Controlling Homo- & Heterodimerization of ErbB Receptors Using Synthetic Ligands & Understanding the RTK Heterodimer Signaling Specificity in Breast Cancer.
Annual rept. 1 Aug 97-31 Jul 98
ARIAD INST FOR BIOMEDICAL RESEARCH CAMBRIDGE MA
Pagination or Media Count:
The four members of the ErbB family of receptor tyrosine kinases are involved in a complex array of combinatorial interactions involving homo and heterodimers. Since most cell types express more than one member of the ErbB family, it is difficult to distinguish the biological activities of different homo and heterodimers. Here we describe a method for including homo or heterodimerization of ErbB receptors using synthetic ligands without interference from the endogenous receptors, ErbB receptor chimeras containing synthetic ligand binding domains FKBPor FRB were homodimerized using the bivalent FKBP ligand, APl5lO, and heterodimerized using the bifunctional FKBP-FRB ligand, rapamycin. APl5lO treatment induced tyrosine phosphorylation of ErbBl and ErbB2 homodimers and recruitment of SH2 domain containing proteins She and Grb2. In addition, ErbBl and ErbB2 homodimers activated downstream signaling pathways leading to Erk2 and Akt phosphorylation. However, only ErbB1 homodimers were internalized upon APl5lO stimulation and only ErbBl homodimers were able to associate and induce phosphorylation of c-Cbl. Cells expressing AP1510-induced ErbBl homodimers were able to form foci, however cells expressing ErbB2 homodimers displayed 5-7 fold higher focus forming ability. Using rapamycin-inducible heterodimerization we show that c-Cbl is unable to associate with ErbBl in a ErbB1ErbB2 heterodimer most likely because ErbB2 is unable to phosphorylate the c-Cbl binding site on ErbBl. Thus, we demonstrate that ErbB 1 and ErbB2 homodimers differ in their ability to transform fibroblasts and provide evidence for differential signaling by ErbB homodimers and heterodimers. These observations also validate the use of synthetic ligands to study signaling and biological specificity of selected ErbB dimers in any cell type.
- Medicine and Medical Research