The pro-apoptotic drug camptothecin stimulates phospholipase D activity and diacylglycerol production in the nucleus of HL-60 human promyelocytic leukemia cells

It has recently been reported (T. Shimizu et at, J. Biol. Chem., 273: 8669- 8674, 1998) that the pro-apoptotic drug, camptothecin, an inhibitor of topo isomerase I, induces a protein kinase C-alpha-mediated phosphorylation of l amin B in HL-60 cells, which precedes both degradation of lamin B and fragm entation of DNA, In this paper, we report that, in HL-60 cells exposed to c amptothecin, there is a rapid and sustained increase of nuclear protein kin ase C-alpha activity that is due to an increase in the amount of protein ki nase C-alpha present in the nucleus. The enhancement of nuclear kinase C ac tivity is preceded by an increase in the mass of nuclear diacylglycerol. As demonstrated by its sensitivity to propranolol, the nuclear diacylglycerol mass increase is due to the activation of a phospholipase D. Indeed, inhib itors of neither phosphatidylcholine-specific phospholipase C nor phosphoin ositide-specific phospholipase C blocked the rise in nuclear diacylglycerol , In vitro assays also demonstrated the activation of a nuclear phospholipa se D, but not of a phosphoinositide-specific phospholipase C, after treatme nt with camptothecin. Propranolol was also able to block the rise in nuclea r protein kinase C-alpha activity, thus suggesting that the increase in dia cylglycerol mass is important for the activation of the kinase at the nucle ar level. Moreover, propranolol was capable of drastically reducing the num ber of HL-60 cells that underwent apoptosis after treatment with camptothec in, Our results show the activation during apoptosis of a phospholipase D-m ediated signaling pathway operating at the nuclear level. This pathway may represent an attractive therapeutic target for the modulation of apoptotic events in human disease.