Aβ46 is processed to Aβ40 and Aβ43, but not to Aβ42, in the low density membrane domains

gamma-Secretase cleaves the transmembrane domain of beta-amyloid precursor protein at multiple sites referred to as gamma-, epsilon-, and zeta-cleavage sites. We previously showed that N-[N-( 3,5-difluoro-phenacetyl)-L- alanyl]-S-phenylglycine t-butyl ester (DAPT), a potent dipeptide gamma- secretase inhibitor, causes differential accumulation of longer amyloid beta-proteins (A beta s) within Chinese hamster ovary cells co-expressing beta C-terminal fragment and wild-type presenilin 1(C99/wtPS1 cells). In this study, we used sucrose density gradient centrifugation to fractionate the membranes from C99/wtPS1 cells that had been pretreated with DAPT. We found that accumulating A beta 46 localized exclusively to low density membrane (LDM) domains. Incubating the A beta 46-accumulating LDM domains at 37 degrees C produced A beta 40, A beta 42, A beta 43, and beta-amyloid precursor protein intracellular domain. The addition of L685,458 completely prevented beta-amyloid precursor protein intracellular domain generation and resulted in a large decrease in the level of A beta 46 and the concomitant appearance of A beta 40 and A beta 43 but not A beta 42. Further addition of DAPT suppressed the production of A beta 40/43 and abolished the decrease in the amount of A beta 46. These data indicate that preaccumulated A beta 46 is processed by gamma-secretase to A beta 40/43 but not to A beta 42 in the LDM domains. The amount of newly produced A beta 40 and A beta 43 was roughly equivalent to the decrease in the amount of A beta 46. Temporal profiles did not show a maximal concentration for A beta 43, suggesting that A beta 46 is processed to A beta 40 and A beta 43 through a nonsuccessive process.