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  • 1. Acevedo, Nathalie
    et al.
    Bornacelly, Adriana
    Mercado, Dilia
    Unneberg, Per
    Stockholm University, Science for Life Laboratory (SciLifeLab). Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
    Mittermann, Irene
    Valenta, Rudolf
    Kennedy, Malcolm
    Scheynius, Annika
    Caraballo, Luis
    Genetic Variants in CHIA and CHI3L1 Are Associated with the IgE Response to the Ascaris Resistance Marker ABA-1 and the Birch Pollen Allergen Bet v 12016In: plos one, ISSN 1932-6203, Vol. 11, no 12, e0167453Article in journal (Refereed)
    Abstract [en]

    Helminth infections and allergic diseases are associated with IgE hyperresponsiveness but the genetics of this phenotype remain to be defined. Susceptibility to Ascaris lumbricoides infection and antibody levels to this helminth are associated with polymorphisms in locus 13q33-34. We aimed to explore this and other genomic regions to identify genetic variants associated with the IgE responsiveness in humans. Forty-eight subjects from Cartagena, Colombia, with extreme values of specific IgE to Ascaris and ABA-1, a resistance marker of this nematode, were selected for targeted resequencing. Burden analyses were done comparing extreme groups for IgE values. One-hundred one SNPs were genotyped in 1258 individuals of two well-characterized populations from Colombia and Sweden. Two low-frequency coding variants in the gene encoding the Acidic Mammalian Chitinase (CHIA rs79500525, rs139812869, tagged by rs10494133) were found enriched in high IgE responders to ABA-1 and confirmed by genetic association analyses. The SNP rs4950928 in the Chitinase 3 Like 1 gene (CHI3L1) was associated with high IgE to ABA-1 in Colombians and with high IgE to Bet v 1 in the Swedish population. CHIA rs10494133 and ABDH13 rs3783118 were associated with IgE responses to Ascaris. SNPs in the Tumor Necrosis Factor Superfamily Member 13b gene (TNFSF13B) encoding the cytokine B cell activating Factor were associated with high levels of total IgE in both populations. This is the first report on the association between low-frequency and common variants in the chitinases- related genes CHIA and CHI3L1 with the intensity of specific IgE to ABA-1 in a population naturally exposed to Ascaris and with Bet v 1 in a Swedish population. Our results add new information about the genetic influences of human IgE responsiveness; since the genes encode for enzymes involved in the immune response to parasitic infections, they could be helpful for understanding helminth immunity and allergic responses. We also confirmed that TNFSF13B has an important and conserved role in the regulation of total IgE levels, which supports potential evolutionary links between helminth immunity and allergic response.

  • 2.
    Ahmed, Engy
    et al.
    Stockholm University, Faculty of Science, Department of Geological Sciences. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Abdulla, Hesham M.
    Mohamed, Amy H.
    El-Bassuony, Ahmed D.
    Remediation and recycling of chromium from tannery wastewater using combined chemical-biological treatment system2016In: Process Safety and Environmental Protection, ISSN 0957-5820, E-ISSN 1744-3598, Vol. 104, 1-10 p.Article in journal (Refereed)
    Abstract [en]

    Tannery wastewater containing chromium (Cr) is one of the most serious problems in leather industry. In order to develop an effective and eco-friendly treatment technology, a combined chemical-biological treatment system was performed for Cr remediation and recycling. The aim of the present study is to design a laboratory scale system using chemical precipitation of Cr(III) combined with biological removal of Cr(VI) from tannery wastewater, and to investigate the possibility of recycling the recovered Cr(III) in the tanning industry. Chemical precipitation of Cr(III) was carried out using lime and cement dust. The actinomycete strain Kitasatosporia sp. was used in microcosm studies for Cr(VI) bioremoval. Moreover, parameters such as type of porous medium, inoculum size, flow rate and culture conditions were investigated. The precipitated Cr(III) that was recovered from the chemical precipitation stage was recycled in the leather tanning industry. Our findings indicate that the maximum Cr(III) precipitation (98%) was achieved using 2 g/100 mL of lime and 2 h of settling rate. On the other hand, microcosm columns using sand that was inoculated with induced culture (OD600 = 2.43) and flow rate (2 mL/min) gave the maximum recovery (99%) of Cr(VI). The experimental Cr(III) was successfully recycled in the tanning process and the experimental leathers showed comparable properties as same as the leathers tanned with commercial Cr(III). Thus, we concluded that using combined chemical-biological treatment system for Cr remediation from tanning wastewater together with recycling process for the recovered Cr(III) is a promising strategy for economic and environmental friendly tanning industry.

  • 3. Ahrentorp, Fredrik
    et al.
    Blomgren, Jakob
    Jonasson, Christian
    Sarwe, Anna
    Sepehri, Sobhan
    Eriksson, Emil
    Kalaboukhov, Alexei
    Jesorka, Aldo
    Winkler, Dag
    Schneiderman, Justin F.
    Nilsson, Mats
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Albert, Jan
    Gómez de la Torre, Teresa Zardán
    Strømme, Maria
    Johansson, Christer
    Sensitive magnetic biodetection using magnetic multi-core nanoparticles and RCA coils2017In: Journal of Magnetism and Magnetic Materials, ISSN 0304-8853, E-ISSN 1873-4766, Vol. 427, 14-18 p.Article in journal (Refereed)
    Abstract [en]

    We use functionalized iron oxide magnetic multi-core particles of 100 nm in size (hydrodynamic particle diameter) and AC susceptometry (ACS) methods to measure the binding reactions between the magnetic nanoparticles (MNPs) and bio-analyte products produced from DNA segments using the rolling circle amplification (RCA) method. We use sensitive induction detection techniques in order to measure the ACS response. The DNA is amplified via RCA to generate RCA coils with a specific size that is dependent on the amplification time. After about 75 min of amplification we obtain an average RCA coil diameter of about 1 mu m. We determine a theoretical limit of detection (LOD) in the range of 11 attomole (corresponding to an analyte concentration of 55 fM for a sample volume of 200 mu L) from the ACS dynamic response after the MNPs have bound to the RCA coils and the measured ACS readout noise. We also discuss further possible improvements of the LOD.

  • 4. Alcamán, M. Estrella
    et al.
    Alcorta, Jaime
    Bergman, Birgitta
    Stockholm University, Faculty of Science, Department of Ecology, Environment and Plant Sciences. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Vásquez, Mónica
    Polz, Martin
    Díez, Beatriz
    Physiological and gene expression responses to nitrogen regimes and temperatures in Mastigocladus sp strain CHP1, a predominant thermotolerant cyanobacterium of hot springs2017In: Systematic and Applied Microbiology, ISSN 0723-2020, E-ISSN 1618-0984, Vol. 40, no 2, 102-113 p.Article in journal (Refereed)
    Abstract [en]

    Cyanobacteria are widely distributed primary producers with significant implications for the global biogeochemical cycles of carbon and nitrogen. Diazotrophic cyanobacteria of subsection V (Order Stigonematales) are particularly ubiquitous in photoautotrophic microbial mats of hot springs. The Stigonematal cyanobacterium strain CHPI isolated from the Porcelana hot spring (Chile) was one of the major contributors of the new nitrogen through nitrogen fixation. Further morphological and genetic characterization verified that the strain CHP1 belongs to Stigonematales, and it formed a separate Glade together with other thermophiles of the genera Fischerella and Mastigocladus. Strain CHP1 fixed maximum N-2 in the light, independent of the temperature range. At 50 degrees C niJH gene transcripts showed high expression during the light period, whereas the nifH gene expression at 45 degrees C was arrhythmic. The strain displayed a high affinity for nitrate and a low tolerance for high ammonium concentrations, whereas the narB and glnA genes showed higher expression in light and at the beginning of the dark phase. It is proposed that Mastigocladus sp. strain CHPI would represent a good model for the study of subsection V thermophilic cyanobacteria, and for understanding the adaptations of these photoautotrophic organisms inhabiting microbial mats in hot springs globally.

