Antizyme, a natural ornithine decarboxylase inhibitor, induces apoptosis of haematopoietic cells through mitochondrial membrane depolarization and caspases' cascade

Antizymes delicately regulate ornithine decarboxylase (ODC) enzyme activity and polyamine transportation. One member of the family, antizyme-1, plays vital roles in molecular and cellular functions, including developmental regulation, cell cycle, proliferation, cell death, differentiation and tumorigenesis. However, the question of how does it participate in the cell apoptotic mechanism is still unsolved. To elucidate the contribution of human antizyme-1 in haematopoietic cell death, we examine whether inducible overexpression of antizyme enhances apoptotic cell death. Antizyme reduced the viability in a dose- and time-dependent manner of human leukemia HL-60 cells, acute T leukemia Jurkat cells and mouse macrophage RAW 264.7 cells. The apoptosis-inducing activities were determined by nuclear condensation, DNA fragmentation, sub-G(1) appearance, loss of mitochondrial membrane potential (Delta psi(m) ), release of mitochondrial cytochrome c into cytoplasm and proteolytic activation of caspase 9 and 3. Following conditional antizyme overexpression, all protein levels of cyclin-dependent kinases (Cdks) and cyclins are not significantly reduced, except cyclin D, before their entrance into apoptotic cell death. However, introduced cyclin D1 into Jurkat T tetracycline (Tet)-On cell system still couldn't rescue cells from apoptosis. Antizyme doesn't influence the expression of tumor suppressor p53 and its downstream p21, but it interferes in the expressions of Bcl-2 family. Inducible antizyme largely enters mitochondria resulting in cytochrome c release from mitochondria to cytosol following Bcl-xL decrease and Bax increase. According to these data, we suggest that antizyme induces apoptosis mainly through mitochondria-mediated and cell cycle-independent pathway. Furthermore, antizyme induces apoptosis not only by Bax accumulation reducing the function of the Bcl-2 family, destroying the Delta psi(m) , and releasing cytochrome c to cytoplasm but also by the activation of apoptosomal caspase cascade.