Dilysine retrieval signal‐containing p24 proteins collaborate in inhibiting γ‐cleavage of amyloid precursor protein

J. Neurochem. (2010) 115 , 771–781. Abstract γ‐Secretase mediates intramembranous γ‐cleavage and ε‐cleavage of β‐amyloid precursor protein (APP) to liberate β‐amyloid peptide (Aβ) and APP intracellular domain respectively from the membrane. Although the regulatory mechanism of γ‐secretase cleavage remains unresolved, a member of the p24 cargo protein family, named p24δ 1 or TMP21, has been identified as an activity‐modulating component. The p24 family proteins are divided into four subfamilies (p24α, β, δ and γ). In contrast to p24δ 1 , p24β 1 has reportedly no effect on γ‐cleavage. In this study, we determined whether p24α 2 , p24γ 3 or p24γ 4 modulates APP processing. Knockdown of cellular p24α 2 induced a significant increase in Aβ generation but not in APP intracellular domain production in cell‐based and cell‐free assays, whereas p24α 2 over‐expression suppressed Aβ secretion. By contrast, Aβ secretion was not altered by p24γ 3 or p24γ 4 knockdown. Endogenous p24α 2 co‐immunoprecipitated with core components of the γ‐secretase complex, and the anti‐p24α 2 immunoprecipitate exhibited γ‐secretase activity. Mutational disruption of the conserved dilysine ER‐retrieval motifs of p24α 2 and p24δ 1 perturbed inhibition of γ‐cleavage. Simultaneous knockdown, or co‐over‐expression, of these proteins had no additive or synergistic effect on Aβ generation. Our findings suggest that dilysine ER‐retrieval signal‐containing p24 proteins, p24α 2 and p24δ 1 , bind with γ‐secretase complexes and collaborate in attenuating γ‐cleavage of APP.