority of the countries accurately but overestimates the impact of NPIs in Sweden and Denmark and underestimates them in France and Belgium. We also note that the exponential nature of all epidemiological models based on the basic reproductive number, R-0 cause small errors to have extensive effects on the predicted outcome.

  • 129.
    Bryant, Patrick
    et al.
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Elofsson, Arne
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Peptide binder design with inverse folding and protein structure prediction2023In: Communications Chemistry, E-ISSN 2399-3669, Vol. 6, no 1, article id 229Article in journal (Refereed)
    Abstract [en]

    The computational design of peptide binders towards a specific protein interface can aid diagnostic and therapeutic efforts. Here, we design peptide binders by combining the known structural space searched with Foldseek, the protein design method ESM-IF1, and AlphaFold2 (AF) in a joint framework. Foldseek generates backbone seeds for a modified version of ESM-IF1 adapted to protein complexes. The resulting sequences are evaluated with AF using an MSA representation for the receptor structure and a single sequence for the binder. We show that AF can accurately evaluate protein binders and that our bind score can select these (ROC AUC = 0.96 for the heterodimeric case). We find that designs created from seeds with more contacts per residue are more successful and tend to be short. There is a relationship between the sequence recovery in interface positions and the plDDT of the designs, where designs with >= 80% recovery have an average plDDT of 84 compared to 55 at 0%. Designed sequences have 60% higher median plDDT values towards intended receptors than non-intended ones. Successful binders (predicted interface RMSD <= 2 angstrom) are designed towards 185 (6.5%) heteromeric and 42 (3.6%) homomeric protein interfaces with ESM-IF1 compared with 18 (1.5%) using ProteinMPNN from 100 samples. Designing peptides that bind to specific protein targets is crucial for peptidic drug development, however, traditional computer-aided binder design is outperformed by AlphaFold2. Here, the authors develop a peptide binder designing tool by combining Foldseek, ESM-IF1 and AlphaFold2 to increase the success rate.

  • 130.
    Bryant, Patrick
    et al.
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Elofsson, Arne
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    The relationship between ageing and changes in the human blood and brain methylomes 2022In: NAR Genomics and Bioinformatics, E-ISSN 2631-9268, Vol. 4, no 1, article id lqac001Article in journal (Refereed)
    Abstract [en]

    Changes in DNA methylation have been found to be strongly correlated with age, enabling the creation of ‘epigenetic clocks’. Previously, studies on the relationship between ageing and DNA methylation have assumed a linear relationship. Here, we show that several markers show a non-linear behaviour. In particular, we observe a tendency for saturation with age, especially in the cerebellum. Further, we show that the relationships between significant methylation changes and ageing are different in different tissues. We suggest a straightforward method of assessing all methylation-age relationships and cluster them according to their relative fold change. Our fold change selection outperforms the most common epigenetic clocks in predicting age for the cerebellum, but not for Blood or the Frontal Cortex. Further, we find that the saturation of methylation observed at older ages for the cerebellum explains why epigenetic clocks consistently underestimate the age there. The findings imply that assuming linear correlations might cause biologically important markers to be missed. 

  • 131.
    Bryant, Patrick
    et al.
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Pozzati, Gabriele
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Elofsson, Arne
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Improved prediction of protein-protein interactions using AlphaFold22022In: Nature Communications, E-ISSN 2041-1723, Vol. 13, no 1, article id 1265Article in journal (Refereed)
    Abstract [en]

    Predicting the structure of interacting protein chains is a fundamental step towards understanding protein function. Unfortunately, no computational method can produce accurate structures of protein complexes. AlphaFold2, has shown unprecedented levels of accuracy in modelling single chain protein structures. Here, we apply AlphaFold2 for the prediction of heterodimeric protein complexes. We find that the AlphaFold2 protocol together with optimised multiple sequence alignments, generate models with acceptable quality (DockQ >= 0.23) for 63% of the dimers. From the predicted interfaces we create a simple function to predict the DockQ score which distinguishes acceptable from incorrect models as well as interacting from non-interacting proteins with state-of-art accuracy. We find that, using the predicted DockQ scores, we can identify 51% of all interacting pairs at 1% FPR. Predicting the structure of protein complexes is extremely difficult. Here, authors apply AlphaFold2 with optimized multiple sequence alignments to model complexes of interacting proteins, enabling prediction of both if and how proteins interact with state-of-art accuracy.