  • 5. Alexeyenko, Andrey
    et al.
    Nystedt, Björn
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Vezzi, Francesco
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Sherwood, Ellen
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Ye, Rosa
    Knudsen, Bjarne
    Simonsen, Martin
    Turner, Benjamin
    de Jong, Pieter
    Wu, Cheng-Cang
    Lundeberg, Joakim
    Efficient de novo assembly of large and complex genomes by massively parallel sequencing of Fosmid pools2014In: BMC Genomics, ISSN 1471-2164, Vol. 15, 439- p.Article in journal (Refereed)
    Abstract [en]

    Background: Sampling genomes with Fosmid vectors and sequencing of pooled Fosmid libraries on the Illumina platform for massive parallel sequencing is a novel and promising approach to optimizing the trade-off between sequencing costs and assembly quality. Results: In order to sequence the genome of Norway spruce, which is of great size and complexity, we developed and applied a new technology based on the massive production, sequencing, and assembly of Fosmid pools (FP). The spruce chromosomes were sampled with similar to 40,000 bp Fosmid inserts to obtain around two-fold genome coverage, in parallel with traditional whole genome shotgun sequencing (WGS) of haploid and diploid genomes. Compared to the WGS results, the contiguity and quality of the FP assemblies were high, and they allowed us to fill WGS gaps resulting from repeats, low coverage, and allelic differences. The FP contig sets were further merged with WGS data using a novel software package GAM-NGS. Conclusions: By exploiting FP technology, the first published assembly of a conifer genome was sequenced entirely with massively parallel sequencing. Here we provide a comprehensive report on the different features of the approach and the optimization of the process. We have made public the input data (FASTQ format) for the set of pools used in this study: ftp://congenie.org/congenie/Nystedt_2013/Assembly/ProcessedData/FosmidPools/.(alternatively accessible via http://congenie.org/downloads).The software used for running the assembly process is available at http://research.scilifelab.se/andrej_alexeyenko/downloads/fpools/.

  • 6.
    Alexeyenko, Andrey
    et al.
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Schmitt, Thomas
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Tjärnberg, Andreas
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Guala, Dmitri
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Frings, Oliver
    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).
    Comparative interactomics with Funcoup 2.02012In: Nucleic Acids Research, ISSN 0305-1048, E-ISSN 1362-4962, Vol. 40, no D1, D821-D828 p.Article in journal (Refereed)
    Abstract [en]

    FunCoup (http://FunCoup.sbc.su.se) is a database that maintains and visualizes global gene/protein networks of functional coupling that have been constructed by Bayesian integration of diverse high-throughput data. FunCoup achieves high coverage by orthology-based integration of data sources from different model organisms and from different platforms. We here present release 2.0 in which the data sources have been updated and the methodology has been refined. It contains a new data type Genetic Interaction, and three new species: chicken, dog and zebra fish. As FunCoup extensively transfers functional coupling information between species, the new input datasets have considerably improved both coverage and quality of the networks. The number of high-confidence network links has increased dramatically. For instance, the human network has more than eight times as many links above confidence 0.5 as the previous release. FunCoup provides facilities for analysing the conservation of subnetworks in multiple species. We here explain how to do comparative interactomics on the FunCoup website.

  • 7. Ali, Raja H.
    et al.
    Bark, Mikael
    Miró, Jorge
    Muhammad, Sayyed A.
    Sjöstrand, Joel
    Stockholm University, Faculty of Science, Numerical Analysis and Computer Science (NADA). Stockholm University, Science for Life Laboratory (SciLifeLab). Swedish e-Science Research Centre, Sweden.
    Zubair, Syed M.
    Abbas, Raja M.
    Arvestad, Lars
    Stockholm University, Faculty of Science, Numerical Analysis and Computer Science (NADA). Stockholm University, Science for Life Laboratory (SciLifeLab). Swedish e-Science Research Centre, Sweden.
    VMCMC: a graphical and statistical analysis tool for Markov chain Monte Carlo traces2017In: BMC Bioinformatics, ISSN 1471-2105, E-ISSN 1471-2105, Vol. 18, 97Article in journal (Refereed)
    Abstract [en]

    Background: MCMC-based methods are important for Bayesian inference of phylogeny and related parameters. Although being computationally expensive, MCMC yields estimates of posterior distributions that are useful for estimating parameter values and are easy to use in subsequent analysis. There are, however, sometimes practical difficulties with MCMC, relating to convergence assessment and determining burn-in, especially in large-scale analyses. Currently, multiple software are required to perform, e.g., convergence, mixing and interactive exploration of both continuous and tree parameters.

    Results: We have written a software called VMCMC to simplify post-processing of MCMC traces with, for example, automatic burn-in estimation. VMCMC can also be used both as a GUI-based application, supporting interactive exploration, and as a command-line tool suitable for automated pipelines.

    Conclusions: VMCMC is a free software available under the New BSD License. Executable jar files, tutorial manual and source code can be downloaded from https://bitbucket. org/rhali/visualmcmc/.

  • 8. Ali, Raja H.
    et al.
    Muhammad, Sayyed A.
    Arvestad, Lars
    Stockholm University, Faculty of Science, Numerical Analysis and Computer Science (NADA). Stockholm University, Science for Life Laboratory (SciLifeLab). Swedish e-Science Research Centre, Sweden.
    GenFamClust: an accurate, synteny-aware and reliable homology inference algorithm2016In: BMC Evolutionary Biology, ISSN 1471-2148, E-ISSN 1471-2148, Vol. 16, 120Article in journal (Refereed)
    Abstract [en]

    Background: Homology inference is pivotal to evolutionary biology and is primarily based on significant sequence similarity, which, in general, is a good indicator of homology. Algorithms have also been designed to utilize conservation in gene order as an indication of homologous regions. We have developed GenFamClust, a method based on quantification of both gene order conservation and sequence similarity. Results: In this study, we validate GenFamClust by comparing it to well known homology inference algorithms on a synthetic dataset. We applied several popular clustering algorithms on homologs inferred by GenFamClust and other algorithms on a metazoan dataset and studied the outcomes. Accuracy, similarity, dependence, and other characteristics were investigated for gene families yielded by the clustering algorithms. GenFamClust was also applied to genes from a set of complete fungal genomes and gene families were inferred using clustering. The resulting gene families were compared with a manually curated gold standard of pillars from the Yeast Gene Order Browser. We found that the gene-order component of GenFamClust is simple, yet biologically realistic, and captures local synteny information for homologs. Conclusions: The study shows that GenFamClust is a more accurate, informed, and comprehensive pipeline to infer homologs and gene families than other commonly used homology and gene-family inference methods.

  • 9. Ali, Raja Hashim
    et al.
    Muhammad, Sayyed Auwn
    Khan, Mehmood Alam
    Arvestad, Lars
    Stockholm University, Faculty of Science, Numerical Analysis and Computer Science (NADA). Stockholm University, Science for Life Laboratory (SciLifeLab). Swedish e-Science Research Center, Sweden .
    Quantitative synteny scoring improves homology inference and partitioning of gene families2013In: BMC Bioinformatics, ISSN 1471-2105, Vol. 14, no Suppl,15, S12- p.Article in journal (Refereed)
    Abstract [en]

    Background

    Clustering sequences into families has long been an important step in characterization of genes and proteins. There are many algorithms developed for this purpose, most of which are based on either direct similarity between gene pairs or some sort of network structure, where weights on edges of constructed graphs are based on similarity. However, conserved synteny is an important signal that can help distinguish homology and it has not been utilized to its fullest potential.

    Results

    Here, we present GenFamClust, a pipeline that combines the network properties of sequence similarity and synteny to assess homology relationship and merge known homologs into groups of gene families. GenFamClust identifies homologs in a more informed and accurate manner as compared to similarity based approaches. We tested our method against the Neighborhood Correlation method on two diverse datasets consisting of fully sequenced genomes of eukaryotes and synthetic data.

    Conclusions

    The results obtained from both datasets confirm that synteny helps determine homology and GenFamClust improves on Neighborhood Correlation method. The accuracy as well as the definition of synteny scores is the most valuable contribution of GenFamClust.

  • 10. Allen, Lisa Zeigler
    et al.
    McCrow, John P.
    Ininbergs, Karolina
    Stockholm University, Faculty of Science, Department of Ecology, Environment and Plant Sciences. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Dupont, Christopher L.
    Badger, Jonathan H.
    Hoffman, Jeffery M.
    Ekman, Martin
    Stockholm University, Faculty of Science, Department of Ecology, Environment and Plant Sciences. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Allen, Andrew E.
    Bergman, Birgitta
    Stockholm University, Faculty of Science, Department of Ecology, Environment and Plant Sciences. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Venter, J. Craig
    The Baltic Sea Virome: Diversity and Transcriptional Activity of DNA and RNA Viruses2017In: mSystems, ISSN 2379-5077, Vol. 2, no 1, UNSP e00125-16Article in journal (Refereed)
    Abstract [en]

    Metagenomic and metatranscriptomic data were generated from size-fractionated samples from 11 sites within the Baltic Sea and adjacent marine waters of Kattegat and freshwater Lake Tornetrask in order to investigate the diversity, distribution, and transcriptional activity of virioplankton. Such a transect, spanning a salinity gradient from freshwater to the open sea, facilitated a broad genome-enabled investigation of natural as well as impacted aspects of Baltic Sea viral communities. Taxonomic signatures representative of phages within the widely distributed order Caudovirales were identified with enrichments in lesser-known families such as Podoviridae and Siphoviridae. The distribution of phage reported to infect diverse and ubiquitous heterotrophic bacteria (SAR11 clades) and cyanobacteria (Synechococcus sp.) displayed population-level shifts in diversity. Samples from higher-salinity conditions (>14 practical salinity units [PSU]) had increased abundances of viruses for picoeukaryotes, i.e., Ostreococcus. These data, combined with host diversity estimates, suggest viral modulation of diversity on the whole-community scale, as well as in specific prokaryotic and eukaryotic lineages. RNA libraries revealed single-stranded DNA (ssDNA) and RNA viral populations throughout the Baltic Sea, with ssDNA phage highly represented in Lake Tornetrask. Further, our data suggest relatively high transcriptional activity of fish viruses within diverse families known to have broad host ranges, such as Nodoviridae (RNA), Iridoviridae (DNA), and predicted zoonotic viruses that can cause ecological and economic damage as well as impact human health. IMPORTANCE Inferred virus-host relationships, community structures of ubiquitous ecologically relevant groups, and identification of transcriptionally active populations have been achieved with our Baltic Sea study. Further, these data, highlighting the transcriptional activity of viruses, represent one of the more powerful uses of omics concerning ecosystem health. The use of omics-related data to assess ecosystem health holds great promise for rapid and relatively inexpensive determination of perturbations and risk, explicitly with regard to viral assemblages, as no single marker gene is suitable for widespread taxonomic coverage.

