Beta-tubulin binds Src homology 2 domains through a region different from the tyrosine-phosphorylated protein-recognizing site.

Src homology 2 (SH2) domains have been demonstrated to bind tyrosine-phosphorylated proteins that participate in signaling by growth factors and oncogenes by recognizing amino acid sequences containing phosphotyrosine residue. We found that SH2 domains such as Ash/Grb2, the 85-kDa subunit of phosphatidylinositol 3-kinase, and phospholipase Cgamma1 also bind beta-tubulin through a different region that recognizes phosphotyrosine in vitro and in vivo. Furthermore, binding occurs even when the SH2 domain is occupied by tyrosine-phosphorylated epidermal growth factor receptors. Using deleted constructs of Ash/Grb2 SH2, we found that carboxyl-terminal beta strands E and F, and alpha helix B (region "c") are required for binding. A synthetic peptide (FLWVVKFNSLNELVDYH) composed of region c inhibited the binding of beta-tubulin to the SH2 domains of Ash/Grb2, phosphatidylinositol 3-kinase, and phospholipase Cgamma1. The co-localization of SH2 proteins and microtubules is also confirmed by immunostaining. These data suggest that microtubules play important roles in the assembly of signaling molecules complexes containing SH2 proteins.