INVOLVEMENT OF PP60(C-SRC) WITH 2 MAJOR SIGNALING PATHWAYS IN HUMAN BREAST-CANCER

The phosphotyrosine residues of receptor tyrosine kinases serve as uni que binding sites for proteins involved in intracellular signaling, wh ich contain SRC homology 2 (SH2) domains. Since overexpression or acti vation of the pp60c-src kinase has been reported in a number of human tumors, including primary human breast carcinomas, we examined the int eractions of the SH2 and SH3 domains of human SRC with target proteins in human carcinoma cell lines. Glutathione S-transferase fusion prote ins containing either the SH2, SH3, or the entire SH3/SH2 region of hu man SRC were used to affinity purify tyrosine-phosphorylated proteins from human breast carcinoma cell lines. We show here that in human bre ast carcinoma cell lines, the SRC SH2 domain binds to activated epider mal growth factor receptor (EGFR) and p185HER2/neu. SRC SH2 binding to EGFR was also observed in a nontumorigenic cell line after hormone st imulation. Endogenous pp60c-src was found to tightly associate with ty rosine-phosphorylated EGFR. Association of the SRC SH2 with the EGFR w as blocked by tyrosyl phosphopeptides containing the sequences surroun ding tyrosine-530, the regulatory site in the SRC C terminus, or seque nces surrounding the major sites of autophosphorylation in the EGFR. T hese results raise the possibility that association of pp60c-src with these receptor tyrosine kinases is an integral part of the signaling e vents mediated by these receptors and may contribute to malignant tran sformation.