  • 132.
    Bryant, Patrick
    et al.
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Pozzati, Gabriele
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Zhu, Wensi
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Shenoy, Aditi
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Kundrotas, Petras
    Elofsson, Arne
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Predicting the structure of large protein complexes using AlphaFold and Monte Carlo tree search2022In: Nature Communications, E-ISSN 2041-1723, Vol. 13, no 1, article id 6028Article in journal (Refereed)
    Abstract [en]

    AlphaFold can predict the structure of single- and multiple-chain proteins with very high accuracy. However, the accuracy decreases with the number of chains, and the available GPU memory limits the size of protein complexes which can be predicted. Here we show that one can predict the structure of large complexes starting from predictions of subcomponents. We assemble 91 out of 175 complexes with 10–30 chains from predicted subcomponents using Monte Carlo tree search, with a median TM-score of 0.51. There are 30 highly accurate complexes (TM-score ≥0.8, 33% of complete assemblies). We create a scoring function, mpDockQ, that can distinguish if assemblies are complete and predict their accuracy. We find that complexes containing symmetry are accurately assembled, while asymmetrical complexes remain challenging. The method is freely available and accesible as a Colab notebook https://colab.research.google.com/github/patrickbryant1/MoLPC/blob/master/MoLPC.ipynb.

  • 133.
    Bryant, Patrick
    et al.
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab). Karolinska Institutet, Sweden.
    Walton Bernstedt, Sophie
    Thutkawkorapin, Jessada
    Backman, Ann-Sofie
    Lindblom, Annika
    Lagerstedt-Robinson, Kristina
    Exome sequencing in a Swedish family with PMS2 mutation with varying penetrance of colorectal cancer: investigating the presence of genetic risk modifiers in colorectal cancer risk2023In: European Journal of Cancer Prevention, ISSN 0959-8278, E-ISSN 1473-5709, Vol. 32, no 2, p. 113-118Article in journal (Refereed)
    Abstract [en]

    Objective  Lynch syndrome is caused by germline mutations in the mismatch repair (MMR) genes, such as the PMS2 gene, and is characterised by a familial accumulation of colorectal cancer. The penetrance of cancer in PMS2 carriers is still not fully elucidated as a colorectal cancer risk has been shown to vary between PMS2 carriers, suggesting the presence of risk modifiers.

    Methods  Whole exome sequencing was performed in a Swedish family carrying a PMS2 missense mutation [c.2113G>A, p.(Glu705Lys)]. Thirteen genetic sequence variants were further selected and analysed in a case-control study (724 cases and 711 controls).

    Results  The most interesting variant was an 18 bp deletion in gene BAG1. BAG1 has been linked to colorectal tumour progression with poor prognosis and is thought to promote colorectal tumour cell survival through increased NF-κB activity.

    Conclusions  We conclude the genetic architecture behind the incomplete penetrance of PMS2 is complicated and must be assessed in a genome wide manner using large families and multifactorial analysis.

  • 134. Buck, Moritz
    et al.
    Garcia, Sarahi L.
    Stockholm University, Faculty of Science, Department of Ecology, Environment and Plant Sciences. Stockholm University, Science for Life Laboratory (SciLifeLab). Uppsala University, Sweden.
    Fernandez, Leyden
    Martin, Gaëtan
    Martinez-Rodriguez, Gustavo
    Saarenheimo, Jatta
    Zopfi, Jakob
    Bertilsson, Stefan
    Peura, Sari
    Comprehensive dataset of shotgun metagenomes from oxygen stratified freshwater lakes and ponds2021In: Scientific Data, E-ISSN 2052-4463, Vol. 8, no 1, article id 131Article in journal (Refereed)
    Abstract [en]

    Stratified lakes and ponds featuring steep oxygen gradients are significant net sources of greenhouse gases and hotspots in the carbon cycle. Despite their significant biogeochemical roles, the microbial communities, especially in the oxygen depleted compartments, are poorly known. Here, we present a comprehensive dataset including 267 shotgun metagenomes from 41 stratified lakes and ponds mainly located in the boreal and subarctic regions, but also including one tropical reservoir and one temperate lake. For most lakes and ponds, the data includes a vertical sample set spanning from the oxic surface to the anoxic bottom layer. The majority of the samples were collected during the open water period, but also a total of 29 samples were collected from under the ice. In addition to the metagenomic sequences, the dataset includes environmental variables for the samples, such as oxygen, nutrient and organic carbon concentrations. The dataset is ideal for further exploring the microbial taxonomic and functional diversity in freshwater environments and potential climate change impacts on the functioning of these ecosystems.

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  • 135. Buetti-Dinh, Antoine
    et al.
    Dethlefsen, Olga
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Friedman, Ran
    Dopson, Mark
    Transcriptomic analysis reveals how a lack of potassium ions increases Sulfolobus acidocaldarius sensitivity to pH changes2016In: Microbiology, ISSN 1350-0872, E-ISSN 1465-2080, Vol. 162, no 8, p. 1422-1434Article in journal (Refereed)
    Abstract [en]

