Caveolae, Plasma Membrane Microdomains for α-Secretase-mediated Processing of the Amyloid Precursor Protein

Abstract

Caveolae are plasma membrane invaginations where key signaling elements are concentrated. In this report, both biochemical and histochemical analyses demonstrate that the amyloid precursor protein (APP), a source of A␤ amyloid peptide, is enriched within caveolae. Caveolin-1, a principal component of caveolae, is physically associated with APP, and the cytoplasmic domain of APP directly participates in this binding. The characteristic C-terminal fragment that results from APP processing by ␣-secretase, an as yet unidentified enzyme that cleaves APP within the A␤ amyloid sequence, was also localized within these caveolae-enriched fractions. Further analysis by cell surface biotinylation revealed that this cleavage event occurs at the cell surface. Importantly, ␣-secretase processing was significantly promoted by recombinant overexpression of caveolin in intact cells, resulting in increased secretion of the soluble extracellular domain of APP. Conversely, caveolin depletion using antisense oligonucletotides prevented this cleavage event. Our current results indicate that caveolae and caveolins may play a pivotal role in the ␣-secretase-mediated proteolysis of APP in vivo.Senile plaques and paired helical filaments are the hallmarks of the brain pathology of Alzheimer's disease (1). The principal component of the senile plaque is the A␤ amyloid peptide, which is composed of 39 -43 amino acid residues. The A␤ amyloid peptide is derived from a full-length precursor protein, termed APP 1 (amyloid precursor protein) (2). Alternate splicing of the APP gene generates at least 10 distinct isoforms; APP 695 is the brain-specific isoform. The A␤ amyloid peptide is generated by the processing of APP with ␤-and ␥-secretases (3). Alternatively, APP is processed by ␣-secretase, which cleaves APP within the A␤ sequence, thereby precluding the formation of A␤ (4). The identities of these secretases remain unknown.In some inherited forms of Alzheimer's disease, point mutations have been identified within the coding sequence of the APP gene (5). These mutations co-segregate with the disease phenotype and cause Alzheimer's disease. Understanding the molecular function and processing of APP is therefore critical to unraveling the molecular basis of Alzheimer's disease.One approach to elucidate the function of APP is to identify APP-interacting proteins. At least five distinct classes of molecules have been identified as APP binding partners as follows: G o (6), Fe 65 (7), X11 protein (8), Fe 65 -like protein (9), and APP-BP1 (10). The APP domain that interacts with G o has been localized to residues His 657 -Lys 676 within the cytoplasmic domain of APP 695 . G o is a brain-specific member of heterotrimeric GTP-binding protein (G-protein) family. The in vivo interaction between APP 695 and G o results in apoptotic cell death (11) and inhibition of cAMP response element trans-activation (12). In contrast, functional consequences of interactions between APP and other binding partners have not yet been described. However, it is likely th...

Caveolin proteins and estrogen signaling in the brain

“…A wealth of information regarding caveolins and their influence on brain function has recently emerged. Topics include, but are not limited to, caveolin regulation of synaptic strength, motor control, and Alzheimer's disease [101][102][103][104][105][106]. These studies demonstrate that caveolins play critical roles in both neuronal and glial processes.…”

Amyloid precursor protein, although partially detergent-insoluble in mouse cerebral cortex, behaves as an atypical lipid raft protein

“…Thus, although APP, alkaline phosphodiesterase I, aminopeptidase A and clathrin are present in DIGs isolated from mouse cerebral cortex, they do not behave as typical DIG proteins. The observation that altering the protein\ detergent ratio used during solubilization of the membranes can alter the apparent distribution of APP and other atypical DIGs proteins in the sucrose density gradients may account for the apparently conflicting reports on the presence of APP in DIGs [27,31,32,[35][36][37][38]. One possible explanation to reconcile these conflicting data is that APP, alkaline phosphodiesterase I and clathrin are ineffectively solubilized contaminants of the DIGs fraction.…”

“…DIGs isolated from COS-7, HEK293 and Madin-Darby canine kidney (' MDCK ') cells have been shown to retain the entire cell complement of APP [36]. In this instance APP was shown to interact directly with caveolin, and depletion of the latter protein using antisense oligonucleotides prevented α-secretase cleavage, suggesting that caveolin may have a role in the α-secretase-mediated processing of APP in non-neuronal cells.…”

“…It has been reported also that a large proportion of Aβ, along with various levels of presenilin-1 and presenilin-2, are present in DIGs isolated from either brain tissue or neuronal cell cultures [31,32,40]. The presence of these Alzheimer's-disease-related proteins in DIGs has led to the hypothesis that lipid rafts may be a site for the proteolytic processing of APP [31,36]. However, our observation that APP is present in DIGs from brain tissue at levels no higher than several other non-DIGs proteins brings into question whether lipid rafts are focal points for APP proteolytic processing.…”

“…Currently there are conflicting reports as to whether APP is present in lipid rafts, where its processing by the secretases may occur. Some studies have reported that a minor amount of APP is present in DIGs [31,[35][36][37], while others have failed to detect it in such fractions [27,32,38].…”