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PLOS ONE, April 2013; Vol8/Issue4/e61299

Generation of Amyloid-b Is Reduced by the Interaction of Calreticulin with Amyloid Precursor Protein, Presenilin and Nicastrin

By: Nina Stemmer, Elena Strekalova, Nevena Djogo, Frank Plöger, Gabriele Loers, David Lutz, Friedrich Buck, Marek Michalak, Melitta Schachner, Ralf Kleene

Dysregulation of the proteolytic processing of amyloid precursor protein by c-secretase and the ensuing generation of amyloid-b is associated with the pathogenesis of Alzheimer’s disease. Thus, the identification of amyloid precursor protein binding proteins involved in regulating processing of amyloid precursor protein by the c-secretase complex is essential for understanding the mechanisms underlying the molecular pathology of the disease. We identified calreticulin as novel amyloid precursor protein interaction partner that binds to the c-secretase cleavage site within amyloid precursor protein and showed that this Ca2+- and N-glycan-independent interaction is mediated by amino acids 330–344 in the C-terminal Cdomain of calreticulin. Co-immunoprecipitation confirmed that calreticulin is not only associated with amyloid precursor protein but also with the c-secretase complex members presenilin and nicastrin. Calreticulin was detected at the cell surface by surface biotinylation of cells overexpressing amyloid precursor protein and was co-localized by immunostaining with amyloid precursor protein and presenilin at the cell surface of hippocampal neurons. The P-domain of calreticulin located between the N-terminal N-domain and the C-domain interacts with presenilin, the catalytic subunit of the c-secretase complex. The P- and C-domains also interact with nicastrin, another functionally important subunit of this complex. Transfection of amyloid precursor protein overexpressing cells with full-length calreticulin leads to a decrease in amyloid-b42 levels in culture supernatants, while transfection with the P-domain increases amyloid-b40 levels. Similarly, application of the recombinant P- or C-domains and of a synthetic calreticulin peptide comprising amino acid 330–344 to amyloid precursor protein overexpressing cells result in elevated amyloid-b40 and amyloid-b42 levels, respectively. These findings indicate that the interaction of calreticulin with amyloid precursor protein and the c-secretase complex regulates the proteolytic processing of amyloid precursor protein by the c-secretase complex, pointing to calreticulin as a potential target for therapy in Alzheimer’s disease.

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