    Extremely acidophilic microorganisms (optimum growth pH of <= 3) maintain a near neutral cytoplasmic pH via several homeostatic mechanisms, including an inside positive membrane potential created by potassium ions. Transcriptomic responses to pH stress in the thermoacidophilic archaeon, Sulfolobus acidocaldarius were investigated by growing cells without added sodium and/or potassium ions at both optimal and sub-optimal pH. Culturing the cells in the absence of added sodium or potassium ions resulted in a reduced growth rate compared to full-salt conditions as well as 43 and 75 significantly different RNA transcript ratios, respectively. Differentially expressed RNA transcripts during growth in the absence of added sodium ions included genes coding for permeases, a sodium/proline transporter and electron transport proteins. In contrast, culturing without added potassium ions resulted in higher RNA transcripts for similar genes as a lack of sodium ions plus genes related to spermidine that has a general role in response to stress and a decarboxylase that potentially consumes protons. The greatest RNA transcript response occurred when S. acidocaldarius cells were grown in the absence of potassium and/or sodium at a sub-optimal pH. These adaptations included those listed above plus osmoregulated glucans and mechanosensitive channels that have previously been shown to respond to osmotic stress. In addition, data analyses revealed two co-expressed IcIR family transcriptional regulator genes with a previously unknown role in the S. acidocaldarius pH stress response. Our study provides additional evidence towards the importance of potassium in acidophile growth at acidic pH.

  • 136. Bugiardini, Enrico
    et al.
    Mitchell, Alice L.
    Dalla Rosa, Ilaria
    Horning-Do, Hue-Tran
    Pitmann, Alan M.
    Poole, Olivia
    Holton, Janice L.
    Shah, Sachit
    Woodward, Cathy
    Hargreaves, Iain
    Quinlivan, Rosaline
    Amunts, Alexey
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab). Karolinska Institutet, Sweden.
    Wiesner, Rudolf J.
    Houlden, Henry
    Holt, Ian J.
    Hanna, Michael G.
    Pitceathly, Robert D. S.
    Spinazzola, Antonella
    MRPS25 mutations impair mitochondrial translation and cause encephalomyopathy2019In: Human Molecular Genetics, ISSN 0964-6906, E-ISSN 1460-2083, Vol. 28, no 16, p. 2711-2719Article in journal (Refereed)
    Abstract [en]

    Mitochondrial disorders are clinically and genetically heterogeneous and are associated with a variety of disease mechanisms. Defects of mitochondrial protein synthesis account for the largest subgroup of disorders manifesting with impaired respiratory chain capacity; yet, only a few have been linked to dysfunction in the protein components of the mitochondrial ribosomes. Here, we report a subject presenting with dyskinetic cerebral palsy and partial agenesis of the corpus callosum, while histochemical and biochemical analyses of skeletal muscle revealed signs of mitochondrial myopathy. Using exome sequencing, we identified a homozygous variant c.215C>T in MRPS25, which encodes for a structural component of the 28S small subunit of the mitochondrial ribosome (mS25). The variant segregated with the disease and substitutes a highly conserved proline residue with leucine (p.P72L) that, based on the high-resolution structure of the 28S ribosome, is predicted to compromise inter-protein contacts and destabilize the small subunit. Concordant with the in silico analysis, patient's fibroblasts showed decreased levels of MRPS25 and other components of the 28S subunit. Moreover, assembled 28S subunits were scarce in the fibroblasts with mutant mS25 leading to impaired mitochondrial translation and decreased levels of multiple respiratory chain subunits. Crucially, these abnormalities were rescued by transgenic expression of wild-type MRPS25 in the mutant fibroblasts. Collectively, our data demonstrate the pathogenicity of the p.P72L variant and identify MRPS25 mutations as a new cause of mitochondrial translation defect.

  • 137.
    Bui, Thuy T.
    et al.
    Stockholm University, Faculty of Science, Department of Environmental Science.
    Aasa, Jenny
    Abass, Khaled
    Ågerstrand, Marlene
    Stockholm University, Faculty of Science, Department of Environmental Science.
    Beronius, Anna
    Castro, Mafalda
    Escrivá, Laura
    Galizia, Audrey
    Gliga, Anda
    Karlsson, Oskar
    Stockholm University, Science for Life Laboratory (SciLifeLab). Stockholm University, Faculty of Science, Department of Environmental Science.
    Whaley, Paul
    Yost, Erin
    Rudén, Christina
    Stockholm University, Faculty of Science, Department of Environmental Science.
    Applying a modified systematic review and integrated assessment framework (SYRINA) - a case study on triphenyl phosphate2023In: Environmental Science: Processes & Impacts, ISSN 2050-7887, E-ISSN 2050-7895, Vol. 26, no 2, p. 380-399Article in journal (Refereed)
    Abstract [en]

