The Oligomerization of Amyloid β-Protein Begins Intracellularly in Cells Derived from Human Brain

Walsh, Dominic M.; Tseng, Bertrand P.; Rydel, Russell E.; Podlisny, Marcia B.; Selkoe, Dennis J.

doi:10.1021/bi001048s

Cited by 465 publications

(372 citation statements)

References 39 publications

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“…This finding is consistent with the observation that stable A␤ dimers are found intracellularly in neurons and in vivo in brain (26). Taken together, these results have led to the idea that APP dimerization can positively regulate A␤ production.…”

supporting

confidence: 91%

Induced Dimerization of the Amyloid Precursor Protein Leads to Decreased Amyloid-β Protein Production

Eggert

Midthune

Cottrell

et al. 2009

Journal of Biological Chemistry

142

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The amyloid precursor protein (APP) plays a central role in Alzheimer disease (AD) pathogenesis because sequential cleavages by ␤-and ␥-secretase lead to the generation of the amyloid-␤ (A␤) peptide, a key constituent in the amyloid plaques present in brains of AD individuals. In several studies APP has recently been shown to form homodimers, and this event appears to influence A␤ generation. However, these studies have relied on APP mutations within the A␤ sequence itself that may affect APP processing by interfering with secretase cleavages independent of dimerization. Therefore, the impact of APP dimerization on A␤ production remains unclear. To address this question, we compared the approach of constitutive cysteine-induced APP dimerization with a regulatable dimerization system that does not require the introduction of mutations within the A␤ sequence. To this end we generated an APP chimeric molecule by fusing a domain of the FK506-binding protein (FKBP) to the C terminus of APP. The addition of the synthetic membrane-permeant drug AP20187 induces rapid dimerization of the APP-FKBP chimera. Using this system we were able to induce up to 70% APP dimers. Our results showed that controlled homodimerization of APP-FKBP leads to a 50% reduction in total A␤ levels in transfected N2a cells. Similar results were obtained with the direct precursor of ␤-secretase cleavage, C99/SPA4CT-FKBP. Furthermore, there was no modulation of different A␤ peptide species after APP dimerization in this system. Taken together, our results suggest that APP dimerization can directly affect ␥-secretase processing and that dimerization is not required for A␤ production.The mechanism of ␤-amyloid protein (A␤) 2 generation from the amyloid precursor protein is of major interest in Alzheimer disease research because A␤ is the major constituent of senile plaques, one of the neuropathological hallmarks of Alzheimer disease (1, 2). In the amyloidogenic pathway A␤ is released from the amyloid precursor protein (APP) (3) after sequential cleavages by ␤-secretase BACE1 (4 -6) and by the ␥-secretase complex (7,8). BACE1 cleavage releases the large ectodomain of APP while generating the membrane-anchored C-terminal APP fragment (CTF) of 99 amino acids (C99). Cleavage of ␤-CTF by ␥-secretase leads to the secretion of A␤ peptides of various lengths and the release of the APP intracellular domain (AICD) into the cytosol (9 -11). The ␥-secretase complex consists of at least four proteins: presenilin, nicastrin, Aph-I, and Pen-2 (12). Presenilin is thought to be the catalytic subunit of the enzyme complex (13), but how the intramembrane scission is carried out remains to be elucidated. Alternatively, APP can first be cleaved in the non-amyloidogenic pathway by ␣-secretase within the A␤ domain between Lys-16 and 15). This cleavage releases the APP ectodomain (APPs␣) while generating the membrane-bound C-terminal fragment (␣-CTF) of 83 amino acids (C83). The latter can be further processed by the ␥-secretase complex, resulting in the secretion of the sm...

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supporting

confidence: 91%

Induced Dimerization of the Amyloid Precursor Protein Leads to Decreased Amyloid-β Protein Production

Eggert

Midthune

Cottrell

et al. 2009

Journal of Biological Chemistry

142

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“…We hypothesize that heme binding to the monomer form of A␤ peptides may change the conformation of the peptide, its hydrophobicity, or mask the site that binds free metals (i.e., histidine), thus, inhibiting A␤ oligomerization (20) and keeping intracellular A␤ in monomer form (Scheme 1). Intracellular accumulation of A␤ has been hypothesized to form prefibrillar species (e.g., oligomers) (40), which are resistant to proteolysis. Recent research on AD implicates intracellular prefibrillar species of A␤, rather than the senile plaques or monomers, in neurotoxicity and neuronal dysfunction of AD (41)(42)(43).…”

Section: Discussionmentioning

confidence: 99%

A role for heme in Alzheimer's disease: Heme binds amyloid β and has altered metabolism

Atamna

Frey

2004

Proc. Natl. Acad. Sci. U.S.A.