  • 11. Altenhoff, Adrian M.
    et al.
    Boeckmann, Brigitte
    Capella-Gutierrez, Salvador
    Dalquen, Daniel A.
    DeLuca, Todd
    Forslund, Kristoffer
    Huerta-Cepas, Jaime
    Linard, Benjamin
    Pereira, Cecile
    Pryszcz, Leszek P.
    Schreiber, Fabian
    da Silva, Alan Sousa
    Szklarczyk, Damian
    Train, Clement-Marie
    Bork, Peer
    Lecompte, Odile
    von Mering, Christian
    Xenarios, Ioannis
    Sjölander, Kimmen
    Juhl Jensen, Lars
    Martin, Maria J.
    Muffato, Matthieu
    Gabaldon, Toni
    Lewis, Suzanna E.
    Thomas, Paul D.
    Sonnhammer, Erik
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Dessimoz, Christophe
    Standardized benchmarking in the quest for orthologs2016In: Nature Methods, ISSN 1548-7091, E-ISSN 1548-7105, Vol. 13, no 5, 425-+ p.Article in journal (Refereed)
    Abstract [en]

    Achieving high accuracy in orthology inference is essential for many comparative, evolutionary and functional genomic analyses, yet the true evolutionary history of genes is generally unknown and orthologs are used for very different applications across phyla, requiring different precision-recall trade-offs. As a result, it is difficult to assess the performance of orthology inference methods. Here, we present a community effort to establish standards and an automated web-based service to facilitate orthology benchmarking. Using this service, we characterize 15 well-established inference methods and resources on a battery of 20 different benchmarks. Standardized benchmarking provides a way for users to identify the most effective methods for the problem at hand, sets a minimum requirement for new tools and resources, and guides the development of more accurate orthology inference methods.

  • 12. Angleby, Helen
    et al.
    Oskarsson, Mattias
    Pang, Junfeng
    Zhang, Ya-ping
    Leitner, Thomas
    Braham, Caitlyn
    Arvestad, Lars
    Stockholm University, Faculty of Science, Numerical Analysis and Computer Science (NADA). Stockholm University, Science for Life Laboratory (SciLifeLab).
    Lundeberg, Joakim
    Webb, Kristen M.
    Savolainen, Peter
    Forensic Informativity of similar to 3000bp of Coding Sequence of Domestic Dog mtDNA2014In: Journal of Forensic Sciences, ISSN 0022-1198, E-ISSN 1556-4029, Vol. 59, no 4, 898-908 p.Article in journal (Refereed)
    Abstract [en]

    The discriminatory power of the noncoding control region (CR) of domestic dog mitochondrial DNA alone is relatively low. The extent to which the discriminatory power could be increased by analyzing additional highly variable coding regions of the mitochondrial genome (mtGenome) was therefore investigated. Genetic variability across the mtGenome was evaluated by phylogenetic analysis, and the three most variable similar to 1kb coding regions identified. We then sampled 100 Swedish dogs to represent breeds in accordance with their frequency in the Swedish population. A previously published dataset of 59 dog mtGenomes collected in the United States was also analyzed. Inclusion of the three coding regions increased the exclusion capacity considerably for the Swedish sample, from 0.920 for the CR alone to 0.964 for all four regions. The number of mtDNA types among all 159 dogs increased from 41 to 72, the four most frequent CR haplotypes being resolved into 22 different haplotypes.

  • 13. Atzori, Alessio
    et al.
    Liggi, Sonia
    Laaksonen, Aatto
    Stockholm University, Faculty of Science, Department of Materials and Environmental Chemistry (MMK). Stockholm University, Science for Life Laboratory (SciLifeLab). University of Cagliari, Italy.
    Porcu, Massimiliano
    Stockholm University, Faculty of Science, Department of Materials and Environmental Chemistry (MMK). Università di Cagliari, Italy.
    Lyubartsev, Alexander P.
    Stockholm University, Faculty of Science, Department of Materials and Environmental Chemistry (MMK).
    Saba, Giuseppe
    Mocci, Francesca
    Stockholm University, Faculty of Science, Department of Materials and Environmental Chemistry (MMK). Stockholm University, Science for Life Laboratory (SciLifeLab). Università di Cagliari, Italy.
    Base sequence specificity of counterion binding to DNA: what can MD simulations tell us?2016In: Canadian journal of chemistry (Print), ISSN 0008-4042, E-ISSN 1480-3291, Vol. 94, no 12, 1181-1188 p.Article in journal (Refereed)
    Abstract [en]

    Nucleic acids are highly charged biopolymers whose secondary structure is strongly dependent on electrostatic interactions. Solvent molecules and ions are also believed to play an important role in mediating and directing both sequence recognition and interactions with other molecules, such as proteins and a variety of ligands. Therefore, to fully understand the biological functions of DNA, it is necessary to understand the interactions with the surrounding counterions. It is well known that monovalent counterions can bind to the minor groove of DNA with consecutive sequences of four, or more, adenine and thymine (A-tracts) with relatively long residence times. However, much less is known about their binding to the backbone and to the major groove. In this work, we used molecular dynamics simulations to both investigate the interactions between the backbone and major groove of DNA and one of its physiological counterions (Na+) and evaluate the relationship between these interactions and the nucleotide sequence. Three dodecamers, namely CGAAAATTTTCG, CGCTCTAGAGCG, and CGCGAATTCGCG, were simulated using the Toukan-Rahman flexible SPC water model and Smith and Dang parameters for Na+, revealing a significant sequence dependence on the ion binding to both backbone and major groove. In the absence of experimental data on the atomistic details of the studied interactions, the reliability of the results was evaluated performing the simulations with additional sets of potential parameters for ions and solvent, namely the A. qvist or the Joung and Cheatham ion parameters and the TIP3P water model. This allowed us to evaluate the results by verifying which features are preserved independently from the parameters adopted.

  • 14. Ayoglu, Burcu
    et al.
    Birgersson, Elin
    Mezger, Anja
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Nilsson, Mats
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Uhlén, Mathias
    Nilsson, Peter
    Schwenk, Jochen M.
    Multiplexed protein profiling by sequential affinity capture2016In: Proteomics, ISSN 1615-9853, E-ISSN 1615-9861, Vol. 16, no 8, 1251-1256 p.Article in journal (Refereed)
    Abstract [en]

    Antibody microarrays enable parallelized and miniaturized analysis of clinical samples, and have proven to provide novel insights for the analysis of different proteomes. However, there are concerns that the performance of such direct labeling and single antibody assays are prone to off-target binding due to the sample context. To improve selectivity and sensitivity while maintaining the possibility to conduct multiplexed protein profiling, we developed a multiplexed and semi-automated sequential capture assay. This novel bead-based procedure encompasses a first antigen capture, labeling of captured protein targets on magnetic particles, combinatorial target elution and a read-out by a secondary capture bead array. We demonstrate in a proof-of-concept setting that target detection via two sequential affinity interactions reduced off-target contribution, while lowered background and noise levels, improved correlation to clinical values compared to single binder assays. We also compared sensitivity levels with single binder and classical sandwich assays, explored the possibility for DNA-based signal amplification, and demonstrate the applicability of the dual capturebead-based antibody microarray for biomarker analysis. Hence, the described concept enhances the possibilities for antibody array assays to be utilized for protein profiling in body fluids and beyond.

