Diphtheria Toxin Binds to the Epidermal Growth Factor (EGF)-like Domain of Human Heparin-binding EGF-like Growth Factor/Diphtheria Toxin Receptor and Inhibits Specifically Its Mitogenic Activity

Abstract

The membrane anchored form of human heparin-binding epidermal growth factor-like growth factor (HB-EGF) acts as the diphtheria toxin (DT) receptor. Transfection of human HB-EGF cDNA into mouse LC cells, L cells stably expressing DRAP27, conferred sensitivity to DT, but transfection of mouse HB-EGF cDNA did not. To define the essential regions of HB-EGF that serve as the functional DT receptor, we examined the sensitivity to DT and DT binding of cells expressing several human/mouse HB-EGF chimeras. It was found that DT binds to the EGF-like domain of the human HB-EGF. However, mouse HB-EGF does not serve as a functional DT receptor due to non-conserved amino acid substitutions in this domain. In addition, CRM197, a non-toxic mutant of DT, inhibited strongly the mitogenic activity of the secreted form of human HB-EGF, but not of mouse HB-EGF and other EGF receptor-binding growth factors. These results confirmed further that DT interacts with the EGF-like domain of HB-EGF and that this interaction is specific for human HB-EGF.

Transcutaneous Immunization with Cross-Reacting Material CRM 197 of Diphtheria Toxin Boosts Functional Antibody Levels in Mice Primed Parenterally with Adsorbed Diphtheria Toxoid Vaccine

“…Despite the fact that mice lack the required receptors for DTx binding, which significantly reduces the immunogenicity of CRM 197 in this model (13,23), we have shown that topical application of CRM 197 , without additional stabilization or formulation, is highly immunogenic for boosting diphtheria immunity in mice after parenteral priming with a conventional toxoid vaccine. Although initial neutralizing antibody responses were significantly enhanced in the presence of a mucosal adjuvant (LTR72), longer-term immunity induced by TCI using CRM 197 alone was comparable to immunity induced by TCI with adjuvant and to boosting with a conventional toxoid vaccine given by the classical parenteral route.…”

Suppression of the Biological Activities of the Epidermal Growth Factor (EGF)-like Domain by the Heparin-binding Domain of Heparin-binding EGF-like Growth Factor

“…This concentration (1.7 nM) is close to the concentration required for half-saturation of DT binding to human pro-HB-EGF (18,37). Nonspecific binding of 125 I-DT was assessed in the presence of 10 g/ml unlabeled DT.…”

“…DT Binding Assay-HB-EGF has been shown to act as a diphtheria toxin receptor (DTR) and binds to DT through its EGF-like domain (37). Previously, we reported that pro-HB-EGF associates with cell surface HSPGs, which increases its binding affinity for DT (32).…”

Mitogenic effects of gastrin-releasing peptide in head and neck squamous cancer cells are mediated by activation of the epidermal growth factor receptor

“…Extracellular processing of the EGFR ligand, proheparin-binding epidermal growth factor (pro-HB-EGF) has been primarily implicated in EGFR activation by GPCRs (Prenzel et al, 1999). A mutated form of diphtheria toxin, CRM197, has been shown to bind specifically to HB-EGF and inhibit its mitogenic activity (Mitamura et al, 1995). Prenzel et al reported that CRM197 pretreatment could completely inhibit EGFR activation induced by GPCRs in COS-7 cells (Prenzel et al, 1999).…”