PROTEOLYTIC CLEAVAGE AND ACTIVATION OF PAK2 DURING UV IRRADIATION-INDUCED APOPTOSIS IN A431 CELLS

Exposure of mammalian cells to ultraviolet (UV) light elicits a cellul ar response and can also lead to apoptotic cell death. In this report, we show that a 36-kDa myelin basic protein (MBP) kinase detected by a n in-gel kinase assay can be dramatically activated during the early s tages of UV irradiation-triggered apoptosis of A431 cells. Immunoblot analysis revealed that this 36-kDa MBP kinase could be recognized by a n antibody against the C-terminal regions of a family of p(21Cdc42/Rac )-activated kinases (PAKs). By using this antibody and a PAK2-specific antibody against the N-terminal region of PAK2 as studying tools, we further demonstrated that UV irradiation caused cleavage of PAK2 to ge nerate a 36-kDa C-terminal catalytic fragment and a 30-kDa N-terminal fragment in A431 cells. The appearance of the 36-kDa C-terminal cataly tic fragment of PAK2 matched exactly with the activation of the 36-kDa MBP kinase in A431 cells upon UV irradiation. In addition, UV irradia tion also led to activation of CPP32/caspase-3, but not ICH-1L/caspase -2 and ICE/caspase-1, in A431 cells and the kinetics of activation of CPP32/caspase-3 appeared to correlate well with that of DNA fragmentat ion and of cleavage/activation of PAK2, respectively. Moreover, blocka ge of activation of CPP32/caspase-3 by pretreating the cells with two specific tetrapeptidic inhibitors for caspases (Ac-DEVD-cho and Ac-YVA D-cmk) could significantly attenuate the extent of cleavage/activation of PAK2 induced by UV irradiation. Collectively, the results demonstr ate that cleavage and activation of PAK2 can be induced during the ear ly stages of UV UV-irradiation-triggered apoptosis and indicate the in volvement of CPP32/caspase-3 in this process. J. Cell. Biochcm. 70:442 -454, 1998 (C) 1998 Wiley-Liss, Inc.