  • 15. Babbitt, Patricia C.
    et al.
    Bagos, Pantelis G.
    Bairoch, Amos
    Bateman, Alex
    Chatonnet, Arnaud
    Chen, Mark Jinan
    Craik, David J.
    Finn, Robert D.
    Gloriam, David
    Haft, Daniel H.
    Henrissat, Bernard
    Holliday, Gemma L.
    Isberg, Vignir
    Kaas, Quentin
    Landsman, David
    Lenfant, Nicolas
    Manning, Gerard
    Nagano, Nozomi
    Srinivasan, Narayanaswamy
    O'Donovan, Claire
    Pruitt, Kim D.
    Sowdhamini, Ramanathan
    Rawlings, Neil D.
    Saier, Milton H.
    Sharman, Joanna L.
    Spedding, Michael
    Tsirigos, Konstantinos D.
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Vastermark, Ake
    Vriend, Gerrit
    Creating a specialist protein resource network: a meeting report for the protein bioinformatics and community resources retreat2015In: Database: The Journal of Biological Databases and Curation, ISSN 1758-0463, E-ISSN 1758-0463, bav063Article in journal (Refereed)
    Abstract [en]

    During 11-12 August 2014, a Protein Bioinformatics and Community Resources Retreat was held at the Wellcome Trust Genome Campus in Hinxton, UK. This meeting brought together the principal investigators of several specialized protein resources (such as CAZy, TCDB and MEROPS) as well as those from protein databases from the large Bioinformatics centres (including UniProt and RefSeq). The retreat was divided into five sessions: (1) key challenges, (2) the databases represented, (3) best practices for maintenance and curation, (4) information flow to and from large data centers and (5) communication and funding. An important outcome of this meeting was the creation of a Specialist Protein Resource Network that we believe will improve coordination of the activities of its member resources. We invite further protein database resources to join the network and continue the dialogue.

  • 16. Baeza-Delgado, Carlos
    et al.
    von Heijne, Gunnar
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Marti-Renom, Marc A.
    Mingarro, Ismael
    Biological insertion of computationally designed short transmembrane segments2016In: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 6, 23397Article in journal (Refereed)
    Abstract [en]

    The great majority of helical membrane proteins are inserted co-translationally into the ER membrane through a continuous ribosome-translocon channel. The efficiency of membrane insertion depends on transmembrane (TM) helix amino acid composition, the helix length and the position of the amino acids within the helix. In this work, we conducted a computational analysis of the composition and location of amino acids in transmembrane helices found in membrane proteins of known structure to obtain an extensive set of designed polypeptide segments with naturally occurring amino acid distributions. Then, using an in vitro translation system in the presence of biological membranes, we experimentally validated our predictions by analyzing its membrane integration capacity. Coupled with known strategies to control membrane protein topology, these findings may pave the way to de novo membrane protein design.

  • 17. Bano-Polo, Manuel
    et al.
    Martinez-Gill, Luis
    Wallner, Björn
    Nieva, Jose L.
    Elofsson, Arne
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Mingarro, Ismael
    Charge Pair Interactions in Transmembrane Helices and Turn Propensity of the Connecting Sequence Promote Helical Hairpin Insertion2013In: Journal of Molecular Biology, ISSN 0022-2836, E-ISSN 1089-8638, Vol. 425, no 4, 830-840 p.Article in journal (Refereed)
    Abstract [en]

    alpha-Helical hairpins, consisting of a pair of closely spaced transmembrane (TM) helices that are connected by a short interfacial turn, are the simplest structural motifs found in multi-spanning membrane proteins. In naturally occurring hairpins, the presence of polar residues is common and predicted to complicate membrane insertion. We postulate that the pre-packing process offsets any energetic cost of allocating polar and charged residues within the hydrophobic environment of biological membranes. Consistent with this idea, we provide here experimental evidence demonstrating that helical hairpin insertion into biological membranes can be driven by electrostatic interactions between closely separated, poorly hydrophobic sequences. Additionally, we observe that the integral hairpin can be stabilized by a short loop heavily populated by turn-promoting residues. We conclude that the combined effect of TM-TM electrostatic interactions and tight turns plays an important role in generating the functional architecture of membrane proteins and propose that helical hairpin motifs can be acquired within the context of the Sec61 translocon at the early stages of membrane protein biosynthesis. Taken together, these data further underline the potential complexities involved in accurately predicting TM domains from primary structures.

  • 18. Barisic, Ivan
    et al.
    Schoenthaler, Silvia
    Ke, Rongqin
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Nilsson, Mats
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Noehammer, Christa
    Wiesinger-Mayr, Herbert
    Multiplex detection of antibiotic resistance genes using padlock probes2013In: Diagnostic microbiology and infectious disease, ISSN 0732-8893, E-ISSN 1879-0070, Vol. 77, no 2, 118-125 p.Article in journal (Refereed)
    Abstract [en]

    The elucidation of resistance mechanisms is of central importance to providing and maintaining efficient medical treatment. However, molecular detection methods covering the complete set of resistance genes with a single test are still missing. Here, we present a novel 100-plex assay based on padlock probes in combination with a microarray that allows the simultaneous large-scale identification of highly diverse beta-lactamases. The specificity of the assay was performed using 70 clinical bacterial isolates, recovering 98% of the beta-lactamase nucleotide sequences present. Additionally, the sensitivity was evaluated with PCR products and genomic bacterial DNA, revealing a detection limit of 10(4) DNA copies per reaction when using PCR products as the template. Pre-amplification of genomic DNA in a 25-multiplex PCR further facilitated the detection of beta-lactamase genes in dilutions of 10(7) cells/mL. In summary, we present an efficient, highly specific, and highly sensitive multiplex detection method for any gene.

  • 19. Barrio, Alvaro Martinez
    et al.
    Lamichhaney, Sangeet
    Fan, Guangyi
    Rafati, Nima
    Pettersson, Mats
    Zhang, He
    Dainat, Jacques
    Ekman, Diana
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Hoppner, Marc
    Jern, Patric
    Martin, Marcel
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Nystedt, Björn
    Liu, Xin
    Chen, Wenbin
    Liang, Xinming
    Shi, Chengcheng
    Fu, Yuanyuan
    Ma, Kailong
    Zhan, Xiao
    Feng, Chungang
    Gustafson, Ulla
    Rubin, Carl-Johan
    Almen, Markus Sallman
    Blass, Martina
    Casini, Michele
    Folkvord, Arild
    Laikre, Linda
    Stockholm University, Faculty of Science, Department of Zoology.
    Ryman, Nils
    Stockholm University, Faculty of Science, Department of Zoology.
    Lee, Simon Ming-Yuen
    Xu, Xun
    Andersson, Leif
    The genetic basis for ecological adaptation of the Atlantic herring revealed by genome sequencing2016In: eLIFE, E-ISSN 2050-084X, Vol. 5, e12081Article in journal (Refereed)
    Abstract [en]

    Ecological adaptation is of major relevance to speciation and sustainable population management, but the underlying genetic factors are typically hard to study in natural populations due to genetic differentiation caused by natural selection being confounded with genetic drift in subdivided populations. Here, we use whole genome population sequencing of Atlantic and Baltic herring to reveal the underlying genetic architecture at an unprecedented detailed resolution for both adaptation to a new niche environment and timing of reproduction. We identify almost 500 independent loci associated with a recent niche expansion from marine (Atlantic Ocean) to brackish waters (Baltic Sea), and more than 100 independent loci showing genetic differentiation between spring- and autumn-spawning populations irrespective of geographic origin. Our results show that both coding and non-coding changes contribute to adaptation. Haplotype blocks, often spanning multiple genes and maintained by selection, are associated with genetic differentiation.

  • 20.
    Basile, Walter
    et al.
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Sachenkova, Oxana
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Light, Sara
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab). Linköping University, Sweden.
    Elofsson, Arne
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab). Kungliga Tekniska Högskolan, Sweden.
    High GC content causes orphan proteins to be intrinsically disordered2017In: PloS Computational Biology, ISSN 1553-734X, E-ISSN 1553-7358, Vol. 13, no 3, e1005375Article in journal (Refereed)
    Abstract [en]

    De novo creation of protein coding genes involves the formation of short ORFs from noncoding regions; some of these ORFs might then become fixed in the population These orphan proteins need to, at the bare minimum, not cause serious harm to the organism, meaning that they should for instance not aggregate. Therefore, although the creation of short ORFs could be truly random, the fixation should be subjected to some selective pressure. The selective forces acting on orphan proteins have been elusive, and contradictory results have been reported. In Drosophila young proteins are more disordered than ancient ones, while the opposite trend is present in yeast. To the best of our knowledge no valid explanation for this difference has been proposed. To solve this riddle we studied structural properties and age of proteins in 187 eukaryotic organisms. We find that, with the exception of length, there are only small differences in the properties between proteins of different ages. However, when we take the GC content into account we noted that it could explain the opposite trends observed for orphans in yeast (low GC) and Drosophila (high GC). GC content is correlated with codons coding for disorder promoting amino acids. This leads us to propose that intrinsic disorder is not a strong determining factor for fixation of orphan proteins. Instead these proteins largely resemble random proteins given a particular GC level. During evolution the properties of a protein change faster than the GC level causing the relationship between disorder and GC to gradually weaken.