    This work presents a case study in applying a systematic review framework (SYRINA) to the identification of chemicals as endocrine disruptors. The suitability and performance of the framework is tested with regard to the widely accepted World Health Organization definition of an endocrine disruptor (ED). The endocrine disrupting potential of triphenyl phosphate (TPP), a well-studied flame retardant reported to exhibit various endocrine related effects was assessed. We followed the 7 steps of the SYRINA framework, articulating the research objective via Populations, Exposures, Comparators, Outcomes (PECO) statements, performed literature search and screening, conducted study evaluation, performed data extraction and summarized and integrated the evidence. Overall, 66 studies, consisting of in vivoin vitro and epidemiological data, were included. We concluded that triphenyl phosphate could be identified as an ED based on metabolic disruption and reproductive function. We found that the tools used in this case study and the optimizations performed on the framework were suitable to assess properties of EDs. A number of challenges and areas for methodological development in systematic appraisal of evidence relating to endocrine disrupting potential were identified; significant time and effort were needed for the analysis of in vitro mechanistic data in this case study, thus increasing the workload and time needed to perform the systematic review process. Further research and development of this framework with regards to grey literature (non-peer-reviewed literature) search, harmonization of study evaluation methods, more consistent evidence integration approaches and a pre-defined method to assess links between adverse effect and endocrine activity are recommended. It would also be advantageous to conduct more case studies for a chemical with less data than TPP.

  • 138. Burke, David F.
    et al.
    Bryant, Patrick
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Barrio-Hernandez, Inigo
    Memon, Danish
    Pozzati, Gabriele
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Shenoy, Aditi
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Zhu, Wensi
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Dunham, Alistair S.
    Albanese, Pascal
    Keller, Andrew
    Scheltema, Richard A.
    Bruce, James E.
    Leitner, Alexander
    Kundrotas, Petras
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab). The University of Kansas, Lawrence, USA.
    Beltrao, Pedro
    Elofsson, Arne
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Towards a structurally resolved human protein interaction network2023In: Nature Structural & Molecular Biology, ISSN 1545-9993, E-ISSN 1545-9985, Vol. 30, no 2, p. 216-225Article in journal (Refereed)
    Abstract [en]

    Cellular functions are governed by molecular machines that assemble through protein-protein interactions. Their atomic details are critical to studying their molecular mechanisms. However, fewer than 5% of hundreds of thousands of human protein interactions have been structurally characterized. Here we test the potential and limitations of recent progress in deep-learning methods using AlphaFold2 to predict structures for 65,484 human protein interactions. We show that experiments can orthogonally confirm higher-confidence models. We identify 3,137 high-confidence models, of which 1,371 have no homology to a known structure. We identify interface residues harboring disease mutations, suggesting potential mechanisms for pathogenic variants. Groups of interface phosphorylation sites show patterns of co-regulation across conditions, suggestive of coordinated tuning of multiple protein interactions as signaling responses. Finally, we provide examples of how the predicted binary complexes can be used to build larger assemblies helping to expand our understanding of human cell biology.

  • 139. Buslaev, Pavel
    et al.
    Aho, Noora
    Jansen, Anton
    Bauer, Paul
    Hess, Berk
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Groenhof, Gerrit
    Scalable Constant pH Molecular Dynamics in GROMACS2022In: Journal of Chemical Theory and Computation, ISSN 1549-9618, E-ISSN 1549-9626, Vol. 18, no 10, p. 6148-6160Article in journal (Refereed)
    Abstract [en]

    Molecular dynamics (MD) computer simulations are used routinely to compute atomistic trajectories of complex systems. Systems are simulated in various ensembles, depending on the experimental conditions one aims to mimic. While constant energy, temperature, volume, and pressure are rather straightforward to model, pH, which is an equally important parameter in experiments, is more difficult to account for in simulations. Although a constant pH algorithm based on the λ-dynamics approach by Brooks and co-workers [Kong, X.; Brooks III, C. L. J. Chem. Phys.1996105, 2414–2423] was implemented in a fork of the GROMACS molecular dynamics program, uptake has been rather limited, presumably due to the poor scaling of that code with respect to the number of titratable sites. To overcome this limitation, we implemented an alternative scheme for interpolating the Hamiltonians of the protonation states that makes the constant pH molecular dynamics simulations almost as fast as a normal MD simulation with GROMACS. In addition, we implemented a simpler scheme, called multisite representation, for modeling side chains with multiple titratable sites, such as imidazole rings. This scheme, which is based on constraining the sum of the λ-coordinates, not only reduces the complexity associated with parametrizing the intramolecular interactions between the sites but also is easily extendable to other molecules with multiple titratable sites. With the combination of a more efficient interpolation scheme and multisite representation of titratable groups, we anticipate a rapid uptake of constant pH molecular dynamics simulations within the GROMACS user community.

  • 140. Bustamante, M.
    et al.
    Hernandez-Ferrer, C.
    Tewari, A.
    Sarria, Y.
    Harrison, G. I.
    Puigdecanet, E.
    Nonell, L.
    Kang, Wenjing
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Friedländer, Marc R.
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Estivill, X.
    González, J. R.
    Nieuwenhuijsen, M.
    Young, A. R.
    Dose and time effects of solar-simulated ultraviolet radiation on the in vivo human skin transcriptome2020In: British Journal of Dermatology, ISSN 0007-0963, E-ISSN 1365-2133, Vol. 182, no 6, p. 1458-1468Article in journal (Refereed)
    Abstract [en]

    Background Terrestrial ultraviolet (UV) radiation causes erythema, oxidative stress, DNA mutations and skin cancer. Skin can adapt to these adverse effects by DNA repair, apoptosis, keratinization and tanning.

