Use of Bid-ribozymes to characterize and Bid-affiliated apoptotic pathway.
Multiple cell and stimulus specific apoptosis pathways have been identified (1) revealing a complexity of molecular events involved. Tumor necrosis factor (TNF alpha) is known to activate mitochondria dependent or independent pathways for cell death. These pathways converge to activate effector caspases, caspases 3, 6 and 7 (2). The intermediate events including the substrates and effectors however are not clearly understood. In this report, we have employed ribozyme technology to elucidate some of these intermediate events. We show here that Bid, a pro-apoptotic facilitator that plays an important role in the mitochondrial pathway, can be targeted by Bid-specific ribozymes. MCF7 breast carcinoma cells stably transfected with expression plasmids encoding active (but not inactive or mock) Bid ribozymes showed delayed response to TNF alpha. This was accompanied by decreased activation of caspases 7 and 9, but not of caspase 8. The data assign caspase 9 as an upstream activator of caspase 7 in TNF alpha-induced Bid-mediated apoptosis.
Onuki
R
R
Gene Discovery Research Center, National Institute of Advanced Industrial Science and Technology (AIST), Institute of Applied Biochemistry, University of Tsukuba, Tsukuba Science City, Japan.
Kawasaki
H
H
Baba
T
T
Taira
K
K
D017209:Apoptosis; D051023:BH3 Interacting Domain Death Agonist Protein; D002352:Carrier Proteins / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D053179:Caspase 7; D053453:Caspase 9; D020169:Caspases / Q000378:metabolism; D006801:Humans; D008954:Models, Biological; D016337:RNA, Catalytic / Q000378:metabolism*; D015398:Signal Transduction; D014407:Tumor Cells, Cultured; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D051023:BH3 Interacting Domain Death Agonist Protein; C494814:BID protein, human; D002352:Carrier Proteins; D016337:RNA, Catalytic; D014409:Tumor Necrosis Factor-alpha; C505520:CASP7 protein, human; C506433:CASP9 protein, human; D053179:Caspase 7; D053453:Caspase 9; D020169:Caspases