Individual epidermal growth factor receptor autophosphorylation sites do not stringently define association motifs for several SH2-containing proteins.

To determine whether individual autophosphorylation sites in the epidermal growth factor (EGF) receptor define specific interaction sites for the in vivo association of signal transduction proteins that contain src homology 2 (SH2) domains, the capacity of wild-type and mutant EGF receptors to associate with several SH2 domain-containing proteins has been assayed. Mutants included receptors with single autophosphorylation site mutations at each of five autophosphorylation sites and receptors in which multiple autophosphorylation sites were removed by point mutation or deletion of carboxyl-terminal residues. Receptor association, as measured by coimmunoprecipitation, has been determined for phospholipase C-gamma 1, the ras GTPase-activating protein, the p85 subunit of phosphatidylinositol 3-kinase, and the src homology and collagen protein. In contrast to data obtained with single autophosphorylation site mutants of other receptor tyrosine kinases, none of the EGF receptor single site mutants was dramatically impaired in its capacity to associate with any of these SH2-containing proteins. However, association was completely abrogated when all five autophosphorylation sites were mutated or removed by deletion. These results indicate that individual autophosphorylation sites in the EGF receptor are not stringently required for the recognition and association of different SH2-containing substrates. Thus, EGF receptor autophosphorylation sites seem to be flexible and/or compensatory in their capacity to mediate association with these four SH2-containing substrates.