Identification of Src phosphorylation sites in the catenin p120(ctn)

p120-catenin (p120(ctn)) interacts with the cytoplasmic tail of cadherins a nd is thought to regulate cadherin clustering during formation of adherens junctions. Several observations suggest that p120 can both positively and n egatively regulate cadherin adhesiveness depending on signals that so far r emain unidentified. Although p120 tyrosine phosphorylation is a leading can didate, the role of this modification in normal and Src-transformed cells r emains unknown. Here, as a first step toward pinpointing this role, we have employed two-dimensional tryptic mapping to directly identify the major si tes of Src-induced p120 phosphorylation. Eight sites were identified by dir ect mutation of candidate tyrosines to phenylalanine and elimination of the accompanying spots on the two-dimensional maps. Identical sites were obser ved in vitro and in vivo, strongly suggesting that the physiologically impo rtant sites have been correctly identified. Changing all of these sites to phenylalanine resulted in a p120 mutant, p120-8F, that could not be efficie ntly phosphorylated by Src and failed to interact with SHP-1, a tyrosine ph osphatase shown previously to interact selectively with tyrosine-phosphoryl ated p120 in cells stimulated with epidermal growth factor. Using selected tyrosine to phenylalanine p120 mutants as dominant negative reagents, it ma y now be possible to selectively block events postulated to be dependent on p120 tyrosine phosphorylation.