    Objectives To investigate the transcriptional response to fluorescent solar-simulated radiation (FSSR) in sun-sensitive human skin in vivo.

    Methods Seven healthy male volunteers were exposed to 0, 3 and 6 standard erythemal doses (SED). Skin biopsies were taken at 6 h and 24 h after exposure. Gene and microRNA expression were quantified with next generation sequencing. A set of candidate genes was validated by quantitative polymerase chain reaction (qPCR); and wavelength dependence was examined in other volunteers through microarrays.

    Results The number of differentially expressed genes increased with FSSR dose and decreased between 6 and 24 h. Six hours after 6 SED, 4071 genes were differentially expressed, but only 16 genes were affected at 24 h after 3 SED. Genes for apoptosis and keratinization were prominent at 6 h, whereas inflammation and immunoregulation genes were predominant at 24 h. Validation by qPCR confirmed the altered expression of nine genes detected under all conditions; genes related to DNA repair and apoptosis; immunity and inflammation; pigmentation; and vitamin D synthesis. In general, candidate genes also responded to UVA1 (340-400 nm) and/or UVB (300 nm), but with variations in wavelength dependence and peak expression time. Only four microRNAs were differentially expressed by FSSR.

    Conclusions The UV radiation doses of this acute study are readily achieved daily during holidays in the sun, suggesting that the skin transcriptional profile of 'typical' holiday makers is markedly deregulated.

  • 141. Bustamante, Mariona
    et al.
    Hernandez-Ferrer, Caries
    Sarria, Yaris
    Harrison, Graham I.
    Nonell, Lara
    Kang, Wenjing
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Friedlander, Marc R.
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Estivill, Xavier
    Gonzalez, Juan R.
    Nieuwenhuijsen, Mark
    Young, Antony R.
    The acute effects of ultraviolet radiation on the blood transcriptome are independent of plasma 25OHD(3)2017In: Environmental Research, ISSN 0013-9351, E-ISSN 1096-0953, Vol. 159, p. 239-248Article in journal (Refereed)
    Abstract [en]

    The molecular basis of many health outcomes attributed to solar ultraviolet radiation (UVR) is unknown. We tested the hypothesis that they may originate from transcriptional changes in blood cells. This was determined by assessing the effect of fluorescent solar simulated radiation (FSSR) on the transcriptional profile of peripheral blood pre- and 6 h, 24 h and 48 h post-exposure in nine healthy volunteers. Expression of 20 genes was down regulated and one was up-regulated at 6 h after FSSR. All recovered to baseline expression at 24 h or 48 h. These genes have been associated with immune regulation, cancer and blood pressure; health effects attributed to vitamin D via solar UVR exposure. Plasma 25-hydroxyvitamin D-3 [250HD(3)] levels increased over time after FSSR and were maximal at 48 h. The increase was more pronounced in participants with low basal 250HD(3) levels. Mediation analyses suggested that changes in gene expression due to FSSR were independent of 250HD(3) and blood cell subpopulations.

  • 142.
    Buzzao, Davide
    et al.
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Castresana-Aguirre, Miguel
    Guala, Dimitri
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Sonnhammer, Erik L. L.
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    TOPAS, a network-based approach to detect disease modules in a top-down fashion 2022In: NAR Genomics and Bioinformatics, E-ISSN 2631-9268, Vol. 4, no 4, article id lqac093Article in journal (Refereed)
    Abstract [en]

    A vast scenario of potential disease mechanisms and remedies is yet to be discovered. The field of Network Medicine has grown thanks to the massive amount of high-throughput data and the emerging evidence that disease-related proteins form ‘disease modules’. Relying on prior disease knowledge, network-based disease module detection algorithms aim at connecting the list of known disease associated genes by exploiting interaction networks. Most existing methods extend disease modules by iteratively adding connector genes in a bottom-up fashion, while top-down approaches remain largely unexplored. We have created TOPAS, an iterative approach that aims at connecting the largest number of seed nodes in a top-down fashion through connectors that guarantee the highest flow of a Random Walk with Restart in a network of functional associations. We used a corpus of 382 manually selected functional gene sets to benchmark our algorithm against SCA, DIAMOnD, MaxLink and ROBUST across four interactomes. We demonstrate that TOPAS outperforms competing methods in terms of Seed Recovery Rate, Seed to Connector Ratio and consistency during module detection. We also show that TOPAS achieves competitive performance in terms of biological relevance of detected modules and scalability. 

