A Versatile Toolkit for Semi-Automated Production of Fluorescent Chemokines to Study CCR7 Expression and Functions

Lade...
Vorschaubild
Dateien
Artinger_2-ri82jl5e530c6.pdf
Artinger_2-ri82jl5e530c6.pdfGröße: 3.51 MBDownloads: 214
Datum
2021
Herausgeber:innen
Kontakt
ISSN der Zeitschrift
Electronic ISSN
ISBN
Bibliografische Daten
Verlag
Schriftenreihe
Auflagebezeichnung
ArXiv-ID
Internationale Patentnummer
Link zur Lizenz
Angaben zur Forschungsförderung
Projekt
Open Access-Veröffentlichung
Open Access Gold
Sammlungen
Core Facility der Universität Konstanz
Gesperrt bis
Titel in einer weiteren Sprache
Publikationstyp
Zeitschriftenartikel
Publikationsstatus
Published
Erschienen in
International Journal of Molecular Sciences (IJMS). MDPI. 2021, 22(8), 4158. ISSN 1661-6596. eISSN 1422-0067. Available under: doi: 10.3390/ijms22084158
Zusammenfassung

Chemokines guide leukocyte migration in different contexts, including homeostasis, immune surveillance and immunity. The chemokines CCL19 and CCL21 control lymphocyte and dendritic cell migration and homing to lymphoid organs. Thereby they orchestrate adaptive immunity in a chemokine receptor CCR7-dependent manner. Likewise, cancer cells that upregulate CCR7 expression are attracted by these chemokines and metastasize to lymphoid organs. In-depth investigation of CCR7 expression and chemokine-mediated signaling is pivotal to understand their role in health and disease. Appropriate fluorescent probes to track these events are increasingly in demand. Here, we present an approach to cost-effectively produce and fluorescently label CCL19 and CCL21 in a semi-automated process. We established a versatile protocol for the production of recombinant chemokines harboring a small C-terminal S6-tag for efficient and site-specific enzymatic labelling with an inorganic fluorescent dye of choice. We demonstrate that the fluorescently labeled chemokines CCL19-S6Dy649P1 and CCL21-S6Dy649P1 retain their full biological function as assessed by their abilities to mobilize intracellular calcium, to recruit β-arrestin to engaged receptors and to attract CCR7-expressing leukocytes. Moreover, we show that CCL19-S6Dy649P1 serves as powerful reagent to monitor CCR7 internalization by time-lapse confocal video microscopy and to stain CCR7-positive primary human and mouse T cell sub-populations.

