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pubmed:abstractText |
Bax is cleaved by calpain at aspartate 33 (Asp33) to yield p18 Bax during stress-induced apoptosis. To assess the role of p18 Bax in apoptosis, an ecdysone-inducible expression system was generated. Similar levels of wild-type (WT) and noncleavable Asp33Ala (Asp-->Ala) Bax are induced in 293 cells while expression of N-terminal-deleted p18 (Delta1-33) Bax remains low (20% of full-length p21 Bax) due to a reduced half-life (2 hours versus 12 hours for p21 Bax) resulting from increased sensitivity to cathepsin-like proteolytic degradation. Expression of p18 Bax is enhanced to levels comparable to p21 Bax when induction is carried out in the presence of cathepsin inhibitors, Z-Phe-Gly-NHO-Bz or N-Acetyl-Leu-Leu-Met-CHO. Compared with WT Bax, expression of similar levels of p18 Bax and, surprisingly, Asp33Ala Bax more potently induces apoptosis as indicated by increased cytochrome c release, caspase-9/-3 activation, and DNA fragmentation, potentially due to their increased homo-oligomerization in mitochondrial membranes. Studies in A-549, U-937, K-562, and HL-60 cells confirm that inhibition of Bax cleavage results in 25% to 35% reduction of drug-induced apoptosis, while inhibition of p18 Bax degradation enhances apoptosis by 25% to 40%. Results indicate that although cleavage to p18 Bax is not required for Bax to initiate apoptosis, p18 Bax potently accelerates the apoptotic process.
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