Abstract
Alzheimer disease (AD) is characterized by senile plaques, which are mainly composed of beta amyloid (Abeta) peptides. Abeta is cleaved off from amyloid precursor protein (APP) with consecutive proteolytic processing: beta-secretase, followed by gamma-secretase. Here, we show that BRI3, a member of the BRI gene family that includes the familial British and Danish dementia gene BRI2, interacts with APP and serves as an endogenous negative regulator of Abeta production. BRI3 colocalizes with APP along neuritis in differentiated N2a cells; endogenous BRI3-APP complexes are readily detectable in mouse brain extract; reducing endogenous BRI3 levels by RNA interference results in increased Abeta secretion. BRI3 resembles BRI2, because BRI3 overexpression reduces both alpha- and beta-APP cleavage. We propose that BRI3 inhibits the various processing of APP by blocking the access of alpha- and beta-secretases to APP. However, unlike BRI2, the binding of BRI3 to the beta-secretase cleaved APP C-terminal fragment is negligible and BRI3 does not cause the massive accumulation of this APP fragment, suggesting that, unlike BRI2, BRI3 is a poor gamma-cleavage inhibitor. Competitive inhibition of APP processing by BRI3 may provide a new approach to AD therapy and prevention.
Highlights
Because BRI2, the homolog of BRI3, interacts with amyloid precursor protein (APP) and inhibits APP processing [20, 21, 24] we investigated the role of BRI3 in regulating the processing of APP
Our observation that BRI3 colovance, only mature APP binds BRI3 endogenously. This calizes with APP in the vesicular structures along the neurites resembles what we observed in BRI3-stable transfected suggests the possibility that BRI3 can regulate APP processing clones (Fig. 5A) and what we have previously described for during APP transport through the neurites
We isolated BRI3, together with the family member BRI2, mouse brain. These data demonstrate that BRI3 is during a genetic screening for membrane-bound APP ligands an important physiological regulator of ␣- and -cleavage of APP and hint to the possibility that factors interfering with the regulation of APP processing by BRI3 may contribute to Alzheimer disease (AD) pathogenesis
Summary
Split-ubiquitin Screening—The construction of APP as bait and screening method were described elsewhere [20]. The BRI3 cDNA obtained in the two-hybrid screening was cloned into a FLAG-tagged mammalian expression vector, and cotransfected into HeLa cells together with APP. FLAG-BRI3 was cotransfected with APP into HeLa cells, and the lysates were immunoprecipitated with the APP CTF antibody (␣APPct) or a control rabbit polyclonal antibody (RP). Each member of the BRI family was transfected into HeLa cells together with APP, and BRIs were precipitated from the prepared lysates with FLAG released by furin cleavage Domain (GFP-AID), an APP construct lacking intracellular C-terminal 31 amino acids (APP-Ncas), two familial AD mutants of APP (APP London and APP Swedish), an APP homolog (APLP2), or unrelated type II transmembrane protein (Fas ligand) was cotransfected to HeLa cells with or without FLAG-BRI3.
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