Figure 3.

Metalloprotease inhibitors prevent generation of both Met-CTF and Met-ICD. (A) MDCK cells were left untreated (−) or were treated overnight with 1 μM E compound and/or 25 μM GM6001 and 50 μM TAPI-1. The following day the cells were treated for 5 h with 10 μM lactacystin. (B) MEF PS1,2 −/− were left untreated or treated overnight with 1 μM E compound and/or 25 μM GM6001. The following day the cells were treated for 5 h with 10 μM lactacystin. (C) MDCK cells transiently transfected with either the empty vector or a vector expressing human Met (h Met) were treated or not overnight with 25 μM GM6001. The following day, the cells were treated for 45 min with 100 ng/ml PMA. The culture medium was collected and immunoprecipitation was performed with an antibody directed against the extracellular domain of human Met. (D) HeLa cells were treated or not overnight with 1 μM E compound and/or 25 μM GM6001. The following day, the cells were treated for 5 h with 10 μM lactacystin and for 45 min with 100 ng/ml PMA. The culture medium was collected, and immunoprecipitation was performed with an antibody directed against the extracellular domain of human Met. (A–D) For each condition, the same amount of protein was resolved by 10% SDS-PAGE or 7% SDS-PAGE for IP and analyzed by Western blotting with different anti-Met antibodies as indicated. Arrows indicate positions of Met-CTF, Met-ICD, and Met-NTF.