ErbB3 (HER3) interaction with the p85 regulatory subunit of phosphoinositide 3-kinase.
ErbB3 (HER3), a unique member of the ErbB receptor family, lacks intrinsic protein tyrosine kinase activity and contains six Tyr-Xaa-Xaa-Met (YXXM) consensus binding sites for the SH2 domains of the p85 regulatory subunit of phosphoinositide 3-kinase. ErbB3 also has a proline-rich sequence that forms a consensus binding site for the SH3 domain of p85. Here we have investigated the interacting domains of ErbB3 and p85 by a unique application of the yeast two-hybrid system. A chimaeric ErbB3 molecule containing the epidermal growth factor receptor protein tyrosine kinase domain was developed so that the C-terminal domain of ErbB3 could become phosphorylated in the yeast system. We also generated several ErbB3 deletion and Tyr-->Phe site-specific mutants, and observed that a single ErbB3 YXXM motif was necessary and sufficient for the association of ErbB3 with p85. The incorporation of multiple YXXM motifs into the ErbB3 C-terminus enabled a stronger ErbB3/p85 interaction. The proline-rich region of ErbB3 was not necessary for interaction with p85. However, either deletion or mutation of the p85 SH3 domain decreased the observed ErbB3/p85 association. Additionally an ErbB3/p85 SH3 domain interaction was detected by an assay in vitro. These results were consistent with a model in which pairs of phosphorylated ErbB3 YXXM motifs co-operate in binding to the tandem SH2 domains of p85. Although a contributing role for the p85 SH3 domain was suggested, the N- and C-terminal SH2 domains seemed to be primarily responsible for the high-affinity association of p85 and ErbB3.
Department of Pharmacology, University of Iowa College of Medicine, Iowa City, IA 52242, USA.
N J
NJ
Hellyer
K
K
Cheng
J G
JG
Koland
D002678:Chimera; D066246:ErbB Receptors / Q000737:chemistry / Q000378:metabolism*; D046911:Macromolecular Substances; D010449:Peptide Mapping; D019869:Phosphatidylinositol 3-Kinases / Q000737:chemistry / Q000378:metabolism*; D011392:Proline / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000737:chemistry / Q000378:metabolism*; D020893:Receptor, ErbB-3; D015003:Yeasts / Q000235:genetics / Q000378:metabolism; D018909:src Homology Domains
D046911:Macromolecular Substances; D011518:Proto-Oncogene Proteins; D011392:Proline; D019869:Phosphatidylinositol 3-Kinases; D066246:ErbB Receptors; D020893:Receptor, ErbB-3