Crosstalk between angiotensin and the nonamyloidogenic pathway of Alzheimer's amyloid precursor protein.

The association between hypertension and an increased risk for Alzheimer's disease (AD) and dementia is well established. Many data suggest that modulation of the renin-angiotensin system may be meaningful for the prevention and therapy of neurodegenerative disorders, in particular AD. Proteolytic cleavage of the amyloid precursor protein (APP) by α-secretase precludes formation of neurotoxic Aβ peptides and is expected to counteract the development of AD.

Statins stimulate the production of a soluble form of the receptor for advanced glycation end products.

The beneficial effects of statin therapy in the reduction of cardiovascular pathogenesis, atherosclerosis, and diabetic complications are well known. The receptor for advanced glycation end products (RAGE) plays an important role in the progression of these diseases. In contrast, soluble forms of RAGE act as decoys for RAGE ligands and may prevent the development of RAGE-mediated disorders.

Induction of RAGE shedding by activation of G protein-coupled receptors.

The multiligand Receptor for Advanced Glycation End products (RAGE) is involved in various pathophysiological processes, including diabetic inflammatory conditions and Alzheimers disease. Full-length RAGE, a cell surface-located type I membrane protein, can proteolytically be converted by metalloproteinases ADAM10 and MMP9 into a soluble RAGE form. Moreover, administration of recombinant soluble RAGE suppresses activation of cell surface-located RAGE by trapping RAGE ligands.

Neuropeptide pituitary adenylate cyclase-activating polypeptide (PACAP) slows down Alzheimer's disease-like pathology in amyloid precursor protein-transgenic mice.

Pituitary adenylate cyclase-activating polypeptide (PACAP) has neuroprotective and neurotrophic properties and is a potent α-secretase activator. As PACAP peptides and their specific receptor PAC1 are localized in central nervous system areas affected by Alzheimer's disease (AD), this study aims to examine the role of the natural peptide PACAP as a valuable approach in AD therapy. We investigated the effect of PACAP in the brain of an AD transgenic mouse model.

Photo-DHEA--a functional photoreactive dehydroepiandrosterone (DHEA) analog.

The steroid hormone dehydroepiandrosterone (DHEA) has beneficial effects on vascular function, survival of neurons, and fatty acid metabolism. However, a specific receptor for DHEA has not been identified to date. Here, we describe the synthesis of a photoreactive DHEA derivative (Photo-DHEA).

Statins and the squalene synthase inhibitor zaragozic acid stimulate the non-amyloidogenic pathway of amyloid-beta protein precursor processing by suppression of cholesterol synthesis.

Cholesterol-lowering drugs such as statins influence the proteolytic processing of the amyloid-beta protein precursor (AbetaPP) and are reported to stimulate the activity of alpha-secretase, the major preventive secretase of Alzheimer's disease. Statins can increase the alpha-secretase activity by their cholesterol-lowering properties as well as by impairment of isoprenoids synthesis. In the present study, we elucidate the contribution of these pathways in alpha-secretase activation.

Influence of ADAM10 on prion protein processing and scrapie infectiosity in vivo.

Both the cellular prion protein (PrP(c)) and the amyloid precursor protein (APP) are physiologically subjected to complex proteolytic processing events. While for APP the proteinases involved--alpha-, beta- and gamma-secretase--have been identified in vitro and in vivo, the cleavage of PrP(c) by now has been linked only to the shedding activity of the metalloproteinase ADAM10 and/or ADAM17 in cell culture. Here we show that neuronal overexpression of the alpha-secretase ADAM10 in mice reduces all PrP(c) species detected in the brain instead of leading to enhanced amounts of specific cleavage products of PrP(c).

Regulated proteolysis of RAGE and AbetaPP as possible link between type 2 diabetes mellitus and Alzheimer's disease.

Epidemiological studies have linked type 2 diabetes mellitus (T2DM) with an increased risk of developing Alzheimer's disease (AD). In T2DM, the elevated blood glucose level promotes formation of advanced glycation end products (AGEs). The receptor for AGEs (RAGE) is a type I membrane-protein and is also able to import amyloid-beta (Abeta) from the blood across the blood-brain-barrier into the brain.

CSF APPs alpha and phosphorylated tau protein levels in mild cognitive impairment and dementia of Alzheimer's type.

We exploratively measured APPs alpha, a secreted fragment of the non-amyloidogenic cleavage of amyloid precursor protein via a-secretase, and tau protein phosphorylated at threonine 181 (p tau) in the cerebrospinal fluid of 10 patients with mild cognitive impairment, 20 patients with dementia of Alzheimer's type, and 10 controls. Cerebrospinal fluid APPs alpha and p tau levels were correlated with cognitive performance. P tau levels were significantly elevated in mild cognitive impairment and in patients with dementia of Alzheimer's type, APPs alpha levels were significantly reduced in patients with dementia of Alzheimer's type compared to the controls.

