The discrepancy between presenilin subcellular localization and γ-secretase processing of amyloid precursor protein

We investigated the relationship between PSI and gamma -secretase processing of amyloid precursor protein (APP) in primary cultures of neurons. Increasing the amount of APP at the cell surface or towards endosomes did not significantly affect PS1-dependent gamma -secretase cleavage, although little PS1 is present in those subcellular compartments. In contrast, almost no gamma -secretase processing was observed when holo-APP or APP-C99, a direct substrate for gamma -secretase, were specifically retained in the endoplasmic reticulum (ER) by a double lysine retention motif. Nevertheless, APP-C99-dilysine (KK) colocalized with PS1 in the ER. In contrast, APP-C99 did not colocalize with PS1, but was efficiently processed by PS1-dependent gamma -secretase. APP-C99 resides in a compartment that is negative for ER, intermediate compartment, and Golgi marker proteins. We conclude that gamma -secretase cleavage of APP-C99 occurs in a specialized subcellular compartment where little or no PS1 is detected. This suggests that at least one other factor than PS1, located downstream of the ER, is required for the gamma -cleavage of APP-C99. In agreement, we found that intracellular gamma -secretase processing of APP-C99-KK both at the gamma 40 and the gamma 42 site could be restored partially after brefeldin A treatment. Our data confirm the "spatial paradox" and raise several questions regarding the PS1 is gamma -secretase hypothesis.