[Show abstract][Hide abstract] ABSTRACT: Abstract As a normal consequence of aging, men experience a significant decline in androgen levels. Although the neural consequences of age-related androgen depletion remain unclear, recent evidence suggests a link between low androgen levels and the development of Alzheimer's disease (AD). Here, we test the hypothesis that androgens act as endogenous modulators of β-amyloid protein (Aβ) levels. To investigate this possibility, brain and plasma levels of Aβ were measured in male rats with varying hormonal conditions. Depletion of endogenous sex steroid hormones via gonadectomy (GDX) resulted in increased brain levels of Aβ in comparison to gonadally intact male rats. This GDX-induced increase in Aβ levels was reversed by DHT supplementation, demonstrating a functional role for androgens in modulating brain levels of Aβ. These findings suggest that age-related androgen depletion may result in accumulation of Aβ in the male brain and thereby act as a risk factor for the development of AD.
Journal of Neurochemistry 11/2004; 87(4):1052-1055. · 3.97 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: As a normal consequence of aging, men experience a significant decline in androgen levels. Although the neural consequences of age-related androgen depletion remain unclear, recent evidence suggests a link between low androgen levels and the development of Alzheimer's disease (AD). Here, we test the hypothesis that androgens act as endogenous modulators of beta-amyloid protein (Abeta) levels. To investigate this possibility, brain and plasma levels of Abeta were measured in male rats with varying hormonal conditions. Depletion of endogenous sex steroid hormones via gonadectomy (GDX) resulted in increased brain levels of Abeta in comparison to gonadally intact male rats. This GDX-induced increase in Abeta levels was reversed by DHT supplementation, demonstrating a functional role for androgens in modulating brain levels of Abeta. These findings suggest that age-related androgen depletion may result in accumulation of Abeta in the male brain and thereby act as a risk factor for the development of AD.
Journal of Neurochemistry 12/2003; 87(4):1052-5. · 3.97 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: ErbB-4 is a transmembrane receptor tyrosine kinase that regulates cell proliferation and differentiation. After binding of its ligand heregulin (HRG) or activation of protein kinase C (PKC) by 12-O-tetradecanoylphorbol-13-acetate (TPA), the ErbB-4 ectodomain is cleaved by a metalloprotease. We now report a subsequent cleavage by gamma-secretase that releases the ErbB-4 intracellular domain from the membrane and facilitates its translocation to the nucleus. gamma-Secretase cleavage was prevented by chemical inhibitors or a dominant negative presenilin. Inhibition of gamma-secretase also prevented growth inhibition by HRG. gamma-Secretase cleavage of ErbB-4 may represent another mechanism for receptor tyrosine kinase-mediated signaling.
[Show abstract][Hide abstract] ABSTRACT: Epidemiological studies have documented a reduced prevalence of Alzheimer's disease among users of nonsteroidal anti-inflammatory drugs (NSAIDs). It has been proposed that NSAIDs exert their beneficial effects in part by reducing neurotoxic inflammatory responses in the brain, although this mechanism has not been proved. Here we report that the NSAIDs ibuprofen, indomethacin and sulindac sulphide preferentially decrease the highly amyloidogenic Abeta42 peptide (the 42-residue isoform of the amyloid-beta peptide) produced from a variety of cultured cells by as much as 80%. This effect was not seen in all NSAIDs and seems not to be mediated by inhibition of cyclooxygenase (COX) activity, the principal pharmacological target of NSAIDs. Furthermore, short-term administration of ibuprofen to mice that produce mutant beta-amyloid precursor protein (APP) lowered their brain levels of Abeta42. In cultured cells, the decrease in Abeta42 secretion was accompanied by an increase in the Abeta(1-38) isoform, indicating that NSAIDs subtly alter gamma-secretase activity without significantly perturbing other APP processing pathways or Notch cleavage. Our findings suggest that NSAIDs directly affect amyloid pathology in the brain by reducing Abeta42 peptide levels independently of COX activity and that this Abeta42-lowering activity could be optimized to selectively target the pathogenic Abeta42 species.
