Regulation of cholesterol and sphingomyelin metabolism by amyloid-beta and presenilin

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).status: publishe

Gender and the University: Stories So Far and Spaces Between

The chapter discusses the application of Doreen Massey's socio-spatial concepts in my theorising of the university. The chapter draws primarily on my research into the impact of gender and intersectional factors on experiences of the workplace and career in a post-1992 university in which Massey's theoretical concepts shape both the research question and the methodology

PS Dependent APP Cleavage Regulates Glucosylceramide Synthase and is Affected in Alzheimer's Disease

Background: Gangliosides were found to be associated with Alzheimer's disease (AD). Here we addressed a potential function of γ-secretase (presenilin) dependent cleavage of the amyloid-precursor-protein (APP) in the regulation of ganglioside de novo synthesis. Methods: To identify a potential role of γ-secretase and APP in ganglioside de novo synthesis we used presenilin (PS) deficient and APP deficient cells and mouse brains, mutated PS as well as transgenic mice and AD post mortem brains. Changes in glucosylceramide synthase (GCS) activity were identified by incorporation of radiolabeled UDP-glucose in glucosylceramide, changes in gene expression via real-time PCR and Western blot analysis. Alterations in ganglioside levels were determined by thin layer chromatography and mass spectrometry. Results: We found that PS and APP deficiency, in vitro and in vivo, resulted in increased GCS gene expression, elevated enzyme activity and thus increased glucosylceramide and total ganglioside level. Using a specific γ-secretase inhibitor revealed that PS proteolytic activity alters ganglioside homeostasis. By the use of mutated PS causing early onset AD in cell culture and transgenic mice we found that GCS is increased in AD, further substantiated by the use of AD post mortem brains, suffering from sporadic AD. Conclusion: APP processing regulates ganglioside de novo synthesis and is affected in AD

Effect of GM3 and GD3 on APP processing.

<p>(<i>A</i>) Dose-dependent effect of GM3 on Aβ production: COS7 cells stably transfected with SPC99, representing the β-secretase cleaved C-terminal fragment of APP, show a dose-dependent decrease in Aβ production in presence of GM3. Aβ levels were determined by IP and Western blot (WB) analysis. (<i>B</i>) Dose-dependent effect of GD3 on Aβ generation in SPC99 expressing COS7 cells. GD3 enhances Aβ generation dose-dependently. Aβ levels were determined by IP and WB analysis.</p

Effect of the APP intracellular domain (AICD) on GD3S.

<p>(<i>A</i>) MEF, deficient in full-length APP expressing a truncated APP construct lacking 15 aa from the C-terminus (APPΔCT15) show increased GD3S expression and activity. The level of the GD3S activity of the APP/APLP2 knock-out cells (MEF APP/APLP2-/-) are indicated (horizontal dotted black line). Interestingly the MEF APP/APLP2 knock-out level showed a slight less effect strength compared to the MEF APPΔCT15 cells. However this difference did not reach a significant level and might be due to clonal heterogeneity. (<i>B</i>) Increased GD3S expression in APPΔCT15 mouse brains. (<i>C</i>) GD3S gene expression: AICD peptide partially rescues elevated GD3S gene expression in MEF cells expressing the C-terminal truncated APP. An AICD peptide consisting of the last 20 aa from the APP C-terminus (AICD) is able to decrease GD3S expression. (<i>D</i>) shRNA generated Fe65 knock-down cells show increased GD3S gene transcription.</p

Hypothetic model of the physiological functions of Aβ and AICD in the regulation of GD3-synthase (GD3S) – the enzyme controlling major brain ganglioside composition.

<p>(<i>A</i>) Amyloidogenic proteolytic processing of the Alzheimer's amyloid precursor protein (APP) releases amyloid-beta peptides (Aβ) and the intracellular domain of APP (AICD). Aβ and AICD inhibit GD3S, resulting in reduced conversion of <i>a</i>- to <i>b</i>-series gangliosides. As a consequence of reduced conversion of <i>a</i>- to <i>b</i>-series gangliosides, GM3 increases whereas GD3 decreases. In return, both gangliosides, GM3 and GD3, themselves regulate the proteolytic cleavage of APP. The <i>b</i>-series ganglioside GD3 increases, whereas the <i>a</i>-series gangliosides GM3 decreases amyloidogenic proteolytic processing of APP.</p

Molecular mechanisms of APP cleavage products in the regulation of GD3S enzyme activity.

<p>(<i>A</i>) In absence of Aβ peptides <i>a</i>-series ganglioside GM3 binds to GD3S and is converted to the <i>b</i>-series ganglioside GD3. In presence of Aβ, Aβ binds ganglioside GM3, forming an Aβ-GM3 complex. This complex still binds to GD3S, but cannot be converted to GD3. (<i>B</i>) Dual function of Aβ and AICD in GD3S regulation. Aβ reduces enzyme activity of GD3S by forming an Aβ-GM3 complex, resulting in reduced turnover of GM3 to GD3. AICD binds the adaptor protein Fe65 and reduces GD3S gene transcription, which also results in reduced turnover of GM3 to GD3.</p