  • 21. Beghini, Alessandro
    et al.
    Corlazzoli, Francesca
    Del Giacco, Luca
    Re, Matteo
    Lazzaroni, Francesca
    Brioschi, Matteo
    Valentini, Giorgio
    Ferrazzi, Fulvia
    Ghilardi, Anna
    Righi, Marco
    Turrini, Mauro
    Mignardi, Marco
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Cesana, Clara
    Bronte, Vincenzo
    Nilsson, Mats
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Morra, Enrica
    Cairoli, Roberto
    Regeneration-associated WNT Signaling Is Activated in Long-term Reconstituting AC133(bright) Acute Myeloid Leukemia Cells2012In: Neoplasia, ISSN 1522-8002, Vol. 14, no 12, 1236-+ p.Article in journal (Refereed)
    Abstract [en]

    Acute myeloid leukemia (AML) is a genetically heterogeneous clonal disorder characterized by two molecularly distinct self-renewing leukemic stem cell (LSC) populations most closely related to normal progenitors and organized as a hierarchy. A requirement for WNT/beta-catenin signaling in the pathogenesis of AML has recently been suggested by a mouse model. However, its relationship to a specific molecular function promoting retention of self-renewing leukemia-initiating cells (LICs) in human remains elusive. To identify transcriptional programs involved in the maintenance of a self-renewing state in LICs, we performed the expression profiling in normal (n = 10) and leukemic (n = 33) human long-term reconstituting AC133(+) cells, which represent an expanded cell population in most AML patients. This study reveals the ligand-dependent WNT pathway activation in AC133(bright) AML cells and shows a diffuse expression and release of WNT 10B, a hematopoietic stem cell regenerative-associated molecule. The establishment of a primary AC133(+) AML cell culture (A46) demonstrated that leukemia cells synthesize and secrete WNT ligands, increasing the levels of dephosphorylated beta-catenin in vivo. We tested the LSC functional activity in AC133(+) cells and found significant levels of engraftment upon transplantation of A46 cells into irradiated Rag2(-/-)gamma c(-/-) mice. Owing to the link between hematopoietic regeneration and developmental signaling, we transplanted A46 cells into developing zebrafish. This system revealed the formation of ectopic structures by activating dorsal organizer markers that act downstream of the WNT pathway. In conclusion, our findings suggest that AC133(bright) LSCs are promoted by misappropriating homeostatic WNT programs that control hematopoietic regeneration. Neoplasia (2012) 14, 1236-1248

  • 22. Bejhed, Rebecca S.
    et al.
    Strömme, Maria
    Svedlindh, Peter
    Ahlford, Annika
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Strömberg, Mattias
    Magnetic nanobeads present during enzymatic amplification and labeling for a simplified DNA detection protocol based on AC susceptometry2015In: AIP Advances, ISSN 2158-3226, E-ISSN 2158-3226, Vol. 5, no 12, 127139Article in journal (Refereed)
    Abstract [en]

    Magnetic biosensors are promising candidates for low-cost point-of-care biodiagnostic devices. For optimal efficiency it is crucial to minimize the time and complexity of the assay protocol including target recognition, amplification, labeling and read-out. In this work, possibilities for protocol simplifications for a DNA biodetection principle relying on hybridization of magnetic nanobeads to rolling circle amplification (RCA) products are investigated. The target DNA is recognized through a padlock ligation assay resulting in DNA circles serving as templates for the RCA process. It is found that beads can be present during amplification without noticeably interfering with the enzyme used for RCA (phi29 polymerase). As a result, the bead-coil hybridization can be performed immediately after amplification in a one-step manner at elevated temperature within a few minutes prior to read-out in an AC susceptometer setup, i.e. a combined protocol approach. Moreover, by recording the phase angle xi = arctan(chi ''/chi'), where chi and chi '' are the in-phase and out-of-phase components of the AC susceptibility, respectively, at one single frequency the total assay time for the optimized combined protocol would be no more than 1.5 hours, often a relevant time frame for diagnosis of cancer and infectious disease. Also, applying the phase angle method normalization of AC susceptibility data is not needed. These findings are useful for the development of point-of-care biodiagnostic devices relying on bead-coil binding and magnetic AC susceptometry.

  • 23.
    Bendz, Maria
    et al.
    Stockholm University, Science for Life Laboratory (SciLifeLab). Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
    Skwark, Marcin
    Stockholm University, Science for Life Laboratory (SciLifeLab). Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
    Nilsson, Daniel
    Stockholm University, Science for Life Laboratory (SciLifeLab). Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
    Granholm, Viktor
    Stockholm University, Science for Life Laboratory (SciLifeLab). Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
    Cristobal, Susana
    Kall, Lukas
    Elofsson, Arne
    Stockholm University, Science for Life Laboratory (SciLifeLab). Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
    Membrane protein shaving with thermolysin can be used to evaluate topology predictors2013In: Proteomics, ISSN 1615-9853, E-ISSN 1615-9861, Vol. 13, no 9, 1467-1480 p.Article in journal (Refereed)
    Abstract [en]

    Topology analysis of membrane proteins can be obtained by enzymatic shaving in combination with MS identification of peptides. Ideally, such analysis could provide quite detailed information about the membrane spanning regions. Here, we examine the ability of some shaving enzymes to provide large-scale analysis of membrane proteome topologies. To compare different shaving enzymes, we first analyzed the detected peptides from two over-expressed proteins. Second, we analyzed the peptides from non-over-expressed Escherichia coli membrane proteins with known structure to evaluate the shaving methods. Finally, the identified peptides were used to test the accuracy of a number of topology predictors. At the end we suggest that the usage of thermolysin, an enzyme working at the natural pH of the cell for membrane shaving, is superior because: (i) we detect a similar number of peptides and proteins using thermolysin and trypsin; (ii) thermolysin shaving can be run at a natural pH and (iii) the incubation time is quite short. (iv) Fewer detected peptides from thermolysin shaving originate from the transmembrane regions. Using thermolysin shaving we can also provide a clear separation between the best and the less accurate topology predictors, indicating that using data from shaving can provide valuable information when developing new topology predictors.

  • 24. Bengtsson-Palme, Johan
    et al.
    Angelin, Martin
    Huss, Mikael
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Kjellqvist, Sanela
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Kristiansson, Erik
    Palmgren, Helena
    Larsson, D. G. Joakim
    Johansson, Anders
    The Human Gut Microbiome as a Transporter of Antibiotic Resistance Genes between Continents2015In: Antimicrobial Agents and Chemotherapy, ISSN 0066-4804, E-ISSN 1098-6596, Vol. 59, no 10, 6551-6560 p.Article in journal (Refereed)
    Abstract [en]

    Previous studies of antibiotic resistance dissemination by travel have, by targeting only a select number of cultivable bacterial species, omitted most of the human microbiome. Here, we used explorative shotgun metagenomic sequencing to address the abundance of >300 antibiotic resistance genes in fecal specimens from 35 Swedish students taken before and after exchange programs on the Indian peninsula or in Central Africa. All specimens were additionally cultured for extended-spectrum beta-lactamase (ESBL)-producing enterobacteria, and the isolates obtained were genome sequenced. The overall taxonomic diversity and composition of the gut microbiome remained stable before and after travel, but there was an increasing abundance of Proteobacteria in 25/35 students. The relative abundance of antibiotic resistance genes increased, most prominently for genes encoding resistance to sulfonamide (2.6-fold increase), trimethoprim (7.7-fold), and beta-lactams (2.6-fold). Importantly, the increase observed occurred without any antibiotic intake. Of 18 students visiting the Indian peninsula, 12 acquired ESBL-producing Escherichia coli, while none returning from Africa were positive. Despite deep sequencing efforts, the sensitivity of metagenomics was not sufficient to detect acquisition of the low-abundant genes responsible for the observed ESBL phenotype. In conclusion, metagenomic sequencing of the intestinal microbiome of Swedish students returning from exchange programs in Central Africa or the Indian peninsula showed increased abundance of genes encoding resistance to widely used antibiotics.

