Abnormalities in neuronal cholesterol homeostasis have been suspected or observed in several neurodegenerative disorders including Alzheimer’s disease, Parkinson’s disease and Huntington’s disease. However, it has not been demonstrated whether an increased abundance of cholesterol in neurons in vivo contributes to neurodegeneration. To address this issue, we used RNA interference methodology to inhibit the expression of cholesterol 24-hydroxylase, encoded by the Cyp46a1 gene, in the hippocampus of normal mice. Cholesterol 24-hydroxylase controls cholesterol efflux from the brain and thereby plays a major role in regulating brain cholesterol homeostasis. We used an adeno-associated virus vector encoding short hairpin RNA directed against the mouse Cyp46a1 mRNA to decrease the expression of the Cyp46a1 gene in hippocampal neurons of normal mice. This increased the cholesterol concentration in neurons, followed by cognitive deficits and hippocampal atrophy due to apoptotic neuronal death. Prior to neuronal death, the recruitment of the amyloid protein precursor to lipid rafts was enhanced leading to the production of β-C-terminal fragment and amyloid-β peptides. Abnormal phosphorylation of tau and endoplasmic reticulum stress were also observed. In the APP23 mouse model of Alzheimer’s disease, the abundance of amyloid-β peptides increased following inhibition of Cyp46a1 expression, and neuronal death was more widespread than in normal mice. Altogether, these results suggest that increased amounts of neuronal cholesterol within the brain may contribute to inducing and/or aggravating Alzheimer’s disease.

Alzheimer's disease (AD) is characterized by both amyloid and Tau pathologies. The amyloid component and altered cholesterol metabolism are closely linked, but the relationship between Tau pathology and cholesterol is currently unclear. Brain cholesterol is synthesized in situ and cannot cross the blood–brain barrier: to be exported from the central nervous system into the blood circuit, excess cholesterol must be converted to 24S-hydroxycholesterol by the cholesterol 24-hydroxylase encoded by the CYP46A1 gene. In AD patients, the concentration of 24S-hydroxycholesterol in the plasma and the cerebrospinal fluid are lower than in healthy controls. The THY-Tau22 mouse is a model of AD-like Tau pathology without amyloid pathology. We used this model to investigate the potential association between Tau pathology and CYP46A1 modulation. The amounts of CYP46A1 and 24S-hydroxycholesterol in the hippocampus were lower in THY-Tau22 than control mice. We used an adeno-associated virus (AAV) gene transfer strategy to increase CYP46A1 expression in order to investigate the consequences on THY-Tau22 mouse phenotype. Injection of the AAV-CYP46A1 vector into the hippocampus of THY-Tau22 mice led to CYP46A1 and 24S-hydroxycholesterol content normalization. The cognitive deficits, impaired long-term depression and spine defects that characterize the THY-Tau22 model were completely rescued, whereas Tau hyperphosphorylation and associated gliosis were unaffected. These results argue for a causal link between CYP46A1 protein content and memory impairments that result from Tau pathology. Therefore, CYP46A1 may be a relevant therapeutic target for Tauopathies and especially for AD.

In Drosophila larvae, growth and developmental timing are regulated by nutrition in a tightly coordinated fashion. The networks that couple these processes are far from understood. Here, we show that the intestine responds to nutrient availability by regulating production of a circulating lipoprotein-associated form of the signaling protein Hedgehog (Hh). Levels of circulating Hh tune the rates of growth and developmental timing in a coordinated fashion. Circulating Hh signals to the fat body to control larval growth. It regulates developmental timing by controlling ecdysteroid production in the prothoracic gland. Circulating Hh is especially important during starvation, when it is also required for mobilization of fat body triacylglycerol (TAG) stores. Thus, we demonstrate that Hh, previously known only for its local morphogenetic functions, also acts as a lipoprotein-associated endocrine hormone, coordinating the response of multiple tissues to nutrient availability.

The activity of membrane proteins such as Na,K-ATPase depends strongly on the surrounding lipid environment. Interactions can be annular, depending on the physical properties of the membrane, or specific with lipids bound in pockets between transmembrane domains. This paper describes three specific lipid-protein interactions using purified recombinant Na,K-ATPase. (a) Thermal stability of the Na,K-ATPase depends crucially on a specific interaction with 18:0/18:1 phosphatidylserine (1-stearoyl-2-oleoyl-sn-glycero-3-phospho-l-serine; SOPS) and cholesterol, which strongly amplifies stabilization. We show here that cholesterol associates with SOPS, FXYD1, and the α subunit between trans-membrane segments αTM8 and -10 to stabilize the protein. (b) Polyunsaturated neutral lipids stimulate Na,K-ATPase turnover by >60%. A screen of the lipid specificity showed that 18:0/20:4 and 18:0/22:6 phosphatidylethanolamine (PE) are the optimal phospholipids for this effect. (c) Saturated phosphatidylcholine and sphingomyelin, but not saturated phosphatidylserine or PE, inhibit Na,K-ATPase activity by 70–80%. This effect depends strongly on the presence of cholesterol. Analysis of the Na,K-ATPase activity and E₁-E₂ conformational transitions reveals the kinetic mechanisms of these effects. Both stimulatory and inhibitory lipids poise the conformational equilibrium toward E₂, but their detailed mechanisms of action are different. PE accelerates the rate of E₁ → E₂P but does not affect E₂(2K)ATP → E₁3NaATP, whereas sphingomyelin inhibits the rate of E₂(2K)ATP → E₁3NaATP, with very little effect on E₁ → E₂P. We discuss these lipid effects in relation to recent crystal structures of Na,K-ATPase and propose that there are three separate sites for the specific lipid interactions, with potential physiological roles to regulate activity and stability of the pump.