title: Cardiotrophin-like cytokine labelling using Bir A biotin ligase: a sensitive tool to study receptor expression by immune and non-immune cells.
abstract: The recently identified IL-6 family member cardiotrophin-like cytokine (also named novel neurotrophin-1 or B cell stimulating factor-3) forms a secreted complex with cytokine-like factor-1 which binds and activates the tripartite ciliary neurotrophic factor receptor. The striking differences between the phenotype of mice in which either the ciliary neurotrophic factor or its receptor are inactivated suggest that the cardiotrophin-like cytokine/cytokine-like factor-1 complex could be the developmentally important ciliary neurotrophic factor receptor ligand. Cardiotrophin-like cytokine is also produced in the immune system and has been reported to activate B cells in vivo and in vitro. B cells do not express the ciliary neurotrophic factor receptor suggesting the existence of an alternative receptor. We produced the cardiotrophin-like cytokine/cytokine-like factor-1 complex tagged with a Bir A biotin ligase AviTag peptide substrate. This cytokine could be efficiently biotinylated in vitro with Bir A. It was subsequently validated as a sensitive tool for ciliary neurotrophic factor receptor detection by flow cytometry and for magnetic-activated cell sorting. It was also shown to allow the detection of a specific receptor by activated B cells. Whereas binding to cells expressing the ciliary neurotrophic factor receptor could be prevented by competition with ciliary neurotrophic factor, binding to B cells was not. The biotinylated cardiotrophin-like cytokine/cytokine-like factor-1 complex therefore represents a new reagent to study ciliary neurotrophic factor and cardiotrophin-like cytokine receptor expression and for the identification of the putative cardiotrophin-like cytokine B cell receptor. It further validates the use of biotin ligase catalysed biotinylation for the detection of cytokine receptors.
authors.affiliation: Département de pharmacologie, Université de Montréal, C.P 6128, succursale Centre-ville, Montreal, QC, Canada H3C 3J7.
authors.forename: Isabelle
authors.initials: I
authors.lastname: Cognet
authors.forename: Florence
authors.initials: F
authors.lastname: Guilhot
authors.forename: Mélanie
authors.initials: M
authors.lastname: Gabriac
authors.forename: Sylvie
authors.initials: S
authors.lastname: Chevalier
authors.forename: Younes
authors.initials: Y
authors.lastname: Chouikh
authors.forename: Alexandra
authors.initials: A
authors.lastname: Herman-Bert
authors.forename: Angélique
authors.initials: A
authors.lastname: Guay-Giroux
authors.forename: Sophie
authors.initials: S
authors.lastname: Corneau
authors.forename: Giovanni
authors.initials: G
authors.lastname: Magistrelli
authors.forename: Greg C
authors.initials: GC
authors.lastname: Elson
authors.forename: Hugues
authors.initials: H
authors.lastname: Gascan
authors.forename: Jean-François
authors.initials: JF
authors.lastname: Gauchat
mesh_terms: D000818:Animals; D001360:Avidin / Q000378:metabolism; D001402:B-Lymphocytes / Q000187:drug effects / Q000378:metabolism; D019731:Carbon-Nitrogen Ligases / Q000737:chemistry / Q000378:metabolism*; D002477:Cells / Q000187:drug effects / Q000276:immunology* / Q000378:metabolism*; D002478:Cells, Cultured; D016207:Cytokines / Q000737:chemistry / Q000378:metabolism*; D000939:Epitopes / Q000276:immunology; D004926:Escherichia coli; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000378:metabolism*; D005260:Female; D005786:Gene Expression Regulation; D006801:Humans; D051379:Mice; D011485:Protein Binding; D020801:Receptor, Ciliary Neurotrophic Factor / Q000378:metabolism; D018121:Receptors, Cytokine / Q000276:immunology / Q000378:metabolism*; D012097:Repressor Proteins / Q000737:chemistry / Q000378:metabolism*; D013154:Spleen / Q000187:drug effects / Q000378:metabolism; D014157:Transcription Factors / Q000737:chemistry / Q000378:metabolism*
chemical_list: D016207:Cytokines; D000939:Epitopes; D029968:Escherichia coli Proteins; D020801:Receptor, Ciliary Neurotrophic Factor; D018121:Receptors, Cytokine; D012097:Repressor Proteins; D014157:Transcription Factors; C121169:cardiotrophin-like cytokine; C113554:cytokine-like factor-1; D001360:Avidin; D019731:Carbon-Nitrogen Ligases; C058125:birA protein, E coli