COORDINATE ACTIVATION OF C-SRC BY SH3-BINDING AND SH2-BINDING SITES ON A NOVEL, P130(CAS)-RELATED PROTEIN, SIN

To understand how protein-protein interactions mediated by the Src-SH3 domain affect c-Src signaling, we screened for proteins that interact with the Src-SH3. We found a novel protein, Sin (Src interacting or s ignal integrating protein), that binds to Src-SH3 with high affinity, contains numerous tyrosine residues in configurations suggestive of SH 2-binding sites, and is related to the v-Src substrate p130(Cas). In c otransfection assays, a small fragment of Sin retaining the Src-SH3-bi nding site and one tyrosine-containing motif induced c-Src activation as measured by a transcriptional reporter. Phosphorylation of the pept ide on tyrosine by c-Src, as a consequence of Src-SH3 binding, was nec essary for its stable interaction with c-Src in vivo and for transcrip tional activation. Phosphorylation of multiple tyrosine-containing mot ifs found on Sin correlated with c-Crk and cellular phosphoprotein bin ding to Sin as well as increased c-Src activity. These data suggest th at (1) SH2 and SH3 ligand sites on Sin cooperatively activate the sign aling potential of c-Src, (2) Sin acts as both an activator and a subs trate for c-Src, and (3) phosphorylated Sin may serve as a signaling e ffector molecule for Src by binding to multiple cellular proteins.