Differential gene expression in ADAM10 and mutant ADAM10 transgenic mice

<p>Abstract</p> <p>Background</p> <p>In a transgenic mouse model of Alzheimer disease (AD), cleavage of the amyloid precursor protein (APP) by the α-secretase ADAM10 prevented amyloid plaque formation, and alleviated cognitive deficits. Furthermore, ADAM10 overexpression increased the cortical synaptogenesis. These results suggest that upregulation of ADAM10 in the brain has beneficial effects on AD pathology.</p> <p>Results</p> <p>To assess the influence of ADAM10 on the gene expression profile in the brain, we performed a microarray analysis using RNA isolated from brains of five months old mice overexpressing either the α-secretase ADAM10, or a dominant-negative mutant (dn) of this enzyme. As compared to non-transgenic wild-type mice, in ADAM10 transgenic mice 355 genes, and in dnADAM10 mice 143 genes were found to be differentially expressed. A higher number of genes was differentially regulated in double-transgenic mouse strains additionally expressing the human APP_[V717I]mutant.</p> <p>Overexpression of proteolytically active ADAM10 affected several physiological pathways, such as cell communication, nervous system development, neuron projection as well as synaptic transmission. Although ADAM10 has been implicated in Notch and β-catenin signaling, no significant changes in the respective target genes were observed in adult ADAM10 transgenic mice.</p> <p>Real-time RT-PCR confirmed a downregulation of genes coding for the inflammation-associated proteins S100a8 and S100a9 induced by moderate ADAM10 overexpression. Overexpression of the dominant-negative form dnADAM10 led to a significant increase in the expression of the fatty acid-binding protein Fabp7, which also has been found in higher amounts in brains of Down syndrome patients.</p> <p>Conclusion</p> <p>In general, there was only a moderate alteration of gene expression in ADAM10 overexpressing mice. Genes coding for pro-inflammatory or pro-apoptotic proteins were not over-represented among differentially regulated genes. Even a decrease of inflammation markers was observed. These results are further supportive for the strategy to treat AD by increasing the α-secretase activity.</p

Ectodomain shedding of L1 adhesion molecule promotes cell migration by autocrine binding to integrins

The L1 adhesion molecule plays an important role in axon guidance and cell migration in the nervous system. L1 is also expressed by many human carcinomas. In addition to cell surface expression, the L1 ectodomain can be released by a metalloproteinase, but the biological function of this process is unknown. Here we demonstrate that membrane-proximal cleavage of L1 can be detected in tumors and in the developing mouse brain. The shedding of L1 involved a disintegrin and metalloproteinase (ADAM)10, as transfection with dominant-negative ADAM10 completely abolishes L1 release. L1-transfected CHO cells (L1-CHO) showed enhanced haptotactic migration on fibronectin and laminin, which was blocked by antibodies to αvβ5 and L1. Migration of L1-CHO cells, but not the basal migration of CHO cells, was blocked by a metalloproteinase inhibitor, indicating a role for L1 shedding in the migration process. CHO and metalloproteinase-inhibited L1-CHO cells were stimulated to migrate by soluble L1-Fc protein. The induction of migration was blocked by αvβ5-specific antibodies and required Arg-Gly-Asp sites in L1. A 150-kD L1 fragment released by plasmin could also stimulate CHO cell migration. We propose that ectodomain-released L1 promotes migration by autocrine/paracrine stimulation via αvβ5. This regulatory loop could be relevant for migratory processes under physiological and pathophysiological conditions

A closer look at alpha-secretase

Accumulation of amyloid beta-peptides (A beta) in the brain is believed to contribute to the development of Alzheimer disease (AD). A beta, a 40-42 amino acid-comprising proteolytical fragment of the amyloid precursor protein (APP), is released from APP by sequential cleavages via beta- and gamma-secretases. However, the predominant route of APP processing consists of successive cleavages by alpha- and gamma-secretases. Alpha-secretase attacks APP inside the A beta sequence, and therefore prevents formation of neurotoxic A beta. After cleavage by alpha-secretase, the soluble N-terminal domain of APP, which possesses neurotrophic and neuroprotective properties, is released. In AD patients, a decrease in alpha-secretase processing of APP has been found and therefore, strategies to improve alpha-secretase activity are obvious. Several years after descriptive reports on alpha-secretase, the responsible enzymes have been identified to belong to the family of A Disintegrin And Metalloproteinase (ADAM). Three of these membrane-anchored zinc-dependent metalloproteinases, ADAM10 as well as ADAM17 and presumably also ADAM9 display alpha-secretase activity. Since the individual knock-out of these proteinases in neither case completely prevented alpha-secretase processing of APP, it seems likely that different ADAMs are compensating mutually, and under different conditions may contribute to alpha-secretase cleavage of APP. In addition to ADAMs, perhaps other membrane-associated metalloproteinases contribute to the shedding of APP. Stimulation of alpha-secretase activities can be achieved via several signaling cascades including phospholipase C, phosphatidylinositol 3-kinase and serine/threonine-specific kinases such as protein kinases C, and mitogen activated protein kinases. Direct activation of protein kinase C and stimulation of distinct G protein-coupled receptors are known to increase alpha-secretase processing of APP. Agonists for M1 muscarinic acetylcholine receptors and serotonin 5-HT4 receptors are currently in clinical trials to test their efficiency in the treatment of AD