[Show abstract][Hide abstract] ABSTRACT: Presenilin 1 (PS1) is linked with Alzheimer's disease but exhibits functional roles regulating growth and development. For instance, PS1 binds to beta-catenin and modulates beta-catenin signaling. In the current study, we observed that knockout of PS1 inhibited beta-catenin-mediated transcription by 35%, as shown by a luciferase reporter driven by the hTcf-4 promoter. Overexpressing wild-type PS1 increased beta-catenin-mediated transcription by 37.5%, and overexpressing PS1 with mutations associated with Alzheimer's disease decreased beta-catenin-mediated transcription by 66%. To examine whether regulation of beta-catenin by PS1 requires phosphorylation by glycogen synthase kinase 3beta (GSK 3beta), we examined whether inhibiting GSK 3beta activity overcomes the inhibition of beta-catenin transcription induced by mutant PS1 constructs. Cells expressing wild-type or mutant PS1 were treated with LiCl, which inhibits GSK 3beta, or transfected with beta-catenin constructs that lack the GSK 3beta phosphorylation sites. Neither treatment overcame PS1-mediated inhibition of beta-catenin signaling, suggesting that regulation of beta-catenin by PS1 was not affected by the activity of GSK 3beta. To investigate how PS1 might regulate beta-catenin signaling, we determined whether PS1 interacts with other elements of the beta-catenin signaling cascade, such as the Tcf-4 transcription factor. Coimmunoprecipitation studies showed binding of PS1 and hTcf-4, and examining nuclear isolates indicated that nuclear hTcf-4 was decreased in cells expressing mutant PS1. These data show that PS1 interacts with multiple components of the beta-catenin signaling cascade and suggest that PS1 regulates beta-catenin in a manner independent of GSK 3beta activity.
Journal of Biological Chemistry 11/2001; 276(42):38563-9. · 4.65 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: In order to develop transgenic animal models that selectively overexpress various Abeta peptides, we have developed a novel expression system that selectively expresses Abeta40 or Abeta42 in the secretory pathway. This system utilizes fusion constructs in which the sequence encoding the 23-amino-acid ABri peptide at the carboxyl terminus of the 266-amino-acid type 2 transmembrane protein BRI is replaced with a sequence encoding either Abeta40 or Abeta42. Constitutive processing of the resultant BRI-Abeta fusion proteins in transfected cells results in high-level expression and secretion of the encoded Abeta peptide. Significantly, expression of Abeta42 from the BRI-Abeta42 construct resulted in no increase in secreted Abeta40, suggesting that the majority of Abeta42 is not trimmed by carboxypeptidase to Abeta40 in the secretory pathway.
Biochimica et Biophysica Acta 08/2001; 1537(1):58-62. · 4.66 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The amyloid b-protein (Ab) deposited in Alzheimer's disease (AD) is a normally secreted proteolytic product of the amyloid b-protein precursor (APP). Generation of Ab from the APP requires two sequential proteolytic events: an initial b-secretase cleavage at the amino terminus of the Ab sequence followed by g-secretase cleavage at the carboxyl terminus of Ab. We describe the development of a robust in vitro assay for g-secretase cleavage by showing de novo Ab production in vitro and establish that this assay monitors authentic gamma-secretase activity by documenting the production of a cognate g-CTF, confirming the size of the Ab produced by mass spectrometry, and inhibiting cleavage in this system with multiple inhibitors that alter g-secretase activity in living cells. Using this assay, we demonstrate that the g-secretase activity 1) is tightly associated with the membrane, 2) can be solubilized, 3) has a pH optimum of 6.8 but is active from pH 6.0 to pH >8.4, and 4) ascertain that activities of the g-40 and g-42 are indeed pharmacologically distinct. These studies should facilitate the purification of the protease or proteases that are responsible for this unusual activity, which is a major therapeutic target for the treatment of AD.
