Activation of DNA-dependent protein kinase may play a role in apoptosis ofhuman neuroblastoma cells

Treating SH-SY5Y human neuroblastoma cells with 1 mu M staurosporine result ed in a three- to fourfold higher DNA-dependent protein kinase (DNA-PK) act ivity compared with untreated cells. Time course studies revealed a biphasi c effect of staurosporine on DNA-PK activity: an initial increase that peak ed by 4 h and a rapid decline that reached similar to 5-10% that of untreat ed cells by 24 h of treatment. Staurosporine induced apoptosis in these cel ls as determined by the appearance of internucleosomal DNA fragmentation an d punctate nuclear morphology. The maximal stimulation of DNA-PK activity p receded significant morphological changes that occurred between 4 and 8 h ( 40% of total number of cells) and increased with time, reaching 70% by 48 h . Staurosporine had no effect on caspase-1 activity but stimulated caspase- 3 activity by I0-15-fold in a time-dependent manner, similar to morphologic al changes. Similar time-dependent changes in DNA-PK activity, morphology, and DNA fragmentation occurred when the cells were exposed to either 100 mu M ceramide or UV radiation. In all these cases the increase in DNA-PK acti vity preceded the appearance of apoptotic markers, whereas the loss in acti vity was coincident with cell death. A cell-permeable inhibitor of DNA-PK, OK-1035, significantly reduced staurosporine-induced punctate nuclear morph ology and DNA fragmentation. Collectively, these results suggest an intrigu ing possibility that activation of DNA-PK may be involved with the inductio n of apoptotic cell death.