FAD mutations in presenilin-1 or amyloid precursor protein decrease the efficacy of a gamma-secretase inhibitor: Evidence for direct involvement of PS1 in the gamma-secretase cleavage complex

To investigate the mechanism of regulation of AO production by familial Alz heimer's disease (FAD)linked presenilin 1 (PS1), we used a cell-free system that allows de novo AO generation to examine whether PS1 participates dire ctly in the gamma -secretase reaction. Optimal AB generation in vitro was a chieved at mildly acidic pH and could be inhibited by the aspartyl protease inhibitor pepstatin A, consistent with the suggestion that gamma -secretas e is an aspartyl protease. Dominant negative mutations of the critical tran smembrane aspartates in PS1 or full deletion of PS1 did not alter the matur ation of APP in the secretory pathway. Instead, PS1 had a direct effect on the inhibition of AO production by a designed peptidomimetic inhibitor: the inhibition was significantly less effective in cells expressing FAD-causin g mutations in either APP or PS1 than in cells expressing the wild-type pro teins. Taken together, these findings suggest that PS1 participates physica lly in a complex with APP during the gamma -secretase cleavage event. (C) 2 000 Academic Press.