Figure 3.

In vitro cleavage of plakoglobin by recombinant caspases. (a) Lysates from control HUVEC were incubated with recombinant CPP32 and Mch2 for 45 min. The reaction products, as well as lysates from control (C) and apoptotic (A) cells, were subjected to 7.5% SDS-PAGE, followed by Western blotting with a monoclonal antibody to plakoglobin (^553–738 C). Mch2 generates a cleavage fragment I, which comigrates with the highest molecular weight cleavage product in apoptotic cells, whereas CPP32 cleavage results in all of the fragments observed in apoptotic cells. (b) Plakoglobin was immunoprecipitated from 250 μg lysate of control HUVEC and incubated with recombinant CPP32, Mch2, or reaction buffer (buffer), with lysates from control (C) or apoptotic (A) cells included in the analysis for comparison. CPP32 cleavage of immunoprecipitated plakoglobin results in fragment I observed in apoptotic cells, whereas Mch2 has no effect.