  • 25.
    Berg, Carlo
    et al.
    Stockholm University, Faculty of Science, Department of Ecology, Environment and Plant Sciences. Stockholm University, Science for Life Laboratory (SciLifeLab). Leibniz Institute for Baltic Sea Research Warnemünde (IOW), Germany.
    Vandieken, Verona
    Thamdrup, Bo
    Juergens, Klaus
    Significance of archaeal nitrification in hypoxic waters of the Baltic Sea2015In: The ISME Journal, ISSN 1751-7362, E-ISSN 1751-7370, Vol. 9, no 6, 1319-1332 p.Article in journal (Refereed)
    Abstract [en]

    Ammonia-oxidizing archaea (AOA) of the phylum Thaumarchaeota are widespread, and their abundance in many terrestrial and aquatic ecosystems suggests a prominent role in nitrification. AOA also occur in high numbers in oxygen-deficient marine environments, such as the pelagic redox gradients of the central Baltic Sea; however, data on archaeal nitrification rates are scarce and little is known about the factors, for example sulfide, that regulate nitrification in this system. In the present work, we assessed the contribution of AOA to ammonia oxidation rates in Baltic deep basins and elucidated the impact of sulfide on this process. Rate measurements with N-15-labeled ammonium, CO2 dark fixation measurements and quantification of AOA by catalyzed reporter deposition-fluorescence in situ hybridization revealed that among the three investigated sites the highest potential nitrification rates (122-884 nmol l(-1) per day) were measured within gradients of decreasing oxygen, where thaumarchaeotal abundance was maximal (2.5-6.9 x 10(5) cells per ml) and CO2 fixation elevated. In the presence of the archaeal-specific inhibitor GC7, nitrification was reduced by 86-100%, confirming the assumed dominance of AOA in this process. In samples spiked with sulfide at concentrations similar to those of in situ conditions, nitrification activity was inhibited but persisted at reduced rates. This result together with the substantial nitrification potential detected in sulfidic waters suggests the tolerance of AOA to periodic mixing of anoxic and sulfidic waters. It begs the question of whether the globally distributed Thaumarchaeota respond similarly in other stratified water columns or whether the observed robustness against sulfide is a specific feature of the thaumarchaeotal subcluster present in the Baltic Deeps.

  • 26.
    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, 2066-2070 p.Article 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.

  • 27. 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, 615-628 p.Article 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.

  • 28.
    Botelho, Salome Calado
    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.
    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, 4792-4798 p.Article 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.

  • 29. 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, 59-+ p.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.

  • 30.
    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, 4442-4455 p.Article 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.

  • 31. 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, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 8, no 7, 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.

  • 32.
    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, 1307-1316 p.Article 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.

  • 33. 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, 1422-1434 p.Article 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.

  • 34.
    Calado Botelho, Salomé
    et al.
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
    Tatsuta, Takashi
    von Heijne, Gunnar
    Stockholm University, Science for Life Laboratory (SciLifeLab).
    Hyun, Kim
    Dislocation by the m-AAA protease increases the threshold hydrophobicity for retention of transmembrane helices in the inner membrane of yeast mitochondria2012Manuscript (preprint) (Other academic)
    Abstract [en]

    Sorting of mitochondrial inner membraneproteins is a complex process where transloconsand proteases function in a concerted way.Many inner membrane proteins insert into themembrane via the TIM23 translocon and someare then further acted upon by themitochondrial m-AAA protease, a molecularmotor capable of dislocating proteins from theinner membrane. This raises the possibility thatthe threshold hydrophobicity for the retentionof transmembrane segments in the innermembrane is different, depending on whetherthey belong to membrane proteins that are m-AAA protease substrate or not. Here, usingmodel transmembrane segments engineeredinto m-AAA protease-dependent proteins, weshow that the threshold hydrophobicity formembrane retention measured in yeast cells inthe absence of a functional m-AAA protease ismarkedly lower than that measured in itspresence. Whether a given hydrophobicsegment in a mitochondrial inner membraneprotein will ultimately form a transmembranehelix may therefore depend on whether or not,during biogenesis, it will be exposed to thepulling force exerted by the m-AAA protease.

  • 35. 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, 97-106 p.Article 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.

  • 36. 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, 65-71 p.Article 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.

  • 37.
    Celepli, Narin
    et al.
    Stockholm University, Faculty of Science, Department of Ecology, Environment and Plant Sciences. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Sundh, John
    Ekman, Martin
    Stockholm University, Faculty of Science, Department of Ecology, Environment and Plant Sciences.
    Dupont, Chris L.
    Yooseph, Shibu
    Bergman, Birgitta
    Stockholm University, Faculty of Science, Department of Ecology, Environment and Plant Sciences.
    Ininbergs, Karolina
    Stockholm University, Faculty of Science, Department of Ecology, Environment and Plant Sciences.
    Meta-omic analyses of Baltic Sea cyanobacteria: diversity, community structure and salt acclimation2017In: Environmental Microbiology, ISSN 1462-2912, E-ISSN 1462-2920, Vol. 19, no 2, 673-686 p.Article in journal (Refereed)
    Abstract [en]

    Cyanobacteria are important phytoplankton in the Baltic Sea, an estuarine-like environment with pronounced north to south gradients in salinity and nutrient concentrations. Here, we present a metagenomic and -transcriptomic survey, with subsequent analyses targeting the genetic identity, phylogenetic diversity, and spatial distribution of Baltic Sea cyanobacteria. The cyanobacterial community constituted close to 12% of the microbial population sampled during a pre-bloom period (June-July 2009). The community was dominated by unicellular picocyanobacteria, specifically a few highly abundant taxa (Synechococcus and Cyanobium) with a long tail of low abundance representatives, and local peaks of bloom-forming heterocystous taxa. Cyanobacteria in the Baltic Sea differed genetically from those in adjacent limnic and marine waters as well as from cultivated and sequenced picocyanobacterial strains. Diversity peaked at brackish salinities 3.5-16psu, with low N:P ratios. A shift in community composition from brackish to marine strains was accompanied by a change in the repertoire and expression of genes involved in salt acclimation. Overall, the pre-bloom cyanobacterial population was more genetically diverse, widespread and abundant than previously documented, with unicellular picocyanobacteria being the most abundant clade along the entire Baltic Sea salinity gradient.

  • 38.
    Celorio-Mancera, Maria de la Paz
    et al.
    Stockholm University, Faculty of Science, Department of Zoology, Animal Ecology.
    Sundmalm, Sara M.
    Stockholm University, Faculty of Science, Department of Zoology, Animal Ecology.
    Vogel, Heiko
    Rutishauser, Dorothea
    Stockholm University, Science for Life Laboratory (SciLifeLab).
    Ytterberg, A. Jimmy
    Zubarev, Roman A.
    Stockholm University, Science for Life Laboratory (SciLifeLab).
    Janz, Niklas
    Stockholm University, Faculty of Science, Department of Zoology, Animal Ecology.
    Chemosensory proteins, major salivary factors in caterpillar mandibular glands2012In: Insect Biochemistry and Molecular Biology, ISSN 0965-1748, E-ISSN 1879-0240, Vol. 42, no 10, 796-805 p.Article in journal (Refereed)
    Abstract [en]

    Research in the field of insect-host plant interactions has indicated that constituents of insect saliva play an important role in digestion and affect host chemical defense responses. However, most efforts have focused on studying the composition and function of regurgitant or saliva produced in the labial glands. Acknowledging the need for understanding the role of the mandibular glands in herbivory, we sought to make a qualitative and semi-quantitative comparison of soluble luminal fractions between mandibular and labial glands of Vanessa gonerilla butterfly larvae. Amylase and lysozyme were inspected as possible major enzymatic activities in the mandibular glands aiding in pre-digestion and antimicrobial defense. Although detected, neither of these enzymatic activities was prominent in the luminal protein preparation of a particular type of gland. Proteins isolated from the glands were identified by mass spectrometry and by searching an EST-library database generated for four other nymphalid butterfly species, in addition to the public NCBI database. The identified proteins were also quantified from thedata using “Quanty”, an in-house program. The proteomic analysis detected chemosensory proteins as the most abundant luminal proteins in the mandibular glands. In comparison to these proteins, the relative amounts of amylase and lysozyme were much lower in both gland types. Therefore, we speculate that the primary role of the mandibular glands in Lepidopteran larvae is chemoreception which may include the detection of microorganisms on plant surfaces, host plant recognition and communication with conspecifics.

  • 39.
    Celorio-Mancera, Maria de la Paz
    et al.
    Stockholm University, Faculty of Science, Department of Zoology, Population Genetics.
    Wheat, Christopher W.
    Stockholm University, Faculty of Science, Department of Zoology, Population Genetics.
    Huss, Mikael
    Stockholm University, Science for Life Laboratory (SciLifeLab).
    Vezzi, Francesco
    Stockholm University, Science for Life Laboratory (SciLifeLab).
    Neethiraj, Ramprasad
    Stockholm University, Faculty of Science, Department of Zoology, Population Genetics.
    Reimegård, Johan
    Stockholm University, Science for Life Laboratory (SciLifeLab).
    Nylin, Sören
    Stockholm University, Faculty of Science, Department of Zoology, Animal Ecology.
    Janz, Niklas
    Stockholm University, Faculty of Science, Department of Zoology, Animal Ecology.
    Evolutionary history of host use, rather than plant phylogeny, determines gene expression in a generalist butterfly2016In: BMC Evolutionary Biology, ISSN 1471-2148, E-ISSN 1471-2148, Vol. 16, 59Article in journal (Refereed)
    Abstract [en]

    Background: Although most insect species are specialized on one or few groups of plants, there are phytophagous insects that seem to use virtually any kind of plant as food. Understanding the nature of this ability to feed on a wide repertoire of plants is crucial for the control of pest species and for the elucidation of the macroevolutionary mechanisms of speciation and diversification of insect herbivores. Here we studied Vanessa cardui, the species with the widest diet breadth among butterflies and a potential insect pest, by comparing tissue-specific transcriptomes from caterpillars that were reared on different host plants. We tested whether the similarities of gene-expression response reflect the evolutionary history of adaptation to these plants in the Vanessa and related genera, against the null hypothesis of transcriptional profiles reflecting plant phylogenetic relatedness. Result: Using both unsupervised and supervised methods of data analysis, we found that the tissue-specific patterns of caterpillar gene expression are better explained by the evolutionary history of adaptation of the insects to the plants than by plant phylogeny. Conclusion: Our findings suggest that V. cardui may use two sets of expressed genes to achieve polyphagy, one associated with the ancestral capability to consume Rosids and Asterids, and another allowing the caterpillar to incorporate a wide range of novel host-plants.

