Caveolae-like membranes and the amyloid precursor protein : a critical reassessment.

Caveolae in non-neuronal cells are specialized membrane regions in &ich a range of signalling mdearles are concentrated, along with a characteristc protein called caveolin. Although caveolin is virtually undetectable in neum. caveolae-like membranes (CLMs) with similar characteristics can be isolated fmm brain tissue. CLMs. like caveolae. contain many signalling molecules and glycosyl phosphatiiylinositoC(GPI)-anchored proteins but also possess a neuron-specif* CLM marker protein. flotillin. These membranes have sparked considerable interest in the fdd of Alzheimeh disease (AD) research because they contain very high levels of ganglioside GM1 and cholesteml. In Alzheimeh disease amyloid beta (Ap) peptides derived from proteoiytic cleavage of the amyloi precursor protein (APP) aggregate to form the amyloidogenic plaques characteristic of the disease. Cholesterol and ganglioside GMI have been shown to accelerate protem aegregation or AD pepiiie fibril formation in v#ro. More recently, APP and Ag pepiiies have been identified as components of CLMs. However, only a minor proportion (I-15%) of APP is located in these membranes. In the cumnt study, we have set out to determine whether APP is a genuine component of CLMs. CLMs were isolated from mouse brain cerebral cortex by solubilization of the tissue in Triton X-100 and subsequent flotation In a sucrose density gradient. Our results show that CLMs isolated under different protein:detergent ratios have similar, yet distinctly different. protein profiles. Flotillin and GPI-anchored proteins (CLM marker proteins) were located exclusively in the CLM fraction under all the prolein:detergent ratios employed. As the pro1ein:detergent ratio was decreased the concentration of flotillin and the GPI-anchored marker proteins increased. However, levels of APP and other 'non-CLM' proteins (clathrin and alkaline phosphodiesterase I) did not increase in parallel with the levels of flotillin and the GPI-anchored proteins. These results indicate that, rather than being a genuine component of caveolae-like membranes, APP in this fraction arises from conlamination of the CLM fradion with distind vesicles of a similar density