Effects of RNA interference-mediated silencing of gamma-secretase complex components on cell sensitivity to caspase-3 activation.

Familial Alzheimer's disease mutations in the presenilin 1 gene (PSEN1) have been previously shown to potentiate caspase activation and apoptosis in transfected cells and transgenic mice. However, the mechanism underlying this effect is not known. We set out to determine whether cellular sensitivity to caspase activation could be affected by modulating presenilin 1 (PS1) processing. PS1 processing was altered using RNA interference (RNAi) aimed at silencing the expression of the genes encoding the four components of the gamma-secretase complex, PSEN1, APH-1, PEN-2, and nicastrin. RNAi for these genes was carried out in naive H4 human neuroglioma cells, as well as H4 cell lines overexpressing either wild-type PSEN1 or the Familial Alzheimer's disease mutant PSEN1-Delta9 (PS1-mutant), that were induced to undergo apoptosis. In wild-type PSEN1 cells, RNAi for PEN-2, as expected, increased levels of full-length PS1 (PS1-FL) and decreased PS1 endoproteolysis. This was accompanied by potentiated caspase-3 activation in response to an apoptotic stimulus. In contrast, nicastrin RNAi, which only decreased levels of PS1-amino-terminal fragment and did not affect PS1-FL levels, had no effect on caspase-3 activation during apoptosis. Surprisingly, in the PS1-mutant cells, RNAi for PEN-2 (and APH-1) did not increase but instead reduced the levels of PS1-FL deleted for exon 9. In turn, this was accompanied by attenuated caspase-3 activation in response to an apoptotic stimulus. Finally, in naive H4 cells, PSEN1 RNAi also attenuated caspase-3 activation in response to an apoptotic stimulus. Collectively, these findings indicate that cellular sensitivity to caspase activation correlates with overall PS1 protein levels, particularly with levels of FL-PS1.