Amyloid beta peptide (Abeta) has a key role in the pathological process of Alzheimer's disease (AD), but the physiological function of Abeta and of the amyloid precursor protein (APP) is unknown. Recently, it was shown that APP processing is sensitive to cholesterol and other lipids. Hydroxymethylglutaryl-CoA reductase (HMGR) and sphingomyelinases (SMases) are the main enzymes that regulate cholesterol biosynthesis and sphingomyelin (SM) levels, respectively. We show that control of cholesterol and SM metabolism involves APP processing. Abeta42 directly activates neutral SMase and downregulates SM levels, whereas Abeta40 reduces cholesterol de novo synthesis by inhibition of HMGR activity. This process strictly depends on gamma-secretase activity. In line with altered Abeta40/42 generation, pathological presenilin mutations result in increased cholesterol and decreased SM levels. Our results demonstrate a biological function for APP processing and also a functional basis for the link that has been observed between lipids and Alzheimer's disease (AD).

Proteolytic processing of the amyloid precursor protein by ␤-secretase yields A4CT (C99), which is cleaved further by the as yet unknown ␥-secretase, yielding the ␤-amyloid (A␤) peptide with 40 (A␤ 40 ) or 42 residues (A␤ 42 ). Because the position of ␥-secretase cleavage is crucial for the pathogenesis of Alzheimer's disease, we individually replaced all membrane-domain residues of A4CT outside the A␤ domain with phenylalanine, stably transfected the constructs in COS7 cells, and determined the effect of these mutations on the cleavage specificity of ␥-secretase (A␤ 42 The main proteinaceous component of the amyloid plaques found in the brains of patients with Alzheimer's disease (AD) is ␤-amyloid (A␤; refs. 1 and 2), an Ϸ4-kDa peptide that is derived from the larger amyloid precursor protein (APP; ref.3). APP processing by the as yet unidentified protease activities, termed ␣-, ␤-, and ␥-secretases, leads to a variety of different soluble and membrane-bound proteins (for reviews, see refs. 4 and 5). The ␣-secretase activity cleaves APP within the A␤ domain and thus precludes the generation of A␤. This cleavage yields secretory ␣-APPs, comprising most of the N-terminal ectodomain of APP, and the remaining membrane-bound C-terminal fragment p3CT. Alternatively, APP can be cleaved by the ␤-secretase activity at the N terminus of A␤, generating a truncated, soluble ␤-APPs and a C-terminal fragment of 99 residues (A4CT, C99). The ␤-secretase product A4CT contains the entire A␤ domain, the transmembrane domain, and the cytoplasmic tail of APP and represents the direct precursor for A␤ (6, 7).Both membrane-bound C-terminal fragments of APP, A4CT and p3CT, are cleaved by the ␥-secretase activity within their transmembrane domains at the C terminus of A␤ or p3, thus releasing the 40-and 42-residue A␤ peptides (A␤ 40 and A␤ 42 ) and the 24-26 residue p3 peptides (p3 40 and p3 42 ) (8-11). Most cells secrete both peptides A␤ and p3 into the conditioned medium. In neuronal cells, as in primary hippocampal neurons and in kidney 293 cells, A␤, but not p3, also can be found intracellularly and does not seem to be secreted (12-16).The major A␤ species secreted by cultured cells expressing wild-type (wt) APP is A␤ 40 ; the minor species is A␤ 42 (17). Mutations in the APP close to the ␥-cleavage site have been shown to alter the cleavage specificity of the ␥-secretase activity (A␤ 42 ͞A␤ 40 ratio; refs. 14 and 18-20). However, the factors that determine this cleavage specificity are unknown. Experiments with inhibitors of ␥-secretase activity suggest that distinct proteases generate the A␤ 40 and A␤ 42 peptides (11,(21)(22)(23)), but it is not known whether these enzymes are related or not.Furthermore, although ␥-cleavage occurs in the transmembrane domain of A4CT, it is not clear whether the cleavage occurs while A4CT is inserted into the membrane or after release of A4CT from the membrane. Understanding the substrate specificity of the ␥-secretase activity is of great importance, because the cleavage at residue 42 ...

Alzheimer disease is characterized by accumulation of the ␤-amyloid peptide (A␤) generated by ␤-and ␥-secretase processing of the amyloid precursor protein (APP). The intake of the polyunsaturated fatty acid docosahexaenoic acid (DHA) has been associated with decreased amyloid deposition and a reduced risk in Alzheimer disease in several epidemiological trials; however, the exact underlying molecular mechanism remains to be elucidated. Here, we systematically investigate the effect of DHA on amyloidogenic and nonamyloidogenic APP processing and the potential cross-links to cholesterol metabolism in vivo and in vitro. DHA reduces amyloidogenic processing by decreasing ␤-and ␥-secretase activity, whereas the expression and protein levels of BACE1 and presenilin1 remain unchanged. In addition, DHA increases protein stability of ␣-secretase resulting in increased nonamyloidogenic processing. Besides the known effect of DHA to decrease cholesterol de novo synthesis, we found cholesterol distribution in plasma membrane to be altered. In the presence of DHA, cholesterol shifts from raft to non-raft domains, and this is accompanied by a shift in ␥-secretase activity and presenilin1 protein levels. Taken together, DHA directs amyloidogenic processing of APP toward nonamyloidogenic processing, effectively reducing A␤ release. DHA has a typical pleiotropic effect; DHA-mediated A␤ reduction is not the consequence of a single major mechanism but is the result of combined multiple effects.