Diphtheria toxin receptor-binding domain substitution with interleukin 6: genetic construction and interleukin 6 receptor-specific action of a diphtheria toxin-related interleukin 6 fusion protein.
We have genetically replaced that portion of the diphtheria toxin structural gene which encodes the native receptor-binding domain with a synthetic gene encoding the cytokine interleukin 6 (IL-6/IFN-beta 2/BSF-2). The resulting gene fusion encodes the chimeric toxin DAB389-IL-6. Following expression and purification, we demonstrate that DAB389-IL-6 is selectively cytotoxic for eukaryotic cells bearing the interleukin 6 receptor. In addition, the cytotoxic action of DAB389-IL-6 is shown to require binding to the IL-6 receptor, internalization by receptor-mediated endocytosis and passage through an acidic compartment. Following the delivery of the catalytically active fragment A to the cytosol of target cells, cellular protein synthesis is inhibited by the ADP-ribosylation of elongation factor 2. While eukaryotic cells which are devoid of the IL-6 receptor are uniformly resistant to the action of this fusion toxin, the data presented suggest that a minimal number of IL-6 receptors may be necessary to mediate the internalization of sufficient levels of DAB389-IL-6 to result in the intoxication of target cells.
Jean
L F
LF
Evans Department of Clinical Research, University Hospital, Boston University School of Medicine, MA 02118.
Murphy
J R
JR
D000818:Animals; D001402:B-Lymphocytes / Q000187:drug effects; D001483:Base Sequence; D001665:Binding Sites; D002460:Cell Line; D004167:Diphtheria Toxin / Q000235:genetics* / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D005057:Eukaryotic Cells / Q000187:drug effects*; D015964:Gene Expression Regulation, Bacterial; D000922:Immunotoxins / Q000235:genetics / Q000378:metabolism; D015850:Interleukin-6 / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D011971:Receptors, Immunologic / Q000378:metabolism*; D019947:Receptors, Interleukin-6; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology
D004167:Diphtheria Toxin; D000922:Immunotoxins; D015850:Interleukin-6; D011971:Receptors, Immunologic; D019947:Receptors, Interleukin-6; D011993:Recombinant Fusion Proteins