Conserved “PAL” sequence in presenilins is essential for γ-secretase activity, but not required for formation or stabilization of γ-secretase complexes

Abstract

Generation of Aβ from the β-amyloid precursor protein (APP) requires a series of proteolytic processes, including an intramembranous cleavage catalyzed by an aspartyl protease, γ-secretase. Two aspartates in presenilins (PS) are required for γ-secretase activity (D257 and D385 of PS1), suggesting that PS may be part of this protease. Little is known concerning the importance of other sequences in PS for activity. We introduced point mutations (P433L, A434D, L435R) into a completely conserved region C-terminal to transmembrane domain eight of PS1. The P433L mutation abolished PS1 endoproteolysis as well as γ-secretase cleavage of APP and Notch in PS1/2 K/O cells. In HEK cells, expression of PS1/P433L reduced Aβ production and caused accumulation of APP C-terminal stubs. When the P433L mutation was introduced into the non-cleavable Δexon 9 (ΔE9) variant of PS1, it abolished γ-secretase cleavage of APP and Notch. The P433L holoprotein is stable and incorporated into the high molecular weight γ-secretase complex, arguing that P433 is not necessary for formation or stabilization of the γ-secretase complex. Other non-conservative mutations in the invariant P433A434L435 sequence also result in a phenotype that is indistinguishable from the aspartate mutants, suggesting a direct involvement of this sequence in γ-secretase activity.

Keywords

• Alzheimer's disease
• Presenilin
• γ-secretase
• APP
• Aβ
• Notch