The FASEB Journal 01/2001; 14(15):2383-6. · 5.70 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Presenilins (PSs) are polytopic membrane proteins that have been implicated as potential therapeutic targets in Alzheimer's disease because of their role in regulating the gamma-secretase cleavage that generates the amyloid beta protein (Abeta). It is not clear how PSs regulate gamma-secretase cleavage, but there is evidence that PSs could be either essential cofactors in the gamma-secretase cleavage, gamma-secretase themselves, or regulators of intracellular trafficking that indirectly influence gamma-secretase cleavage. Using presenilin 1 (PS1) mutants that inhibit Abeta production in conjunction with transmembrane domain mutants of the amyloid protein precursor that are cleaved by pharmacologically distinct gamma-secretases, we show that PS1 regulates multiple pharmacologically distinct gamma-secretase activities as well as inducible alpha-secretase activity. It is likely that PS1 acts indirectly to regulate these activities (as in a trafficking or chaperone role), because these data indicate that for PS1 to be gamma-secretase it must either have multiple active sites or exist in a variety of catalytically active forms that are altered to an equivalent extent by the mutations we have studied.
Journal of Biological Chemistry 09/2000; 275(34):26277-84. · 4.65 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: gamma-Secretase catalyzes the cleavage at the carboxyl terminus of A beta to release it from the APP. While gamma-secretase is a major therapeutic drug target for the treatment of Alzheimer's disease (AD), it appears to be an unusual proteolytic activity, and, to date, no protease responsible for this activity has been identified. Based on studies of APP transmembrane domain (TMD) mutants, it is apparent that there are multiple pharmacologically distinct gamma-secretase activities that are spatially restricted and that presenilins (PS) regulate cleavage by gamma-secretases in a protease independent fashion. Based on these studies, we propose a multiprotease model for gamma-secretase activity and predict that the gamma-secretases are likely to be closely related proteases.
Annals of the New York Academy of Sciences 02/2000; 920:233-40. · 4.38 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: gamma-Secretase activity is the final cleavage event that releases the amyloid beta peptide (Abeta) from the beta-secretase cleaved carboxyl-terminal fragment of the amyloid beta protein precursor (APP). No protease responsible for this highly unusual, purportedly intramembranous, cleavage has been definitively identified. We examined the substrate specificity of gamma-secretase by mutating various residues within or adjacent to the transmembrane domain of the APP and then analyzing Abeta production from cells transfected with these mutant APPs by enzyme-linked immunosorbent assay and mass spectrometry. Abeta production was also analyzed from a subset of transmembrane domain APP mutants that showed dramatic shifts in gamma-secretase cleavage in the presence or absence of pepstatin, an inhibitor of gamma-secretase activity. These studies demonstrate that gamma-secretase's cleavage specificity is primarily determined by location of the gamma-secretase cleavage site of APP with respect to the membrane, and that gamma-secretase activity is due to the action of multiple proteases exhibiting both a pepstatin- sensitive activity and a pepstatin-insensitive activity. Given that gamma-secretase is a major therapeutic target in Alzheimer's disease these studies provide important information with respect to the mechanism of Abeta production that will direct efforts to isolate the gamma-secretases and potentially to develop effective therapeutic inhibitors of pathologically relevant gamma-secretase activities.
Journal of Biological Chemistry 05/1999; 274(17):11914-23. · 4.65 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Presenilins mediate an unusual intramembranous proteolytic activity known as gamma-secretase, two substrates of which are the Notch receptor (Notch) and the beta-amyloid precursor protein (APP). gamma-Secretase-mediated cleavage of APP, like that of Notch, yields an intracellular fragment [APP intracellular domain (AICD)1 that forms a transcriptively active complex. We now demonstrate a functional role for AICD in regulating phosphoinositide-mediated calcium signaling. Genetic ablation of the presenilins or pharmacological inhibition of gamma-secretase activity (and thereby AICD production) attenuated calcium signaling in a dose-dependent and reversible manner through a mechanism involving the modulation of endoplasmic reticulum calcium stores. Cells lacking APP (and hence AICD) exhibited similar calcium signaling deficits, and-notably-these disturbances could be reversed by transfection with APP constructs containing an intact AICD, but not by constructs lacking this domain. Our findings indicate that the AICD regulates phosphoinositide- mediated calcium signaling through a gamma-secretase-dependent signaling pathway, suggesting that the intramembranous proteolysis of APP may play a signaling role analogous to that of Notch.
Proceedings of the National Academy of Sciences of the United States of America, v.99, 4697-4702 (2002).