DISABLED-2 (DAB2) IS AN SH3 DOMAIN-BINDING PARTNER OF GRB2

Disabled-2 (Dab2), a mammalian structural homolog of Drosophila Disabl ed (Dab), is a mitogen-responsive phosphoprotein, It has been speculat ed to be a negative regulator of growth since its expression is lost i n ovarian carcinomas, Dab2 contains a C-terminal proline-rich domain w ith sequences similar to those found in Sos, a guanine nucleotide exch ange factor for Ras, The proline-rich sequences of Sos mediate the int eraction of Sos with Grb2, an adaptor protein which coupled tyrosine k inase receptors to Sos, Herein, we have investigated the possibility t hat Dab2 interacts with Grb2, In experiments of co-immunoprecipitation from BAC1.2F5 macrophage cell lysates, significant quantities of Grb2 were associated with both Sos and Dab2, although Dab2 and Sos were no t present in the same complex, Transfection of Dab2 into a Dab2-negati ve cell line (293 cells) decreased the amount of Grb2 associated with Sos, suggesting that Dab2 competes with Sos for binding to Grb2, Proli ne-rich peptides corresponding to Dab2 (#661-669) and to Sos (#1146-11 61) inhibited the binding of Dab2 to Grb2, but were less effective in disrupting the Grb2-Sos complex, The expressed proline-rich domain of Dab2 (#600-730) bound Grb2, but other regions of Dab2 failed to bind G rb2. Both of the individual SH3 domains of Grb(2) bound to Sos (N-term inal SH3 domain > > C-terminal SH3 domain), but binding to Dab2 requir ed the intact Grb2, suggesting cooperative binding using both SH3 doma ins of Grb2, These data indicate that Dab2 binds to the SH3 domains of Grb2 via its C-terminal proline-rich sequences, Dab2 may modulate gro wth factor/Ras pathways by competing with Sos for binding to Grb2.