Zusammenfassung in einer weiteren Sprache
Fachgebiet (DDC)
570 Biowissenschaften, Biologie
Schlagwörter
fluorescent chemokine; CCL19; CCL21; CCR7; cell migration; leukocytes; flow cytometry; chemokine production
Konferenz
Rezension
undefined / . - undefined, undefined
Forschungsvorhaben
Organisationseinheiten
Zeitschriftenheft
Datensätze
Zitieren
ISO 690ARTINGER, Marc, Christoph MATTI, Oliver J. GERKEN, Christopher T. VELDKAMP, Daniel F. LEGLER, 2021. A Versatile Toolkit for Semi-Automated Production of Fluorescent Chemokines to Study CCR7 Expression and Functions. In: International Journal of Molecular Sciences (IJMS). MDPI. 2021, 22(8), 4158. ISSN 1661-6596. eISSN 1422-0067. Available under: doi: 10.3390/ijms22084158
BibTex
@article{Artinger2021-04-16Versa-54163,
  year={2021},
  doi={10.3390/ijms22084158},
  title={A Versatile Toolkit for Semi-Automated Production of Fluorescent Chemokines to Study CCR7 Expression and Functions},
  number={8},
  volume={22},
  issn={1661-6596},
  journal={International Journal of Molecular Sciences (IJMS)},
  author={Artinger, Marc and Matti, Christoph and Gerken, Oliver J. and Veldkamp, Christopher T. and Legler, Daniel F.},
  note={Article Number: 4158}
}
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/123456789/54163">
    <foaf:homepage rdf:resource="http://localhost:8080/"/>
    <dc:creator>Veldkamp, Christopher T.</dc:creator>
    <dc:creator>Gerken, Oliver J.</dc:creator>
    <dc:date rdf:datatype="http://www.w3.org/2001/XMLSchema#dateTime">2021-06-30T14:40:56Z</dc:date>
    <dc:contributor>Veldkamp, Christopher T.</dc:contributor>
    <dspace:hasBitstream rdf:resource="https://kops.uni-konstanz.de/bitstream/123456789/54163/1/Artinger_2-ri82jl5e530c6.pdf"/>
    <dc:contributor>Legler, Daniel F.</dc:contributor>
    <dcterms:rights rdf:resource="http://creativecommons.org/licenses/by/4.0/"/>
    <dc:creator>Artinger, Marc</dc:creator>
    <dc:contributor>Gerken, Oliver J.</dc:contributor>
    <dcterms:title>A Versatile Toolkit for Semi-Automated Production of Fluorescent Chemokines to Study CCR7 Expression and Functions</dcterms:title>
    <dcterms:hasPart rdf:resource="https://kops.uni-konstanz.de/bitstream/123456789/54163/1/Artinger_2-ri82jl5e530c6.pdf"/>
    <dcterms:abstract xml:lang="eng">Chemokines guide leukocyte migration in different contexts, including homeostasis, immune surveillance and immunity. The chemokines CCL19 and CCL21 control lymphocyte and dendritic cell migration and homing to lymphoid organs. Thereby they orchestrate adaptive immunity in a chemokine receptor CCR7-dependent manner. Likewise, cancer cells that upregulate CCR7 expression are attracted by these chemokines and metastasize to lymphoid organs. In-depth investigation of CCR7 expression and chemokine-mediated signaling is pivotal to understand their role in health and disease. Appropriate fluorescent probes to track these events are increasingly in demand. Here, we present an approach to cost-effectively produce and fluorescently label CCL19 and CCL21 in a semi-automated process. We established a versatile protocol for the production of recombinant chemokines harboring a small C-terminal S6-tag for efficient and site-specific enzymatic labelling with an inorganic fluorescent dye of choice. We demonstrate that the fluorescently labeled chemokines CCL19-S6&lt;sup&gt;Dy649P1&lt;/sup&gt; and CCL21-S6&lt;sup&gt;Dy649P1&lt;/sup&gt; retain their full biological function as assessed by their abilities to mobilize intracellular calcium, to recruit β-arrestin to engaged receptors and to attract CCR7-expressing leukocytes. Moreover, we show that CCL19-S6&lt;sup&gt;Dy649P1&lt;/sup&gt; serves as powerful reagent to monitor CCR7 internalization by time-lapse confocal video microscopy and to stain CCR7-positive primary human and mouse T cell sub-populations.</dcterms:abstract>
    <dc:contributor>Artinger, Marc</dc:contributor>
    <dc:creator>Legler, Daniel F.</dc:creator>
    <dc:creator>Matti, Christoph</dc:creator>
    <bibo:uri rdf:resource="https://kops.uni-konstanz.de/handle/123456789/54163"/>
    <dc:rights>Attribution 4.0 International</dc:rights>
    <dcterms:issued>2021-04-16</dcterms:issued>
    <dcterms:isPartOf rdf:resource="https://kops.uni-konstanz.de/server/rdf/resource/123456789/28"/>
    <dcterms:available rdf:datatype="http://www.w3.org/2001/XMLSchema#dateTime">2021-06-30T14:40:56Z</dcterms:available>
    <void:sparqlEndpoint rdf:resource="http://localhost/fuseki/dspace/sparql"/>
    <dspace:isPartOfCollection rdf:resource="https://kops.uni-konstanz.de/server/rdf/resource/123456789/28"/>
    <dc:language>eng</dc:language>
    <dc:contributor>Matti, Christoph</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