Receptor for advanced glycation end products is subjected to protein ectodomain shedding by metalloproteinases.

The receptor for advanced glycation end products (RAGE) is a 55-kDa type I membrane glycoprotein of the immunoglobulin superfamily. Ligand-induced up-regulation of RAGE is involved in various pathophysiological processes, including late diabetic complications and Alzheimer disease. Application of recombinant soluble RAGE has been shown to block RAGE-mediated pathophysiological conditions.

The neuropeptide PACAP promotes the alpha-secretase pathway for processing the Alzheimer amyloid precursor protein.

The neuropeptide pituitary adenylate cyclase-activating polypeptide (PACAP) has neurotrophic as well as anti-apoptotic properties and is involved in learning and memory processes. Its specific G protein-coupled receptor PAC1 is expressed in several central nervous system (CNS) regions, including the hippocampal formation. Here we examined the effect of PAC1 receptor activation on alpha-secretase cleavage of the amyloid precursor protein (APP) and the production of secreted APP (APPsalpha).

The non-amyloidogenic pathway: structure and function of alpha-secretases.

The amyloid cascade hypothesis is the most accepted explanation for the pathogenesis of Alzheimer's disease (AD). APP is the precursor of the amyloid beta peptide (Abeta), the principal proteinaceous component of amyloid plaques in brains of Alzheimer's disease patients. Proteolytic cleavage of APP by the alpha-secretase within the Abeta sequence precludes formation of amyloidogenic peptides and leads to a release of soluble APPsalpha which has neuroprotective properties.

A disintegrin-metalloproteinase prevents amyloid plaque formation and hippocampal defects in an Alzheimer disease mouse model.

Alzheimer disease (AD) is characterized by excessive deposition of amyloid beta-peptides (A beta peptides) in the brain. In the nonamyloidogenic pathway, the amyloid precursor protein (APP) is cleaved by the alpha-secretase within the A beta peptide sequence. Proteinases of the ADAM family (adisintegrin and metalloproteinase) are the main candidates as physiologically relevant alpha-secretases, but early lethality of knockout animals prevented a detailed analysis in neuronal cells.

Tumor necrosis factor-alpha converting enzyme is processed by proprotein-convertases to its mature form which is degraded upon phorbol ester stimulation.

Tumor necrosis factor-alpha converting enzyme (TACE or ADAM17) is a member of the ADAM (a disintegrin and metalloproteinase) family of type I membrane proteins and mediates the ectodomain shedding of various membrane-anchored signaling and adhesion proteins. TACE is synthesized as an inactive zymogen, which is subsequently proteolytically processed to the catalytically active form. We have identified the proprotein-convertases PC7 and furin to be involved in maturation of TACE.

Low cholesterol stimulates the nonamyloidogenic pathway by its effect on the alpha -secretase ADAM 10.

Biochemical, epidemiological, and genetic findings demonstrate a link between cholesterol levels, processing of the amyloid precursor protein (APP), and Alzheimer's disease. In the present report, we identify the alpha-secretase ADAM 10 (a disintegrin and metalloprotease) as a major target of the cholesterol effects on APP metabolism. Treatment of various peripheral and neural cell lines with either the cholesterol-extracting agent methyl-beta-cyclodextrin or the hydroxymethyl glutaryl-CoA reductase inhibitor lovastatin resulted in a drastic increase of secreted alpha-secretase cleaved soluble APP.

Alpha-secretase activity of the disintegrin metalloprotease ADAM 10. Influences of domain structure.

Disintegrin metalloproteases from different organisms form the ADAM (a disintegrin and metalloprotease) family. All members display a common domain organization and possess four potential functions: proteolysis, cell adhesion, cell fusion, and cell signaling. Members of the ADAM family are responsible for the proteolytic cleavage of transmembrane proteins and release of their extracellular domain.

Structural requirements for V2 vasopressin receptor proteolytic cleavage.

The ligand-induced proteolytic cleavage of the V2 vasopressin receptor transiently expressed in COS cells was investigated. After incubation of the cell membranes with a photoreactive ligand possessing full agonistic properties for V2 receptors, approximately 90% of the porcine and bovine V2 vasopressin receptors were cleaved in the upper part of transmembrane helix 2 at a heptapeptide sequence conserved in both vasopressin and oxytocin receptors. The oxytocin receptor was completely resistant to proteolysis after binding the same photoreactive ligand, which is only a partial agonist for this receptor.

Direct identification of the agonist binding site in the human brain cholecystokininB receptor.

In investigating the agonist binding site of the human brain cholecystokininB receptor (CCKBR), we employed the direct protein chemical approach using a photoreactive tritiated analogue of sulfated cholecystokinin octapeptide, which contains the p-benzoylbenzoyl moiety at the N-terminus, followed by purification of the affinity-labeled receptor to homogeneity. This probe bound specifically, saturably, and with high affinity (KD = 1.2 nM) to the CCKBR and has full agonistic activity.

