CONSTITUTIVE PHOSPHORYLATION OF SHC PROTEINS IN HUMAN TUMORS

The She gene encodes three overlapping proteins which all contain a ca rboxy-terminal SH2 domain. She proteins are ubiquitously expressed and are downstream targets and effecters of activated tyrosine kinases (T K). We investigated tyrosine-phosphorylation of She proteins in normal and transformed cells. In tumor cells with known TK gene alterations She proteins were constitutively phosphorylated and complexed with the activated TK. No constitutive She phosphorylation was found in primar y cell cultures and normal tissues, In 14 of 27 tumor cell lines with no reported TK alterations, She proteins were constitutively phosphory lated and formed stable complexes with novel tyrosine-phosphorylated p olypeptides. Ten distinct She-associated phosphoproteins were identifi ed with molecular weights ranging from 30 to 200 kDa. In a subset of c arcinoma cell lines, phosphorylated She proteins complexed with a p175 phosphoprotein that was identified as the constitutively activated EG FR. In one glioblastoma cell line, a Shc-associated p190 was identifie d as the activated PDGFR. In 13 of 14 acute leukemia samples phosphory lated She proteins were constitutively complexed with a p140 phosphopr otein. Some of the She-associated phosphoproteins (EGFR, PDGFR, erbB-2 , Met, bcr-abl, H4-ret) bound both the Shc- and Grb2-SH2 domains in vi tro; others (p175; p70-p80) only the Shc-SH2 domain and yet others (p1 40) only the Grb2-SH3 domains. These results indicate that She protein s are common substrates of constitutively activated TKs and that the a nalysis of She phosphorylation allow the identification of tumors with constitutive TK activation.