Time and space-resolved quantification of plasma membrane sialylation for measurements of cell function and neurotoxicity

Lade...
Vorschaubild
Dateien
Zu diesem Dokument gibt es keine Dateien.
Datum
2020
Herausgeber:innen
Kontakt
ISSN der Zeitschrift
Electronic ISSN
ISBN
Bibliografische Daten
Verlag
Schriftenreihe
Auflagebezeichnung
ArXiv-ID
Internationale Patentnummer
Link zur Lizenz
Angaben zur Forschungsförderung
European Union (EU): 681002
Projekt
EUToxRisk21
Open Access-Veröffentlichung
Open Access Hybrid
Sammlungen
Core Facility der Universität Konstanz
Gesperrt bis
Titel in einer weiteren Sprache
Forschungsvorhaben
Organisationseinheiten
Zeitschriftenheft
Publikationstyp
Zeitschriftenartikel
Publikationsstatus
Published
Erschienen in
Archives of Toxicology. Springer Science and Business Media. 2020, 94(2), pp. 449-467. ISSN 0340-5761. eISSN 1432-0738. Available under: doi: 10.1007/s00204-019-02642-z
Zusammenfassung

While there are many methods to quantify the synthesis, localization, and pool sizes of proteins and DNA during physiological responses and toxicological stress, only few approaches allow following the fate of carbohydrates. One of them is metabolic glycoengineering (MGE), which makes use of chemically modified sugars (CMS) that enter the cellular biosynthesis pathways leading to glycoproteins and glycolipids. The CMS can subsequently be coupled (via bio-orthogonal chemical reactions) to tags that are quantifiable by microscopic imaging. We asked here, whether MGE can be used in a quantitative and time-resolved way to study neuronal glycoprotein synthesis and its impairment. We focused on the detection of sialic acid (Sia), by feeding human neurons the biosynthetic precursor N-acetyl-mannosamine, modified by an azide tag. Using this system, we identified non-toxic conditions that allowed live cell labeling with high spatial and temporal resolution, as well as the quantification of cell surface Sia. Using combinations of immunostaining, chromatography, and western blotting, we quantified the percentage of cellular label incorporation and effects on glycoproteins such as polysialylated neural cell adhesion molecule. A specific imaging algorithm was used to quantify Sia incorporation into neuronal projections, as potential measure of complex cell function in toxicological studies. When various toxicants were studied, we identified a subgroup (mitochondrial respiration inhibitors) that affected neurite glycan levels several hours before any other viability parameter was affected. The MGE-based neurotoxicity assay, thus allowed the identification of subtle impairments of neurochemical function with very high sensitivity.

