Evidence that Enzyme Processivity Mediates Differential Aβ Production by PS1 and PS2

Abstract

The γ-secretase complex cleaves the carboxy-terminal 99 residue (C99) fragment of the amyloid precursor protein (APP) to generate the amyloid-β (Aβ) peptide. The catalytic activity of this complex is mediated either by the presenilin-1 (PS1) or the presenilin-2 (PS2) subunit. In vitro and in vivo studies have demonstrated that PS1-containing complexes generate more total Aβ product than PS2-containing complexes, indicating greater cleavage activity by PS1-containing γ-secretase complexes at the APP γ-site. However, it remains untested whether γ-secretase cleavage at the APP ε-site, which precedes γ-site cleavage and produces the physiologically active APP intracellular domain (AICD), follows the same rule. Using a novel Swedish APP-GVP substrate to facilitate the parallel detection of Aβ and AICD products from PS1-/-/PS2-/- cells co-transfected with either PS1 or PS2, we observed that while PS1 generates more total Aβ product than PS2, consistent with published reports, PS1 and PS2 unexpectedly generate equal amounts of AICD product. We also observed that PS1 and PS2 produce equivalent amounts of Notch intracellular domain (NICD), indicating equal cleavage activity at the Notch S3-site (the corollary of the APP ε-site). Our findings suggest that processivity differences between PS1 and PS2 underlie the differential production of Aβ peptide. Taken together these findings offer novel insights into γ-secretase biology and have important implications for therapeutically targeting γ-secretase

Specificity of presenilin‐1‐ and presenilin‐2‐dependent γ‐secretases towards substrate processing

“…Several studies have reported a lower activity of PS2 γ‐secretases compared to PS1 γ‐secretases . Our work confirms, both in single‐KO and rescued cells, that APP substrates are more efficiently processed by PS1 than by PS2 and this is in line with published results showing a predominant implication of PS1 in the γ‐secretase activity . However, we observed that PS2‐expressing cells still efficiently cleave C99‐GVP.…”

“…Absence of PS1 (PS1KO) provokes a drastic reduction of APP and Notch cleavages, which are restored by the expression of human PS1 (rPS1wt). These observations are consistent with previously published data [21,22] underlining the predominant role of PS1 in c-secretase-dependent substrate processing. Strikingly, our results further indicate that pharmacological c-secretase inhibitors differently affect PS1 and PS2 c-secretases.…”

Flexible and Accurate Substrate Processing with Distinct Presenilin/γ-Secretases in Human Cortical Neurons

“…However, mice lacking both PS1 and PS2 die much earlier than PS1 KO mice (Donoviel et al, 1999), and the neurogenesis defects in PS1/PS2-null neural progenitor cells (NPCs) are much more severe than those in PS1-deficient NPCs (Handler et al, 2000;Hitoshi et al, 2002;Kim and Shen, 2008). In terms of Aβ generation, PS1 and PS2 exhibit different properties in nonneuronal cells and/or nonhuman experimental systems, most of which express exogenous PS, and PS2/γsecretase shows less total proteolytic activity than PS1/γ-secretase (Lai et al, 2003;Yonemura et al, 2011;A. Pintchovski et al, 2013).…”

Mechanism of Cellular Production and in Vivo Seeding Effects of Hexameric β-Amyloid Assemblies

“…Our results showed the absence of any signi cant change in the processing of C99 or the release of Aβ when cells have a nearly 50% reduction of PS1 protein levels. This was rather surprising regarding the previous reports on the preponderant importance of PS1 for γ-secretase substrates cleavage and overall Aβ production [33][34][35][36][37]. One can imagine, as mentioned above, that the reduction of PS1 protein levels is not signi cant enough to observe any effect.…”

“…A recent cryo-EM study revealed a similar structure and conformational state between both presenilins (PSs), suggesting similar catalytic activities, but differences in their membraneanchoring motifs [32]. We and others have repeatedly shown a major contribution of PS1 to Aβ production intended as substrate cleavage [33][34][35][36][37], rendering the exact role of PS2 in the amyloid pathology less understood, and in turn PS2 less attractive as a therapeutic target. However, more recent ndings revealed the enrichment of PS2 γ-secretases in endosomal compartments, which might explain its lesser overall contribution to substrate cleavage simply by secondary encounters; substrates such as APP and its CTFs are indeed likely to be e ciently processed in cells rst along tra cking through the secretory pathway and at the plasma membrane before reaching the endosomal pathway [38].…”