Amyloid Beta A4 Precursor Protein-binding Family B Member 1 (FE65) Interactomics Revealed Synaptic Vesicle Glycoprotein 2A (SV2A) and Sarcoplasmic/Endoplasmic Reticulum Calcium ATPase 2 (SERCA2) as New Binding Proteins in the Human Brain

ABSTRACT: FE65 is a cytosolic adapter protein and an important binding partner of the amyloid precursor protein (APP). Dependent on Thr668 phosphorylation in APP, which influences amyloidogenic APP processing, FE65 undergoes nuclear translocation, thereby transmitting a signal from the cell membrane to the nucleus. As this translocation may be relevant in Alzheimer disease (AD) and as FE65 consists of three protein-protein interaction domains able to bind and affect a variety of other proteins and down-stream signaling pathways, the identification of the FE65 interactome is of central interest in AD research. In this study, we identified 121 proteins as new potential FE65 interacting proteins in a pulldown/mass spectrometry approach using human post-mortem brain samples as protein pools for recombinantly expressed FE65. Co-immunoprecipitation assays further validated the interaction of FE65 with the candidates SV2A and SERCA2. In parallel, we investigated the whole cell proteome of primary hippocampal neurons from FE65/FE65L1 double knockout mice. Bioinformatics pipeline: MGF files were generated from RAW files with ProteomDiscoverer 1.3 and imported into ProteinScape 2.1 and analyzed using MASCOT version 2.3.0) with the following parameters: peptide mass tolerance of 10 ppm, fragment mass tolerance of 0.5 Da, enzyme: Trypsin with one missed cleavage site, variable modifications: carbamidomethylation (C), oxidation (M), phosphorylation of S, T and Y, used database: Uniprot/SwissProt (2011_06), taxonomy Homo sapiens + one shuffled decoy per entry (results in 40620 entries, including decoys). For further details please contact Thorsten Müller ([e-mail]), Medizinisches Proteom-Center, Ruhr-Universität Bochum.

INSTRUMENT(S): LTQ Orbitrap, instrument model

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Brain

SUBMITTER: Julian Uszkoreit  

LAB HEAD: Katrin Marcus

PROVIDER: PXD000437 | Pride | 2016-05-18

REPOSITORIES: Pride