γ-secretase cleavage site specificity differs for intracellular and secretory amyloid β

The final step in Aβ generation is the cleavage of the C-terminal 99 amino acid residues of the amyloid precursor protein by γ-secretase. γ-Secretase activity is closely linked to the multi-transmembrane-spanning proteins presenilin 1 and presenilin 2. To elucidate whether the cleavage site specificities of γ-secretase leading to the formation of secreted and intracellular Aβ are identical, we made use of point mutations close to the γ-cleavage site, known to have a dramatic effect on the 42/40 ratio of secreted Aβ. We found that the selected point mutations only marginally influenced the 42/40 ratio of intracellular Aβ, suggesting differences in the γ-secretase cleavage site specificity for the generation of secreted and intracellular Aβ. The analysis of the sub-cellular compartments involved in the generation of intracellular Aβ revealed that Aβ is not generated in the early secretory pathway in the human SH-SY5Y neuroblastoma cell line. In this study we identified late Golgi compartments to be involved in the generation of intracellular Aβ. Moreover, we demonstrate that the presence of processed PS1 is not sufficient to obtain γ-secretase processing of the truncated amyloid precursor protein construct C99, proposing the existence of an additional factor downstream of the endoplasmic reticulum and early Golgi required for the formation of an active γ-secretase complex.