Activation of a-secretase cleavage

Alpha-secretase-mediated cleavage of the amyloid precursor protein (APP) releases the neuroprotective APP fragment saAPP and prevents amyloid beta peptide (A beta) generation. Moreover, a-secretase-like cleavage of the A beta transporter receptor for advanced glycation end products counteracts the import of blood A beta into the brain. Assuming that A beta is responsible for the development of Alzheimers disease (AD), activation of a-secretase should be preventive. a-Secretase-mediated APP cleavage can be activated via several G protein-coupled receptors and receptor tyrosine kinases. Protein kinase C, mitogen-activated protein kinases, phosphatidylinositol 3-kinase, cAMP and calcium are activators of receptor-induced a-secretase cleavage. Selective targeting of receptor subtypes expressed in brain regions affected by AD appears reasonable. Therefore, the PACAP receptor PAC1 and possibly the serotonin 5-HT6 receptor subtype are promising targets. Activation of APP a-secretase cleavage also occurs upon blockade of cholesterol synthesis by statins or zaragozic acid A. Under physiological statin concentrations, the brain cholesterol content is not influenced. Statins likely inhibit A beta production in the blood by a-secretase activation which is possibly sufficient to inhibit AD development. A disintegrin and metalloproteinase 10 (ADAM10) acts as a-secretase on APP. By targeting the nuclear retinoic acid receptor beta, the expression of ADAM10 and non-amyloidogenic APP processing can be enhanced. Excessive activation of ADAM10 should be avoided because ADAM10 and also ADAM17 are not APP-specific. Both ADAM proteins cleave various substrates, and therefore have been associated with tumorigenesis and tumor progression

Regulated proteolysis of rage and a beta pp 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 o

Alpha-secretase activation : an approach to Alzheimer's disease therapy

The nonamyloidogenic pathway of processing the amyloid precursor protein (APP) involves the cleavage within the amyloid-beta peptide sequence, and thus precludes amyloid-beta formation. The identification of a member of the disintegrin and metalloproteinase family, ADAM10, as an alpha-secretase that prevents plaque formation and hippocampal deficits in vivo gave us the possibility to examine the alpha-secretase as a potential target for the therapy of Alzheimer&#039;s disease. Within the priority program Cellular Mechanisms of Alzheimer&#039;s Disease,we investigated several approaches to stimulate the alpha-secretase pathway. Two protein convertases were found to be responsible for the removal of the prodomain, and for the formation of the mature enzyme with alpha-secretase activity. The cloning and characterization of the human ADAM10 promoter provided the basis to examine ADAM10 gene expression. We found a common upregulation of ADAM10, APP, and APP-like protein 2 during differentiation of neuronal cells by retinoic acid, and increased alpha-secretase cleavage of the two substrates. Other approaches for enhancing alpha-secretase activity are the reduction of cellular cholesterol and the stimulation of G protein-coupled neuropeptide receptors. Our results suggest medications and dietary regiments which enhance the nonamyloidogenic pathway of APP processing to be a valuable approach to Alzheimer&#039;s disease therapy

Separate agonist and peptide antagonist binding sites of the oxytocin receptor defined by their transfer into the v-2 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 V-2 vasopressin receptor, chimeric &#039;&#039;gain in function&#039;&#039; V-2/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), Thr(4),Orn(8),Tyr(9)]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. These transmembrane receptor regions could be excluded from participating in OT binding, For agonist binding and selectivity the first three extracellular receptor domains were most important. The interaction of the N-terminal domain and of the first extracellular loop of the OT receptor with the linear C-terminal tripeptidic part of oxytocin was demonstrated, Furthermore, the second extracellular loop of the OT receptor could be identified to interact with the cyclic hormone part. These three domains contribute to OT binding by synergistic interaction with oxytocin but not with the competitive antagonist. Our results provide evidence for the existence of separate domains and different conformations of a peptide hormone receptor involved in binding and selectivity for agonists and peptide antagonists

Ectodomain shedding of the receptor for advanced glycation end products : a novel therapeutic target for Alzheimer's disease

Receptor for advanced glycation end products (RAGE) mediates diverse physiological and pathological effects and is involved in the pathogenesis of Alzheimer&#039;s disease (AD). RAGE is a receptor for amyloid beta peptides (A beta), mediates A beta neurotoxicity and also promotes A beta influx into the brain and contributes to A beta aggregation. Soluble RAGE (sRAGE), a secreted RAGE isoform, acts as a decoy receptor to antagonize RAGE-mediated damages. Accumulating evidence has suggested that sRAGE represents a promising pharmaceutic against RAGE-mediated disorders. Recent studies revealed proteolysis of RAGE as a previously unappreciated means of sRAGE production. In this review we summarize these findings on the proteolytic cleavage of RAGE and discuss the underlying regulatory mechanisms of RAGE shedding. Furthermore, we propose a model in which proteolysis of RAGE could restrain AD development by reducing A beta transport into the brain and A beta production via BACE. Thus, the modulation of RAGE proteolysis provides a novel intervention strategy for AD