FIG. 1.

Stress stimuli generate a p40 MET fragment in primary cultures of mouse hepatocytes and cortical neurons. (A) Hepatocytes were cultured for 3 days and treated for 12 or 18 h with actinomycin D or actinomycin D plus TNF-α (100 ng/ml) or left untreated (−). For each condition, the same amount of protein was resolved by SDS-10% PAGE and analyzed by Western blotting using antibodies directed against the C-terminal tail of mouse MET. The filter was stripped and reprobed using an anti-Erk2 antibody to assess comparable loading. (B) Percentages of TUNEL-positive hepatocytes over total number of hepatocytes were determined (n = 3; plus standard deviation [SD]). Protein was resolved by SDS-10% PAGE and analyzed by Western blotting using anti-cleaved caspase 3 antibody. The filter was stripped and reprobed using an anti-α-tubulin antibody to assess comparable loading. (C) Cortical neurons were cultured for 3 days and treated for 12 h with camptothecin (Campto.; 5 μM) or left untreated (−). For each condition, the same amount of protein was resolved by SDS-10% PAGE and analyzed by Western blotting using antibodies directed against the C-terminal tail of mouse MET. The filter was stripped and reprobed using an anti-Erk2 antibody to assess comparable loading. (D) Percentages of TUNEL-positive cells over total number of cells were determined (n = 3; plus SD). Protein was resolved by SDS-10% PAGE and analyzed by Western blotting using anti-cleaved caspase 3 antibody. The filter was stripped and reprobed using an anti-α-tubulin antibody to assess comparable loading. The arrowheads indicate the positions of the MET precursor p170, MET β subunit p140, and MET p40 fragment. Although expression of p140 MET is higher than that of p170 MET, it appears that degradation of full-length MET is more pronounced for the p170 MET precursor than for the fully glycosylated p140 MET β subunit, suggesting that mature glycosylated receptors are more resistant to degradation.