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.
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Research Article|
August 01 1998
ErbB3 (HER3) interaction with the p85 regulatory subunit of phosphoinositide 3-kinase
Nathan J. HELLYER;
Nathan J. HELLYER
1Department of Pharmacology, University of Iowa College of Medicine, Iowa City, IA 52242, U.S.A.
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Kunrong CHENG;
Kunrong CHENG
1
1Department of Pharmacology, University of Iowa College of Medicine, Iowa City, IA 52242, U.S.A.
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John G. KOLAND
John G. KOLAND
2
1Department of Pharmacology, University of Iowa College of Medicine, Iowa City, IA 52242, U.S.A.
2To whom correspondence should be addressed (e-mail john-koland@uiowa.edu).
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Publisher: Portland Press Ltd
Received:
February 27 1998
Revision Received:
May 11 1998
Accepted:
May 19 1998
Online ISSN: 1470-8728
Print ISSN: 0264-6021
The Biochemical Society, London © 1998
1998
Biochem J (1998) 333 (3): 757–763.
Article history
Received:
February 27 1998
Revision Received:
May 11 1998
Accepted:
May 19 1998
Citation
Nathan J. HELLYER, Kunrong CHENG, John G. KOLAND; ErbB3 (HER3) interaction with the p85 regulatory subunit of phosphoinositide 3-kinase. Biochem J 1 August 1998; 333 (3): 757–763. doi: https://doi.org/10.1042/bj3330757
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