Flexible and accurate substrate processing with distinct presenilin/γ-secretases in human cortical neurons

Abstract

Mutations in the presenilin genes (PS1, PS2) have been linked to the majority of familial Alzheimer’s disease (AD). Although great efforts have been made to investigate pathogenic PS mutations, which ultimately cause an increase in the toxic form of β-amyloid (Aβ), the intrinsic physiological functions of PS in human neurons remain to be determined. In this study, to investigate the physiological roles of PS in human neurons, we generated PS1 conditional knockout induced pluripotent stem cells (iPSCs), in which PS1 can be selectively abrogated under Cre transduction with or without additional PS2 knockout. We showed that iPSC-derived neural progenitor cells do not confer a maintenance ability in the absence of both PS1 and PS2, showing the essential role of PS in Notch signaling. We then generated PS-null human cortical neurons, where PS1 was intact until full neuronal differentiation occurred. Aβ40 production was reduced exclusively in human PS1/PS2-null neurons along with a concomitant accumulation of APP-CTFs, whereas Aβ42 was decreased in neurons devoid of PS2. Unlike previous studies in mice, in which APP cleavage is largely attributable to PS1, γ-secretase activity seemed to be comparable between PS1 and PS2. In contrast, cleavage of another substrate, N-cadherin, was impaired only in neurons devoid of PS1. Moreover, PS2/γ-secretase exists largely in late endosomes/lysosomes, as measured by specific antibody against the γ-secretase complex, in which Aβ42 species are supposedly produced. Using this novel stem cell-based platform, we assessed important physiological PS1/PS2 functions in mature human neurons, the dysfunction of which could underlie AD pathogenesis.

Significance Statement Presenilins are crucial catalytic subunits of γ-secretase, an intramembranous protease complex, whose mutations underlie AD pathogenesis via the dysregulation of Aβ generation. The γ-secretase complex exhibits heterogeneity via the assembly of PS1 or PS2, but the correlation of γ-secretase heterogeneity with substrate processing remains to be established in human neurons. Here, using a novel iPSC-derived cellular model carrying PS1 and/or PS2 conditional knockout alleles, we uncovered the unique processing of three substrates, Notch, APP and N-cadherin, by PS1 or PS2 in human neural cell contexts. Furthermore, the intrinsic subcellular localization of γ-secretase depends on PS1 or PS2, leading to putative differences in the processing of substrates. This novel platform will help ensure the correlation of γ-secretase/substrates in human neurons.