  • 143.
    Calado Botelho, Salomé
    et al.
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
    Tatsuta, Takashi
    von Heijne, Gunnar
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Kim, Hyun
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Seoul National University, South Korea.
    Dislocation by the m-AAA Protease Increases the Threshold Hydrophobicity for Retention of Transmembrane Helices in the Inner Membrane of Yeast Mitochondria2013In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 288, no 7, p. 4792-4798Article in journal (Refereed)
    Abstract [en]

    Sorting of mitochondrial inner membrane proteins is a complex process in which translocons and proteases function in a concerted way. Many inner membrane proteins insert into the membrane via the TIM23 translocon, and some are then further acted upon by the mitochondrial m-AAA protease, a molecular motor capable of dislocating proteins from the inner membrane. This raises the possibility that the threshold hydrophobicity for the retention of transmembrane segments in the inner membrane is different depending on whether they belong to membrane proteins that are m-AAA protease substrates or not. Here, using model transmembrane segments engineered into m-AAA protease-dependent proteins, we show that the threshold hydrophobicity for membrane retention measured in yeast cells in the absence of a functional m-AAA protease is markedly lower than that measured in its presence. Whether a given hydrophobic segment in a mitochondrial inner membrane protein will ultimately form a transmembrane helix may therefore depend on whether or not it will be exposed to the pulling force exerted by the m-AAA protease during biogenesis.

  • 144. Campos, Alexandre
    et al.
    Danielsson, Gabriela
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Farinha, Ana Paula
    Kuruvilla, Jacob
    Warholm, Per
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Cristobal, Susana
    Shotgun proteomics to unravel marine mussel (Mytilus edulis) response to long-term exposure to low salinity and propranolol in a Baltic Sea microcosm2016In: Journal of Proteomics, ISSN 1874-3919, E-ISSN 1876-7737, Vol. 137, p. 97-106Article in journal (Refereed)
    Abstract [en]

    Pharmaceuticals, among them thep-adrenoceptor blocker propranolol, are an important group of environmental contaminants reported in European waters. Laboratory exposure to pharmaceuticals on marine species has been performed without considering the input of the ecosystem flow. To unravel the ecosystem response to long-term exposure to propranolol we have performed long-term exposure to propranolol and low salinity in microcosms. We applied shotgun proteomic analysis to gills of Mytilus edulis from those Baltic Sea microcosms and identified 2071 proteins with a proteogenomic strategy. The proteome profiling patterns from the 587 highly reproductive proteins among groups define salinity as a key factor in the mussel's response to propranolol. Exposure at low salinity drives molecular mechanisms of adaptation based on a decrease in the abundance of several cytoskeletal proteins, signalling and intracellular membrane trafficking pathway combined with a response towards the maintenance of transcription and translation. The exposure to propranolol combined with low salinity modulates the expression of structural proteins including cilia functions and decreases the expression of membrane protein transporters. This study reinforces the environment concerns of the impact of low salinity in combination with anthropogenic pollutants and anticipates critical physiological conditions for the survival of the blue mussel in the northern areas. Biological significance: Applying shotgun proteomic analysis to M. edulis gills samples from a long-term microcosm exposure to propranolol and following a proteogenomic identification strategy, we have identified 2071 proteins. The proteomic analysis unrevealed which molecular mechanisms drive the adaptation to low salinity stress and how salinity modulates the effects of exposure to propranolol. These results reinforce the idea of the impact of low salinity in combination with anthropogenic pollutants and anticipate critical physiological condition.

  • 145. Cao, Lijia
    et al.
    Garcia, Sarahi L.
    Stockholm University, Faculty of Science, Department of Ecology, Environment and Plant Sciences. Stockholm University, Science for Life Laboratory (SciLifeLab). University of Oldenburg, Germany.
    Wurzbacher, Christian
    Establishment of microbial model communities capable of removing trace organic chemicals for biotransformation mechanisms research2023In: Microbial Cell Factories, ISSN 1475-2859, E-ISSN 1475-2859, Vol. 22, no 1, article id 245Article in journal (Refereed)
    Abstract [en]

    Background Removal of trace organic chemicals (TOrCs) in aquatic environments has been intensively studied. Some members of natural microbial communities play a vital role in transforming chemical contaminants, however, complex microbial interactions impede us from gaining adequate understanding of TOrC biotransformation mechanisms. To simplify, in this study, we propose a strategy of establishing reduced-richness model communities capable of removing diverse TOrCs via pre-adaptation and dilution-to-extinction.

    Results Microbial communities were adapted from tap water, soil, sand, sediment deep and sediment surface to changing concentrations of 27 TOrCs mixture. After adaptation, the communities were further diluted to reduce diversity into 96 deep well plates for high-throughput cultivation. After characterizing microbial structure and TOrC removal performance, thirty taxonomically non-redundant model communities with different removal abilities were obtained. The pre-adaptation process was found to reduce the microbial richness but to increase the evenness and phylogenetic diversity of resulting model communities. Moreover, phylogenetic diversity showed a positive effect on the number of TOrCs that can be transformed simultaneously. Pre-adaptation also improved the overall TOrC removal rates, which was found to be positively correlated with the growth rates of model communities.

