Active γ-secretase is enriched in synaptic membranes prepared from rat brain

Synaptic degeneration is the first pathological hallmark in Alzheimer's disease and the one that correlates best with disease progression. The mechanism for synaptic degeneration is unknown but a strong candidate as a causative agent is amyloid β-peptide (Aβ) oligomers that may cause Ca2+ dysregulation and oxidative stress. Aβ is produced by the processing of the amyloid precursor protein (APP) by two sequential proteolytic events. The last cleavage is performed by a large membrane-bound protein complex called γ-secretase consisting of at least for components. In addition to APP, γ-secretase cleaves a large number of substrates which complicates its potential use as a theurapeutical target. In order to design specific γ-secretase inhibitors/ regulators it is important to increase the knowledge about this enzyme. Several studies have determined the subcellular localisation of γ-secretase to the late secretory-endosomal pathways using different cell-lines. However, the localization in brain has not been reported. Since the synapses seem to be particularly vulnerable for Aβ toxicity we wanted to examine if γ-secretase is localized to the synapses. Methods: We prepared synaptic membranes and synaptic vesicles using sucrose gradients and synaptosomal lysis, and assayed these fractions for subcellular markers, γ-secretase components and γ-secretase activity as measured by AICD and Aβ production. Results: The synaptic membrane fraction was enriched in PSD-95, a post-synaptic membrane marker, but also contained endosomal markers. The γ-secretase activity was dramatically enriched in this fraction, as assayed by endogenous AICD and Aβ production. It is possible that the γ-secretase in synaptic membranes could be differently regulated compared to other parts of the brain and we speculate that synaptic membrane preparations could be useful for finding γ-secretase inhibitors/ regulators that specifically interacts with the synaptic Aβ production. Conclusions: Gamma-secretase activity is enriched in synaptic membranes and high local production of Aβ might contribute to the synaptic degeneration seen in Alzheimer's disease