Identification of two novel synaptic γ-secretase associated proteins that affect amyloid β-peptide levels without altering Notch processing

Synaptic degeneration is one of the earliest hallmarks of Alzheimer disease (AD) and results in loss of cognitive function. One of the causative agents for the synaptic degeneration is the amyloid beta-peptide (A beta), which is formed from its precursor protein by two sequential cleavages mediated by beta- and gamma-secretase. We have earlier shown that gamma-secretase activity is enriched in synaptic compartments, suggesting that the synaptotoxic A beta is produced locally. Proteins that interact with gamma-secretase at the synapse and regulate the production of A beta can therefore be potential therapeutic targets. We used a recently developed affinity purification approach to identify gamma-secretase associated proteins (GSAPs) in synaptic membranes and synaptic vesicles prepared from rat brain. Liquid chromatography-tandem mass spectrometry analysis of the affinity purified samples revealed the known gamma-secretase components presenilin-1, nicastrin and Aph-1b along with a number of novel potential GSAPs. To investigate the effect of these GSAPs on APP processing, we performed siRNA experiments to knock down the expression of the GSAPs and measured the A beta levels. Silencing of NADH dehydrogenase [ubiquinone] iron-sulfur protein 7 (NDUFS7) resulted in a decrease in A beta levels whereas silencing of tubulin polymerization promoting protein (TPPP) resulted in an increase in A beta levels. Treatment with gamma-secretase inhibitors often results in Notch-related side effects and therefore we also studied the effect of the siRNAs on Notch processing. Interestingly, silencing of TPPP or NDUFS7 did not affect cleavage of Notch. We also studied the expression of TPPP and NDUFS7 in control and AD brain and found NDUFS7 to be highly expressed in vulnerable neurons such as pyramidal neurons in the hippocampus, whereas TPPP was found to accumulate in intraneuronal granules and fibrous structures in hippocampus from AD cases. In summary, we here report on two proteins, TPPP and NDUFS7, which interact with gamma-secretase and alter the A beta levels without affecting Notch cleavage. (C) 2012 Elsevier Ltd. All rights reserved.