235

239

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Heme is a common factor linking several metabolic perturbations in Alzheimer's disease (AD), including iron metabolism, mitochondrial complex IV, heme oxygenase, and bilirubin. Therefore, we determined whether heme metabolism was altered in temporal lobes obtained at autopsy from AD patients and age-matched nondemented subjects. AD brain demonstrated 2.5-fold more heme-b (P < 0.01) and 26% less heme-a (P ‫؍‬ 0.16) compared with controls, resulting in a highly significant 2.9-fold decrease in heme-a͞heme-b ratio (P < 0.001). Moreover, the strong Pearson correlation between heme-a and heme-b measured in control individuals (r 2 ‫؍‬ 0.66, P < 0.002, n ‫؍‬ 11) was abolished in AD subjects (r 2 ‫؍‬ 0.076, P ‫؍‬ 0.39, n ‫؍‬ 12). The level of ferrochelatase (which makes heme-b in the mitochondrial matrix) in AD subjects was 4.2 times (P < 0.04) that in nondemented controls, suggesting up-regulated heme synthesis. To look for a possible connection between these observations and established mechanisms in AD pathology, we examined possible interactions between amyloid ␤ (A␤) and heme. A␤(1-40) and A␤(1-42) induced a redshift of 15-20 nm in the spectrum of heme-b and heme-a, suggesting that heme binds A␤, likely to one or more of the histidine residues. Lastly, in a tissue culture model, we found that clioquinol, a metal chelator in clinical trials for AD therapy, decreased intracellular heme. In light of these observations, we have proposed a model of AD pathobiology in which intracellular A␤ complexes with free heme, thereby decreasing its bioavailability (e.g., heme-a) and resulting in functional heme deficiency. The model integrates disparate observations, including A␤, mitochondrial dysfunction, cholesterol, and the proposed efficacy of clioquinol.mitochondria ͉ heme-a ͉ iron ͉ clioquinol ͉ ferrochelatase H eme (ferriprotoporphyrin IX) metabolism appears altered in the brains of Alzheimer's disease (AD) patients. Heme oxygenase (HO) increases in AD (1, 2), and the level of bilirubin (one of the products of heme degradation by HO) is increased in AD patients (3). Mitochondrial complex IV, the only enzyme in cells that contains heme-a (see below) (4), declines in AD (5-8). Heme-a is rate limiting for the assembly of complex IV (9). Furthermore, an inhibitor of muscarinic acetylcholine receptor binding, which increased in AD brain, was suggested to be heme (10, 11). Therefore, we studied heme metabolism in AD brain and nondemented age-matched normal subjects.Heme-b, the product of ferrochelatase (FC) (also known as protoheme), is produced in the mitochondrial matrix (12, 13) and is the precursor for heme-c and heme-a. The structure of heme-c is similar to that of heme-b, but it is covalently attached to a few specific proteins (reviewed in ref. 14). Heme-b and heme-a exist in two major pools in the cell, free and protein-associated. Conversion of heme-b to heme-a requires farnasylation and oxidation (15). Farnesyl-pyrophsphate (FPP) is the precursor for the farnesyl moiety in heme-a, cholesterol, dolichol, farnesylation ...

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“…6 and 7). Dimers and aggregates of APP and A␤1-42 (or A␤1-40) have been demonstrated in brain cells (41,42). APP dimers and aggregates induced by heme deficiency are likely to be the abnormal forms of APP.…”

Section: Discussionmentioning

confidence: 99%

Heme deficiency may be a factor in the mitochondrial and neuronal decay of aging

Atamna¹,

Killilea²,

Killilea³

et al. 2002

Proc. Natl. Acad. Sci. U.S.A.

199

150

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Heme, a major functional form of iron in the cell, is synthesized in the mitochondria by ferrochelatase inserting ferrous iron into protoporphyrin IX. Heme deficiency was induced with N-methylprotoporphyrin IX, a selective inhibitor of ferrochelatase, in two human brain cell lines, SHSY5Y (neuroblastoma) and U373 (astrocytoma), as well as in rat primary hippocampal neurons. Heme deficiency in brain cells decreases mitochondrial complex IV, activates nitric oxide synthase, alters amyloid precursor protein, and corrupts iron and zinc homeostasis. The metabolic consequences resulting from heme deficiency seem similar to dysfunctional neurons in patients with Alzheimer's disease. Heme-deficient SHSY5Y or U373 cells die when induced to differentiate or to proliferate, respectively. The role of heme in these observations could result from its interaction with heme regulatory motifs in specific proteins or secondary to the compromised mitochondria. Common causes of heme deficiency include aging, deficiency of iron and vitamin B6, and exposure to toxic metals such as aluminum. Iron and B6 deficiencies are especially important because they are widespread, but they are also preventable with supplementation. Thus, heme deficiency or dysregulation may be an important and preventable component of the neurodegenerative process.iron ͉ complex IV ͉ amyloid precursor protein (APP) ͉ nitric oxide synthase ͉ oxidative stress

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The Oligomerization of Amyloid β-Protein Begins Intracellularly in Cells Derived from Human Brain

Cited by 465 publications

References 39 publications

Induced Dimerization of the Amyloid Precursor Protein Leads to Decreased Amyloid-β Protein Production

Induced Dimerization of the Amyloid Precursor Protein Leads to Decreased Amyloid-β Protein Production

A role for heme in Alzheimer's disease: Heme binds amyloid β and has altered metabolism

Heme deficiency may be a factor in the mitochondrial and neuronal decay of aging

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