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Fig. 3. Constitutive high level phosphorylation of p120 at Y228 in carcinoma cell lines. (A) Variable levels of p120 phosphorylation at Y228. p120 was immunoprecipitated from the indicated human carcinoma cell lines and western blotted with mAb pY228 (pY228) or mAb pp120 (p120). E-cadherin levels were monitored by western blotting of whole cell lysates with mAb HECD-1. (B) Cadherin-dependent p120 phosphorylation. SkBr3 and MDA-MB-453 cells, which lack E-cadherin expression, were infected with control or E-cadherin containing retrovirus. Polyclonal cell lines were generated by selection in G418. Immunoprecipitated p120 was detected by western blotting with mAb pY228 (pY228) or mAb pp120 (p120). E-cadherin was detected in whole cell lysates by western blotting with mAb HECD-1 (E-cadherin). All cells were washed with PBS containing pervanadate for 2 minutes prior to lysis with pervanadate-containing RIPA buffer.
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