Constitutive and regulated alpha-secretase cleavage of Alzheimer's amyloid precursor protein by a disintegrin metalloprotease.

Amyloid beta peptide (Abeta), the principal proteinaceous component of amyloid plaques in brains of Alzheimer's disease patients, is derived by proteolytic cleavage of the amyloid precursor protein (APP). Proteolytic cleavage of APP by a putative alpha-secretase within the Abeta sequence precludes the formation of the amyloidogenic peptides and leads to the release of soluble APPsalpha into the medium. By overexpression of a disintegrin and metalloprotease (ADAM), classified as ADAM 10, in HEK 293 cells, basal and protein kinase C-stimulated alpha-secretase activity was increased severalfold.

Identification of neurohypophysial hormone receptor domains involved in ligand binding and G protein coupling.

Chimeric vasopressin V2/OT receptors were constructed and investigated to identify receptor regions involved in ligand binding or G protein coupling. The fusion sites for one series of hybrid receptors were either located at the C-terminal end of the third extracellular domain or in the centre of the third transmembrane helix, respectively. In each pair of the resulting symmetrical hybrids only one receptor was able to bind arginine vasopressin (AVP) and/or oxytocin (OT).

Identification of binding domains of pituitary adenylate cyclase activating polypeptide (PACAP) for its type 1 receptor by photoaffinity labeling.

Structure-function studies and photoaffinity labeling experiments were performed to identify residues and domains of PACAP involved in the interaction with PACAP receptors. For this purpose, a series of photoreactive analogues of PACAP(1-27) containing a photoreactive benzophenone (BP) residue in different peptide structural domains were utilized to analyze the interaction of PACAP(1-27) with pig PACAP type 1 receptors. Five PACAP derivatives were created with a photoreactive amino acid in the following peptide domains: either the disordered N-terminal or the helical C-terminal domain or a short loop region within the C-terminal helical domain of the peptide.

Photoaffinity labeling analysis of the interaction of pituitary adenylate-cyclase-activating polypeptide (PACAP) with the PACAP type I receptor.

To identify residues and domains of the peptide hormone pituitary adenylate-cyclase-activating polypeptide (PACAP) that interact with the type I receptor, two photoreactive analogues of PACAP-(1-27)-peptide were synthesized using solid-phase peptide synthesis. Phe6 or Tyr22 within the PACAP sequence were replaced by p-benzoyl-L-phenylalanine (Bz-Phe) thus creating two PACAP derivatives with a photoreactive amino acid in either the disordered N-terminal or the helical C-terminal part of the peptide. The ligand-binding properties and the efficiencies of these peptide analogues as photolabels were tested for pig brain PACAP receptors.

Separate agonist and peptide antagonist binding sites of the oxytocin receptor defined by their transfer into the V2 vasopressin receptor.

The neurohypophyseal nonapeptide oxytocin (OT) is the main hormone responsible for the initiation of labor; uterus contraction can be enhanced by application of oxytocin or suppressed by oxytocin antagonists. By transfer of domains from the G protein-coupled OT receptor into the related V2 vasopressin receptor, chimeric "gain in function" V2/OT receptors were produced that were able to bind either OT receptor agonists or a competitive peptide antagonist with high affinity. The binding site for the OT antagonist d(CH2)5[Tyr(Me)2,Thr4,Orn8,Tyr9]vasotocin was found to be formed by transmembrane helices 1, 2, and 7 with a major contribution to binding affinity by the upper part of helix 7.

Reactivity of the N-terminal cysteine residues in active and inactive forms of FNR, and O2-responsive, Fe containing transcriptional regulator of Escherichia coli.

FNR, the O2-responsive gene regulator of anaerobic respiratory genes in Escherichia coli, contains an N-terminal cluster of four cysteine residues (Cys16-X3-Cys20-X2-Cys23-X5-Cys29), three of which are thought to be involved in the binding of an iron cofactor. The accessibility of the cysteine residues for iodoacetate is known to increase upon switch from the active (anaerobic) to the inactive (aerobic or metal depleted) state. It was analyzed which residues become accessible under either condition.

Ligand-induced cleavage of the V2 vasopressin receptor by a plasma membrane metalloproteinase.

The proteolytic cleavage of a G protein-coupled peptide hormone receptor, the renal V2 vasopressin receptor, by a plasma membrane proteinase was investigated. In the absence of protease inhibitors during incubation of bovine kidney membranes with a photoreactive vasopressin agonist, V2 receptor truncation leads to a labeled receptor fragment with M(r) 30,000. The V2 receptor-degrading enzyme could be completely inhibited by zinc ions yielding the native V2 receptor glycoprotein with M(r) 58,000.