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An immunological approach reveals biological differences between the two NDF/heregulin receptors, ErbB-3 and ErbB-4
X Chen1, G Levkowitz, E Tzahar
1Department of Chemical Immunology, Weizmann Institute of Science, Rehovot, Israel.
Abstract:
The group of subtype I transmembrane tyrosine kinases includes the epidermal growth factor (EGF) receptor (ErbB-1), an orphan receptor (ErbB-2), and two receptors for the Neu differentiation factor (NDF/heregulin), namely: ErbB-3 and ErbB-4. Here we addressed the distinct functions of the two NDF receptors by using an immunological approach. Two sets of monoclonal antibodies (mAbs) to ErbB-3 and ErbB-4 were generated through immunization with recombinant ectodomains of the corresponding receptors that were fused to immunoglobulin. We found that the shared ligand binds to highly immunogenic, but immunologically distinct sites of ErbB-3 and ErbB-4. NDF receptors differed also in their kinase activities; whereas the catalytic activity of ErbB-4 was activable by mAbs, ErbB-3 underwent no activation by mAbs in living cells. Likewise, down-regulation of ErbB-4, but not ErbB-3, was induced by certain mAbs. By using the generated mAbs, we found that the major NDF receptor on mammary epithelial cells is a heterodimer of ErbB-3 with ErbB-2, whereas an ErbB-1/ErbB-2 heterodimer, or an ErbB-1 homodimer, is the predominant species that binds EGF. Consistent with ErbB-2 being a shared receptor subunit, its tyrosine phosphorylation was increased by both heterologous ligands and it mediated a trans-inhibitory effect of NDF on EGF binding. Last, we show that the effect of NDF on differentiation of breast tumor cells can be mimicked by anti-ErbB-4 antibodies, but not by mAbs to ErbB-3. Nevertheless, an ErbB-3-specific mAb partially inhibited the effect of NDF on cellular differentiation. These results suggest that homodimers of ErbB-4 are biologically active, but heterodimerization of the kinase-defective ErbB-3, probably with ErbB-2, is essential for transmission of NDF signals through ErbB-3.
Insights
Monoclonal antibodies reveal distinct functions of Neu differentiation factor (NDF) receptors ErbB-3 and ErbB-4. ErbB-4 homodimers are active, while ErbB-3 requires heterodimerization with ErbB-2 for NDF signal transmission.
Area of Science:
- Cell biology
- Molecular biology
- Immunology
Background:
- Subtype I transmembrane tyrosine kinases, including epidermal growth factor (EGF) receptor (ErbB-1), ErbB-2, ErbB-3, and ErbB-4, play critical roles in cellular signaling.
- ErbB-3 and ErbB-4 are receptors for Neu differentiation factor (NDF/heregulin), but their distinct functions are not fully understood.
Purpose of the Study:
- To investigate the distinct functions of ErbB-3 and ErbB-4, the two Neu differentiation factor (NDF) receptors.
- To characterize the roles of ErbB-3 and ErbB-4 in mammary epithelial cells and breast tumor cell differentiation.
Main Methods:
- Generation of monoclonal antibodies (mAbs) against recombinant ErbB-3 and ErbB-4 ectodomains.
- Immunological characterization of ligand binding sites and receptor activation.
- Analysis of receptor dimerization and signaling pathways in living cells.
- Assessment of NDF-induced breast tumor cell differentiation.
Main Results:
- NDF binds to distinct epitopes on ErbB-3 and ErbB-4.
- ErbB-4 kinase activity is activable by mAbs, while ErbB-3 is not; certain mAbs induce ErbB-4 down-regulation.
- ErbB-3/ErbB-2 heterodimers are the major NDF receptors on mammary epithelial cells.
- ErbB-2 phosphorylation is increased by heterologous ligands, mediating trans-inhibition of EGF binding.
- Anti-ErbB-4 antibodies mimic NDF effects on breast tumor cell differentiation, while anti-ErbB-3 partially inhibits them.
Conclusions:
- ErbB-4 homodimers are biologically active and can mediate NDF signaling.
- Heterodimerization of kinase-defective ErbB-3, likely with ErbB-2, is essential for NDF signal transmission.
- Distinct functions of ErbB-3 and ErbB-4 highlight their unique roles in NDF-mediated cellular processes.