  • 40. Chen, Dan
    et al.
    Ranganathan, Anirudh
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Ijzerman, Adriaan P.
    Siegal, Gregg
    Carlsson, Jens
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Complementarity between in Silico and Biophysical Screening Approaches in Fragment-Based Lead Discovery against the A(2A) Adenosine Receptor2013In: Journal of Chemical Information and Modeling, ISSN 1549-9596, E-ISSN 1549-960X, Vol. 53, no 10, 2701-2714 p.Article in journal (Refereed)
    Abstract [en]

    Fragment-based lead discovery (FBLD) is becoming an increasingly important method in drug development. We have explored the potential to complement NMR-based biophysical screening of chemical libraries with molecular docking in FBLD against the A(2A) adenosine receptor (A(2A)AR), a drug target for inflammation and Parkinson's disease. Prior to an NMR-based screen of a fragment library against the A(2A)AR, molecular docking against a crystal structure was used to rank the same set of molecules by their predicted affinities. Molecular docking was able to predict four out of the five orthosteric ligands discovered by NMR among the top 5% of the ranked library, suggesting that structure-based methods could be used to prioritize among primary hits from biophysical screens. In addition, three fragments that were top-ranked by molecular docking, but had not been picked up by the NMR-based method, were demonstrated to be A2AAR ligands. While biophysical approaches for fragment screening are typically limited to a few thousand compounds, the docking screen was extended to include 328,000 commercially available fragments. Twenty-two top-ranked compounds were tested in radioligand binding assays, and 14 of these were A(2A)AR ligands with K-i values ranging from 2 to 240 mu M. Optimization of fragments was guided by molecular dynamics simulations and free energy calculations. The results illuminate strengths and weaknesses of molecular docking and demonstrate that this method can serve as a valuable complementary tool to biophysical screening in FBLD.

  • 41. Chinh, Bkrong
    et al.
    Alsøe, Lene
    Lindvall, Jessica M.
    Stockholm University, Science for Life Laboratory (SciLifeLab). Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. National Bioinformatics Infrastructure Sweden (NBIS), Sweden.
    Sulheim, Dag
    Fagermoen, Even
    Winger, Anette
    Kaarbø, Mari
    Nilsen, Hilde
    Bruun Wyller, Vegard
    Whole blood gene expression in adolescent chronic fatigue syndrome: an exploratory cross-sectional study suggesting altered B cell differentiation and survival2017In: Journal of Translational Medicine, ISSN 1479-5876, E-ISSN 1479-5876, Vol. 15, 102Article in journal (Refereed)
    Abstract [en]

    Background

    Chronic fatigue syndrome (CFS) is a prevalent and disabling condition affecting adolescents. The pathophysiology is poorly understood, but immune alterations might be an important component. This study compared whole blood gene expression in adolescent CFS patients and healthy controls, and explored associations between gene expression and neuroendocrine markers, immune markers and clinical markers within the CFS group.

    Methods

    CFS patients (12-18 years old) were recruited nation-wide to a single referral center as part of the Nor-CAPITAL project. A broad case definition of CFS was applied, requiring 3 months of unexplained, disabling chronic/ relapsing fatigue of new onset, whereas no accompanying symptoms were necessary. Healthy controls having comparable distribution of gender and age were recruited from local schools. Whole blood samples were subjected to RNA sequencing. Immune markers were blood leukocyte counts, plasma cytokines, serum C-reactive protein and immunoglobulins. Neuroendocrine markers encompassed plasma and urine levels of catecholamines and cortisol, as well as heart rate variability indices. Clinical markers consisted of questionnaire scores for symptoms of post-exertional malaise, inflammation, fatigue, depression and trait anxiety, as well as activity recordings.

    Results

    A total of 29 CFS patients and 18 healthy controls were included. We identified 176 genes as differentially expressed in patients compared to controls, adjusting for age and gender factors. Gene set enrichment analyses suggested impairment of B cell differentiation and survival, as well as enhancement of innate antiviral responses and inflammation in the CFS group. A pattern of co-expression could be identified, and this pattern, as well as single gene transcripts, was significantly associated with indices of autonomic nervous activity, plasma cortisol, and blood monocyte and eosinophil counts. Also, an association with symptoms of post-exertional malaise was demonstrated.

    Conclusion

    Adolescent CFS is characterized by differential gene expression pattern in whole blood suggestive of impaired B cell differentiation and survival, and enhanced innate antiviral responses and inflammation. This expression pattern is associated with neuroendocrine markers of altered HPA axis and autonomic nervous activity, and with symptoms of post-exertional malaise.

  • 42. Clausson, Carl-Magnus
    et al.
    Arngården, Linda
    Ishaq, Omer
    Klaesson, Axel
    Kühnemund, Malte
    Grannas, Karin
    Koos, Björn
    Qian, Xiaoyan
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Ranefall, Petter
    Krzywkowski, Tomasz
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Brismar, Hjalmar
    Nilsson, Mats
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Wählby, Carolina
    Söderberg, Ola
    Compaction of rolling circle amplification products increases signal integrity and signal-to-noise ratio2015In: Scientific Reports, ISSN 2045-2322, Vol. 5, 12317Article in journal (Refereed)
    Abstract [en]

    Rolling circle amplification (RCA) for generation of distinct fluorescent signals in situ relies upon the self-collapsing properties of single-stranded DNA in commonly used RCA-based methods. By introducing a cross-hybridizing DNA oligonucleotide during rolling circle amplification, we demonstrate that the fluorophore-labeled RCA products (RCPs) become smaller. The reduced size of RCPs increases the local concentration of fluorophores and as a result, the signal intensity increases together with the signal-to-noise ratio. Furthermore, we have found that RCPs sometimes tend to disintegrate and may be recorded as several RCPs, a trait that is prevented with our cross-hybridizing DNA oligonucleotide. These effects generated by compaction of RCPs improve accuracy of visual as well as automated in situ analysis for RCA based methods, such as proximity ligation assays (PLA) and padlock probes.

  • 43. Conti, Luca
    et al.
    Renhorn, Jakob
    Gabrielsson, Anders
    Turesson, Fredrik
    Liin, Sara I.
    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.
    Elinder, Fredrik
    Reciprocal voltage sensor-to-pore coupling leads to potassium channel C-type inactivation2016In: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 6, 27562Article in journal (Refereed)
    Abstract [en]

    Voltage-gated potassium channels open at depolarized membrane voltages. A prolonged depolarization causes a rearrangement of the selectivity filter which terminates the conduction of ions - a process called slow or C-type inactivation. How structural rearrangements in the voltage-sensor domain (VSD) cause alteration in the selectivity filter, and vice versa, are not fully understood. We show that pulling the pore domain of the Shaker potassium channel towards the VSD by a Cd2+ bridge accelerates C-type inactivation. Molecular dynamics simulations show that such pulling widens the selectivity filter and disrupts the K+ coordination, a hallmark for C-type inactivation. An engineered Cd2+ bridge within the VSD also affect C-type inactivation. Conversely, a pore domain mutation affects VSD gating-charge movement. Finally, C-type inactivation is caused by the concerted action of distant amino acid residues in the pore domain. All together, these data suggest a reciprocal communication between the pore domain and the VSD in the extracellular portion of the channel.

  • 44. Corcoran, Martin M.
    et al.
    Phad, Ganesh E.
    Bernat, Nestor Vazquez
    Stahl-Hennig, Christiane
    Sumida, Noriyuki
    Persson, Mats A. A.
    Martin, Marcel
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Hedestam, Gunilla B. Karlsson
    Production of individualized V gene databases reveals high levels of immunoglobulin genetic diversity2016In: nature communications, ISSN 2041-1723, Vol. 7, 13642Article in journal (Refereed)
    Abstract [en]

    Comprehensive knowledge of immunoglobulin genetics is required to advance our understanding of B cell biology. Validated immunoglobulin variable (V) gene databases are close to completion only for human and mouse. We present a novel computational approach, IgDiscover, that identifies germline V genes from expressed repertoires to a specificity of 100%. IgDiscover uses a cluster identification process to produce candidate sequences that, once filtered, results in individualized germline V gene databases. IgDiscover was tested in multiple species, validated by genomic cloning and cross library comparisons and produces comprehensive gene databases even where limited genomic sequence is available. IgDiscover analysis of the allelic content of the Indian and Chinese-origin rhesus macaques reveals high levels of immunoglobulin gene diversity in this species. Further, we describe a novel human IGHV3-21 allele and confirm significant gene differences between Balb/c and C57BL6 mouse strains, demonstrating the power of IgDiscover as a germline V gene discovery tool.