    Conclusions This is the first study that investigated a wide range of TOrC biotransformation based on different model communities derived from varying natural microbial systems. This study provides a standardized workflow of establishing model communities for different metabolic purposes with changeable inoculum and substrates. The obtained model communities can be further used to find the driving agents of TOrC biotransformation at the enzyme/gene level.

  • 146. Carinelli, S.
    et al.
    Kühnemund, M.
    Nilsson, Mats
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Pividori, M. I.
    Yoctomole electrochemical genosensing of Ebola virus cDNA by rolling circle and circle to circle amplification2017In: Biosensors & bioelectronics, ISSN 0956-5663, E-ISSN 1873-4235, Vol. 93, p. 65-71Article in journal (Refereed)
    Abstract [en]

    This work addresses the design of an Ebola diagnostic test involving a simple, rapid, specific and highly sensitive procedure based on isothermal amplification on magnetic particles with electrochemical readout. Ebola padlock probes were designed to detect a specific L-gene sequence present in the five most common Ebola species. Ebola cDNA was amplified by rolling circle amplification (RCA) on magnetic particles. Further re-amplification was performed by circle-to-circle amplification (C2CA) and the products were detected in a double-tagging approach using a biotinylated capture probe for immobilization on magnetic particles and a readout probe for electrochemical detection by square-wave voltammetry on commercial screen-printed electrodes. The electrochemical genosensor was able to detect as low as 200 ymol, corresponding to 120 cDNA molecules of L-gene Ebola virus with a limit of detection of 33 cDNA molecules. The isothermal double-amplification procedure by C2CA combined with the electrochemical readout and the magnetic actuation enables the high sensitivity, resulting in a rapid, inexpensive, robust and user-friendly sensing strategy that offers a promising approach for the primary care in low resource settings, especially in less developed countries.

  • 147. Carow, Berit
    et al.
    Hauling, Thomas
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Qian, Xiaoyan
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Kramnik, Igor
    Nilsson, Mats
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Rottenberg, Martin E.
    Spatial and temporal localization of immune transcripts defines hallmarks and diversity in the tuberculosis granuloma2019In: Nature Communications, E-ISSN 2041-1723, Vol. 10, article id 1823Article in journal (Refereed)
    Abstract [en]

    Granulomas are the pathological hallmark of tuberculosis (TB) and the niche where bacilli can grow and disseminate or the immunological microenvironment in which host cells interact to prevent bacterial dissemination. Here we show 34 immune transcripts align to the morphology of lung sections from Mycobacterium tuberculosis-infected mice at cellular resolution. Colocalizing transcript networks at <10 mu m in C57BL/6 mouse granulomas increase complexity with time after infection. B-cell clusters develop late after infection. Transcripts from activated macrophages are enriched at subcellular distances from M. tuberculosis. Encapsulated C3HeB/FeJ granulomas show necrotic centers with transcripts associated with immunosuppression (Foxp3, Il10), whereas those in the granuloma rims associate with activated T cells and macrophages. We see highly diverse networks with common interactors in similar lesions. Different immune landscapes of M. tuberculosis granulomas depending on the time after infection, the histopathological features of the lesion, and the proximity to bacteria are here defined.

  • 148. Carow, Berit
    et al.
    Muliadi, Victoria
    Skålén, Kristina
    Yokota, Chika
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Kathamuthu, Gokul Raj
    Setiabudiawan, Todia Pediatama
    Lange, Christoph
    Scheu, Katrin
    Gaede, Karoline I.
    Goldmann, Torsten
    Pandita, Ankur
    Masood, Kiran Iqbal
    Pervez, Shahid
    Grunewald, Johan
    Hasan, Zahra
    Levin, Max
    Rottenberg, Martin E.
    Immune mapping of human tuberculosis and sarcoidosis lung granulomas2024In: Frontiers in Immunology, E-ISSN 1664-3224, Vol. 14, article id 1332733Article in journal (Refereed)
    Abstract [en]

    Tuberculosis (TB) and sarcoidosis are both granulomatous diseases. Here, we compared the immunological microenvironments of granulomas from TB and sarcoidosis patients using in situ sequencing (ISS) transcriptomic analysis and multiplexed immunolabeling of tissue sections. TB lesions consisted of large necrotic and cellular granulomas, whereas “multifocal” granulomas with macrophages or epitheloid cell core and a T-cell rim were observed in sarcoidosis samples. The necrotic core in TB lesions was surrounded by macrophages and encircled by a dense T-cell layer. Within the T-cell layer, compact B-cell aggregates were observed in most TB samples. These B-cell clusters were vascularized and could contain defined B-/T-cell and macrophage-rich areas. The ISS of 40–60 immune transcripts revealed the enriched expression of transcripts involved in homing or migration to lymph nodes, which formed networks at single-cell distances in lymphoid areas of the TB lesions. Instead, myeloid-annotated regions were enriched in CD68CD14ITGAMITGAX, and CD4 mRNA. CXCL8 and IL1B mRNA were observed in granulocytic areas in which M. tuberculosis was also detected. In line with ISS data indicating tertiary lymphoid structures, immune labeling of TB sections expressed markers of high endothelial venules, follicular dendritic cells, follicular helper T cells, and lymph-node homing receptors on T cells. Neither ISS nor immunolabeling showed evidence of tertiary lymphoid aggregates in sarcoidosis samples. Together, our finding suggests that despite their heterogeneity, the formation of tertiary immune structures is a common feature in granulomas from TB patients.

