PITSLRE PROTEIN-KINASE ACTIVITY IS ASSOCIATED WITH APOPTOSIS

Minimal ectopic expression of a 58-kDa protein kinase (PITSLRE beta 1) , distantly related to members of the cdc2 gene family, induces teloph ase delay, abnormal chromosome segregation, and decreased growth rates in Chinese hamster ovary cells. Here eve show that this decrease in c ell growth rate is due to apoptosis. Apoptosis is also induced by ecto pic expression of an amino-terminal deletion mutant containing the cat alytic and C-terminal domains of PITSLRE beta 1 but not by other mutan ts lacking histone H1 kinase activity or by other members of the cdc2 gene family. However, unlike the wild-type PITSLRE beta 1 overexpresso rs, ectopic expression of the N-terminal PITSLRE beta 1 mutant does no t result in telophase delay or abnormal chromosome segregation. These results suggested that the function of this protein kinase could be li nked tb apoptotic signaling. To test this hypothesis, we examined leve ls of PITSLRE mRNA, steady-state protein, and enzyme activity in human T cells undergoing apoptosis after activation with the anti-Fas monoc lonal antibody (MAb). All were substantially elevated shortly after Fa s MAb treatment. In addition to new transcription and translation, pro teolysis contributed to the increased steady-state levels of a novel 5 0-kDa PITSLRE protein, as suggested by the diminution of larger PITSLR E isoforms observed in the same cells. Indeed, treatment of the Fas-ac tivated T cells with a serine protease inhibitor prevented apoptotic d eath and led to the accumulation of larger, less active PITSLRE kinase isoforms but not the enzymatically active 50-kDa PITSLRE isoform. Fin ally, induction of apoptosis by glucocorticoids in the same cell line, as well as by Fas MAb treatment of another T-cell line, led to a simi lar induction of 50-kDa PITSLRE protein levels over time. These findin gs suggest that (i) PITSLRE kinase(s) may lie within apoptotic signali ng pathway(s), (ii) serine protease activation may be an early event i n Fas-activated apoptosis of human T cells, and (iii) some PITSLRE kin ase isoforms may be targets of apoptotic proteases.