Zusammenfassung in einer weiteren Sprache
Fachgebiet (DDC)
570 Biowissenschaften, Biologie
Schlagwörter
Metabolic glycoengineering, Sialic acid, Neurite integrity, Cytotoxicity, Neurotoxicity, Confocal imaging
Konferenz
Rezension
undefined / . - undefined, undefined
Zitieren
ISO 690KRANASTER, Petra, Christiaan KARREMAN, Jeremias E.G.A. DOLD, Alice KREBS, Melina FUNKE, Anna-Katharina HOLZER, Stefanie KLIMA, Johanna NYFFELER, Stefan HELFRICH, Valentin WITTMANN, Marcel LEIST, 2020. Time and space-resolved quantification of plasma membrane sialylation for measurements of cell function and neurotoxicity. In: Archives of Toxicology. Springer Science and Business Media. 2020, 94(2), pp. 449-467. ISSN 0340-5761. eISSN 1432-0738. Available under: doi: 10.1007/s00204-019-02642-z
BibTex
@article{Kranaster2020-02space-48086,
  year={2020},
  doi={10.1007/s00204-019-02642-z},
  title={Time and space-resolved quantification of plasma membrane sialylation for measurements of cell function and neurotoxicity},
  number={2},
  volume={94},
  issn={0340-5761},
  journal={Archives of Toxicology},
  pages={449--467},
  author={Kranaster, Petra and Karreman, Christiaan and Dold, Jeremias E.G.A. and Krebs, Alice and Funke, Melina and Holzer, Anna-Katharina and Klima, Stefanie and Nyffeler, Johanna and Helfrich, Stefan and Wittmann, Valentin and Leist, Marcel}
}
RDF
<rdf:RDF
    xmlns:dcterms="http://purl.org/dc/terms/"
    xmlns:dc="http://purl.org/dc/elements/1.1/"
    xmlns:rdf="http://www.w3.org/1999/02/22-rdf-syntax-ns#"
    xmlns:bibo="http://purl.org/ontology/bibo/"
    xmlns:dspace="http://digital-repositories.org/ontologies/dspace/0.1.0#"
    xmlns:foaf="http://xmlns.com/foaf/0.1/"
    xmlns:void="http://rdfs.org/ns/void#"
    xmlns:xsd="http://www.w3.org/2001/XMLSchema#" > 
  <rdf:Description rdf:about="https://kops.uni-konstanz.de/server/rdf/resource/123456789/48086">
    <bibo:uri rdf:resource="https://kops.uni-konstanz.de/handle/123456789/48086"/>
    <void:sparqlEndpoint rdf:resource="http://localhost/fuseki/dspace/sparql"/>
    <dcterms:abstract xml:lang="eng">While there are many methods to quantify the synthesis, localization, and pool sizes of proteins and DNA during physiological responses and toxicological stress, only few approaches allow following the fate of carbohydrates. One of them is metabolic glycoengineering (MGE), which makes use of chemically modified sugars (CMS) that enter the cellular biosynthesis pathways leading to glycoproteins and glycolipids. The CMS can subsequently be coupled (via bio-orthogonal chemical reactions) to tags that are quantifiable by microscopic imaging. We asked here, whether MGE can be used in a quantitative and time-resolved way to study neuronal glycoprotein synthesis and its impairment. We focused on the detection of sialic acid (Sia), by feeding human neurons the biosynthetic precursor N-acetyl-mannosamine, modified by an azide tag. Using this system, we identified non-toxic conditions that allowed live cell labeling with high spatial and temporal resolution, as well as the quantification of cell surface Sia. Using combinations of immunostaining, chromatography, and western blotting, we quantified the percentage of cellular label incorporation and effects on glycoproteins such as polysialylated neural cell adhesion molecule. A specific imaging algorithm was used to quantify Sia incorporation into neuronal projections, as potential measure of complex cell function in toxicological studies. When various toxicants were studied, we identified a subgroup (mitochondrial respiration inhibitors) that affected neurite glycan levels several hours before any other viability parameter was affected. The MGE-based neurotoxicity assay, thus allowed the identification of subtle impairments of neurochemical function with very high sensitivity.</dcterms:abstract>
    <dc:creator>Karreman, Christiaan</dc:creator>
    <dcterms:isPartOf rdf:resource="https://kops.uni-konstanz.de/server/rdf/resource/123456789/29"/>
    <dc:language>eng</dc:language>
    <dcterms:title>Time and space-resolved quantification of plasma membrane sialylation for measurements of cell function and neurotoxicity</dcterms:title>
    <dcterms:isPartOf rdf:resource="https://kops.uni-konstanz.de/server/rdf/resource/123456789/28"/>
    <dspace:isPartOfCollection rdf:resource="https://kops.uni-konstanz.de/server/rdf/resource/123456789/29"/>
    <dcterms:hasPart rdf:resource="https://kops.uni-konstanz.de/bitstream/123456789/48086/1/Kranaster_2-18vj6nw36y4cq2.pdf"/>
    <dc:creator>Helfrich, Stefan</dc:creator>
    <dc:creator>Funke, Melina</dc:creator>
    <dc:contributor>Helfrich, Stefan</dc:contributor>
    <dc:creator>Leist, Marcel</dc:creator>
    <dc:contributor>Krebs, Alice</dc:contributor>
    <dc:contributor>Dold, Jeremias E.G.A.</dc:contributor>
    <dcterms:rights rdf:resource="http://creativecommons.org/licenses/by/4.0/"/>
    <dc:contributor>Wittmann, Valentin</dc:contributor>
    <dc:creator>Dold, Jeremias E.G.A.</dc:creator>
    <dcterms:issued>2020-02</dcterms:issued>
    <foaf:homepage rdf:resource="http://localhost:8080/"/>
    <dspace:hasBitstream rdf:resource="https://kops.uni-konstanz.de/bitstream/123456789/48086/1/Kranaster_2-18vj6nw36y4cq2.pdf"/>
    <dc:contributor>Kranaster, Petra</dc:contributor>
    <dc:contributor>Klima, Stefanie</dc:contributor>
    <dc:creator>Kranaster, Petra</dc:creator>
    <dc:creator>Holzer, Anna-Katharina</dc:creator>
    <dc:creator>Wittmann, Valentin</dc:creator>
    <dc:contributor>Holzer, Anna-Katharina</dc:contributor>
    <dc:contributor>Leist, Marcel</dc:contributor>
    <dc:date rdf:datatype="http://www.w3.org/2001/XMLSchema#dateTime">2019-12-18T08:11:20Z</dc:date>
    <dc:contributor>Funke, Melina</dc:contributor>
    <dcterms:available rdf:datatype="http://www.w3.org/2001/XMLSchema#dateTime">2019-12-18T08:11:20Z</dcterms:available>
    <dc:rights>Attribution 4.0 International</dc:rights>
    <dc:creator>Krebs, Alice</dc:creator>
    <dc:creator>Nyffeler, Johanna</dc:creator>
    <dc:creator>Klima, Stefanie</dc:creator>
    <dspace:isPartOfCollection rdf:resource="https://kops.uni-konstanz.de/server/rdf/resource/123456789/28"/>
    <dc:contributor>Karreman, Christiaan</dc:contributor>
    <dc:contributor>Nyffeler, Johanna</dc:contributor>
  </rdf:Description>
</rdf:RDF>
Interner Vermerk
xmlui.Submission.submit.DescribeStep.inputForms.label.kops_note_fromSubmitter
Kontakt
URL der Originalveröffentl.
Prüfdatum der URL
Prüfungsdatum der Dissertation
Finanzierungsart
Kommentar zur Publikation
Allianzlizenz
Corresponding Authors der Uni Konstanz vorhanden
Internationale Co-Autor:innen
Universitätsbibliographie
Ja
Begutachtet
Ja
Diese Publikation teilen