  • 149. Carreras-Puigvert, Jordi
    et al.
    Zitnik, Marinka
    Jemth, Ann-Sofie
    Carter, Megan
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
    Unterlass, Judith E.
    Hallström, Björn
    Loseva, Olga
    Karem, Zhir
    Calderón-Montaño, José Manuel
    Lindskog, Cecilia
    Edqvist, Per-Henrik
    Matuszewski, Damian J.
    Blal, Hammou Ait
    Berntsson, Ronnie P. A.
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
    Häggblad, Maria
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Martens, Ulf
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Studham, Matthew
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Lundgren, Bo
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Wählby, Carolina
    Sonnhammer, Erik L. L.
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Lundberg, Emma
    Stenmark, Pål
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
    Zupan, Blaz
    Helleday, Thomas
    A comprehensive structural, biochemical and biological profiling of the human NUDIX hydrolase family2017In: Nature Communications, E-ISSN 2041-1723, Vol. 8, article id 1541Article in journal (Refereed)
    Abstract [en]

    The NUDIX enzymes are involved in cellular metabolism and homeostasis, as well as mRNA processing. Although highly conserved throughout all organisms, their biological roles and biochemical redundancies remain largely unclear. To address this, we globally resolve their individual properties and inter-relationships. We purify 18 of the human NUDIX proteins and screen 52 substrates, providing a substrate redundancy map. Using crystal structures, we generate sequence alignment analyses revealing four major structural classes. To a certain extent, their substrate preference redundancies correlate with structural classes, thus linking structure and activity relationships. To elucidate interdependence among the NUDIX hydrolases, we pairwise deplete them generating an epistatic interaction map, evaluate cell cycle perturbations upon knockdown in normal and cancer cells, and analyse their protein and mRNA expression in normal and cancer tissues. Using a novel FUSION algorithm, we integrate all data creating a comprehensive NUDIX enzyme profile map, which will prove fundamental to understanding their biological functionality.

  • 150. Carstens, Adam
    et al.
    Roos, Annika
    Andreasson, Anna
    Stockholm University, Faculty of Social Sciences, Stress Research Institute. Stockholm University, Science for Life Laboratory (SciLifeLab). Karolinska Institute, Sweden.
    Magnuson, Anders
    Agreus, Lars
    Halfvarson, Jonas
    Engstrand, Lars
    Differential clustering of fecal and mucosa-associated microbiota in 'healthy' individuals2018In: Journal of Digestive Diseases, ISSN 1751-2972, E-ISSN 1751-2980, Vol. 19, no 12, p. 745-752Article in journal (Refereed)
    Abstract [en]

    Objective Fecal samples are often used to characterize gut microbiota. However, whether or not fecal microbiota differs from mucosa-associated microbiota remains largely unknown. This may be specifically relevant in conditions that are characterized by complex mucosal microbe-host interactions, such as Crohn's disease. We aimed to determine the degree of agreement between fecal and mucosal microbiota profiles in 'healthy' individuals, using two commonly used collection procedures. Methods The gut microbiota composition of fecal samples (sent at ambient temperature before storage at -70 degrees C) and of colonic biopsies (obtained at endoscopy and immediately stored at -70 degrees C) was determined by sequencing the 16S rRNA gene. Altogether 31 randomly selected 'healthy' individuals from the population-based colonoscopy (Popcol) study were included. Results The fecal samples were characterized by a reduced degree of richness (P < 0.0001) and diversity (P = 0.016), and also differences in several phyla, including a lower relative abundance of Proteobacteria (P < 0.0001) and Verrucomicrobia (P = 0.008) than in biopsies. Only three of 30 individuals had a similar fecal and mucosal microbiota profile, based on weighted UniFrac analysis. A difference in Crohn's disease dysbiosis-associated bacteria was observed, including a lower relative abundance of Faecalibacterium (P = 0.004) and a higher relative abundance of Ruminococcus (P = 0.001) in feces than in biopsies. Conclusions The observed differences between fecal samples, transported at ambient temperature, and the colonic mucosa-associated microbiota have implications for the interpretation of the previous literature, and may be specifically relevant to studies on Crohn's disease.

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