  • 45.
    Cymer, Florian
    et al.
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
    Hedman, Rickard
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
    Ismail, Nurzian
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
    von Heijne, Gunnar
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Exploration of the Arrest Peptide Sequence Space Reveals Arrest-enhanced Variants2015In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 290, no 16, 10208-10215 p.Article in journal (Refereed)
    Abstract [en]

    Translational arrest peptides (APs) are short stretches of polypeptides that induce translational stalling when synthesized on a ribosome. Mechanical pulling forces acting on the nascent chain can weaken or even abolish stalling. APs can therefore be used as in vivo force sensors, making it possible to measure the forces that act on a nascent chain during translation with single-residue resolution. It is also possible to score the relative strengths of APs by subjecting them to a given pulling force and ranking them according to stalling efficiency. Using the latter approach, we now report an extensive mutagenesis scan of a strong mutant variant of the Mannheimia succiniciproducens SecM AP and identify mutations that further increase the stalling efficiency. Combining three such mutations, we designed an AP that withstands the strongest pulling force we are able to generate at present. We further show that diproline stretches in a nascent protein act as very strong APs when translation is carried out in the absence of elongation factor P. Our findings highlight critical residues in APs, show that certain amino acid sequences induce very strong translational arrest and provide a toolbox of APs of varying strengths that can be used for in vivo force measurements.

  • 46.
    Cymer, Florian
    et al.
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
    Ismail, Nurzian
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
    von Heijne, Gunnar
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Weak pulling forces exerted on N-in-orientated transmembrane segments during co-translational insertion into the inner membrane of Escherichia coli2014In: FEBS Letters, ISSN 0014-5793, E-ISSN 1873-3468, Vol. 588, no 10, 1930-1934 p.Article in journal (Refereed)
    Abstract [en]

    Transmembrane helices (TMHs) in membrane proteins can be orientated with their N-terminus towards the cytoplasm (N-in), or facing the non-cytoplasmic side (N-out). Most membrane proteins are inserted co-translationally into membranes, aided by Sec-type translocons. Since the final orientation of N-in-and N-out-orientated TMHs differs, they could also interact differently with the translocon and the surrounding membrane during insertion. We measured pulling forces exerted on N-in-orientated TMHs during co-translational insertion into the inner membrane of Escherichia coli. Our results demonstrate that Nin-orientated TMHs experience a weaker pulling force but retain the overall biphasic force profile seen previously for Nout-orientated TMHs (Ismail et al., 2012 [1]).

  • 47.
    Cymer, Florian
    et al.
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
    von Heijne, Gunnar
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Cotranslational folding of membrane proteins probed by arrest-peptide-mediated force measurements2013In: Proceedings of the National Academy of Sciences of the United States of America, ISSN 0027-8424, E-ISSN 1091-6490, Vol. 110, no 36, 14640-14645 p.Article in journal (Refereed)
    Abstract [en]

    Polytopic membrane proteins are inserted cotranslationally into target membranes by ribosome-translocon complexes. It is, however, unclear when during the insertion process specific interactions between the transmembrane helices start to form. Here, we use a recently developed in vivo technique to measure pulling forces acting on transmembrane helices during their cotranslational insertion into the inner membrane of Escherichia coli to study the earliest steps of tertiary folding of five polytopic membrane proteins. We find that interactions between residues in a C-terminally located transmembrane helix and in more N-terminally located helices can be detected already at the point when the C-terminal helix partitions from the translocon into the membrane. Our findings pinpoint the earliest steps of tertiary structure formation and open up possibilities to study the cotranslational folding of polytopic membrane proteins.

  • 48.
    Cymer, Florian
    et al.
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
    von Heijne, Gunnar
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    White, Stephen H.
    Mechanisms of Integral Membrane Protein Insertion and Folding2015In: Journal of Molecular Biology, ISSN 0022-2836, E-ISSN 1089-8638, Vol. 427, no 5, 999-1022 p.Article, review/survey (Refereed)
    Abstract [en]

    The biogenesis, folding, and structure of alpha-helical membrane proteins (MPs) are important to understand because they underlie virtually all physiological processes in cells including key metabolic pathways, such as the respiratory chain and the photosystems, as well as the transport of solutes and signals across membranes. Nearly all MPs require translocons-often referred to as protein-conducting channels-for proper insertion into their target membrane. Remarkable progress toward understanding the structure and functioning of translocons has been made during the past decade. Here, we review and assess this progress critically. All available evidence indicates that MPs are equilibrium structures that achieve their final structural states by folding along thermodynamically controlled pathways. The main challenge for cells is the targeting and membrane insertion of highly hydrophobic amino acid sequences. Targeting and insertion are managed in cells principally by interactions between ribosomes and membrane-embedded translocons. Our review examines the biophysical and biological boundaries of MP insertion and the folding of polytopic MPs in vivo. A theme of the review is the under-appreciated role of basic thermodynamic principles in MP folding and assembly. Thermodynamics not only dictates the final folded structure but also is the driving force for the evolution of the ribosome-translocon system of assembly. We conclude the review with a perspective suggesting a new view of translocon-guided MP insertion. (C) 2014 Elsevier Ltd. All rights reserved.

  • 49. Dalin, Martin G.
    et al.
    Engström, Pär G.
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Ivarsson, Emil G.
    Unneberg, Per
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Light, Sara
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Schaufelberger, Maria
    Gilljama, Thomas
    Andersson, Bert
    Bergo, Martin O.
    Massive parallel sequencing questions the pathogenic role of missense variants in dilated cardiomyopathy2017In: International Journal of Cardiology, ISSN 0167-5273, E-ISSN 1874-1754, Vol. 228, 742-748 p.Article in journal (Refereed)
    Abstract [en]

    Background: Germline genetic variants are an important cause of dilated cardiomyopathy (DCM). However, recent sequencing studies have revealed rare variants in DCM-associated genes also in individuals without known heart disease. In this study, we investigate variant prevalence and genotype-phenotype correlations in Swedish DCM patients, and compare their genetic variants to those detected in reference cohorts. Methods and results: We sequenced the coding regions of 41 DCM-associated genes in 176 unrelated patients with idiopathic DCM and found 102 protein-altering variants with an allele frequency of <0.04% in reference cohorts; the majority were missense variants not previously described in DCM. Fifty-five (31%) patients had one variant, and 24 (14%) patients had two or more variants in the analysed genes. Detection of genetic variants in any gene, and in LMNA, MYII7 or TTN alone, was associated with early onset disease and reduced transplant-free survival. As expected, nonsense and frameshift variants were more common in DCM patients than in healthy individuals of the reference cohort 1000 Genomes Europeans. Surprisingly however, the prevalence, conservation and pathogenicity scores, and localization of missense variants were similar in DCM patients and healthy reference individuals. Conclusion: To our knowledge, this is the first study to identify correlations between genotype and prognosis when sequencing a large number of genes in unselected DCM patients. The similar distribution of missense variants in DCM patients and healthy reference individuals questions the pathogenic role of many variants, and suggests that results from genetic testing of DCM patients should be interpreted with caution.

  • 50. Danielsson, Frida
    et al.
    James, Tojo
    Gomez-Cabrero, David
    Huss, Mikael
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Assessing the consistency of public human tissue RNA-seq data sets2015In: Briefings in Bioinformatics, ISSN 1467-5463, E-ISSN 1477-4054, Vol. 16, no 6, 941-949 p.Article in journal (Refereed)
    Abstract [en]

    Sequencing-based gene expression methods like RNA-sequencing (RNA-seq) have become increasingly common, but it is often claimed that results obtained in different studies are not comparable owing to the influence of laboratory batch effects, differences in RNA extraction and sequencing library preparation methods and bioinformatics processing pipelines. It would be unfortunate if different experiments were in fact incomparable, as there is great promise in data fusion and meta-analysis applied to sequencing data sets. We therefore compared reported gene expression measurements for ostensibly similar samples (specifically, human brain, heart and kidney samples) in several different RNA-seq studies to assess their overall consistency and to examine the factors contributing most to systematic differences. The same comparisons were also performed after preprocessing all data in a consistent way, eliminating potential bias from bioinformatics pipelines. We conclude that published human tissue RNA-seq expression measurements appear relatively consistent in the sense that samples cluster by tissue rather than laboratory of origin given simple preprocessing transformations. The article is supplemented by a detailed walkthrough with embedded R code and figures.

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