Inhibitors of γ-secretase stabilize the complex and differentially affect processing of amyloid precursor protein and other substrates

gamma-Secretase inhibitors (GSIs) are drugs used in research to inhibit production of A beta and in clinical trials to treat Alzheimer's disease (AD). They inhibit proteolytic activities of gamma-secretase noncompetitively by unknown mechanisms. Here, we used cortical neuronal cultures expressing endogenous levels of enzymes and substrates to study the effects of GSIs on the structure and function of gamma-secretase. We show that GSIs stabilize the interactions between the C-terminal fragment of presenilin (PS-CTF), the central component of the gamma-secretase complex, and its partners the APH-1/nicastrin and PS1-NTF/PEN-2 subcomplexes. This stabilization dose-dependently correlates with inhibition of N-cadherin cleavage, a process limited by enzyme availability. In contrast, production of amyloid precursor protein (APP) intracellular domain (AICD) is insensitive to low concentrations of GSIs and is limited by substrate availability. Interestingly, APP is processed by both PS1- and PS2-containing gamma-secretase complexes, while N-cadherin and ephrinB1 are processed only by PS1-containing complexes. Paradoxically, low concentrations of GSIs specifically increased the levels of A beta without affecting its catabolism, indicating increased A beta production. Our data reveal a mechanism of gamma-secretase inhibition by GSIs and provide evidence that distinct gamma-secretase complexes process specific substrates. Furthermore, our observations have implications for GSIs as therapeutics because processing of functionally important substrates may be inhibited at lower concentrations than A beta.-Barthet, G., Shioi, J., Shao, Z., Ren, Y., Georgakopoulos, A., Robakis, N. K. Inhibitors of gamma-secretase stabilize the complex and differentially affect processing of amyloid precursor protein and other substrates. FASEB J. 25, 2937-2946 (2011). www.fasebj.org