γ-Secretase activity requires the presenilin-dependent trafficking of nicastrin through the Golgi apparatus but not its complex glycosylation

Herreman, An; Gassen, Geert Van; Bentahir, Mustapha; Nyabi, Omar; Craessaerts, Katleen; Mueller, Ulrike; Annaert, Wim; Strooper, Bart De

doi:10.1242/jcs.00292

Cited by 176 publications

(214 citation statements)

References 57 publications

(76 reference statements)

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“…As expected, and as reported previously (17,18), compared with vehicle-treated cells, kifunensine treatment affected neither the levels of APP-carboxylterminal fragments (CTFs) nor levels of secreted Aβ peptides (Fig. 1A, APP-CTFs and Aβ; compare lanes 3 and 4 with 5 and 6, respectively), suggesting that kifunensine-mediated inhibition of oligosaccharide maturation does not impair γ-secretase activity.…”

supporting

confidence: 89%

Identification of a tetratricopeptide repeat-like domain in the nicastrin subunit of γ-secretase using synthetic antibodies

Zhang¹,

Hoey

Lin³

et al. 2012

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

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The γ-secretase complex, composed of presenilin, anterior-pharynxdefective 1, nicastrin, and presenilin enhancer 2, catalyzes the intramembranous processing of a wide variety of type I membrane proteins, including amyloid precursor protein (APP) and Notch. Earlier studies have revealed that nicastrin, a type I membrane-anchored glycoprotein, plays a role in γ-secretase assembly and trafficking and has been proposed to bind substrates. To gain more insights regarding nicastrin structure and function, we generated a conformationspecific synthetic antibody and used it as a molecular probe to map functional domains within nicastrin ectodomain. The antibody bound to a conformational epitope within a nicastrin segment encompassing residues 245-630 and inhibited the processing of APP and Notch substrates in in vitro γ-secretase activity assays, suggesting that a functional domain pertinent to γ-secretase activity resides within this region. Epitope mapping and database searches revealed the presence of a structured segment, located downstream of the previously identified DAP domain (DYIGS and peptidase; residues 261-502), that is homologous to a tetratricopeptide repeat (TPR) domain commonly involved in peptide recognition. Mutagenesis analyses within the predicted TPR-like domain showed that disruption of the signature helical structure resulted in the loss of γ-secretase activity but not the assembly of the γ-secretase and that Leu571 within the TPR-like domain plays an important role in mediating substrate binding. Taken together, these studies offer provocative insights pertaining to the structural basis for nicastrin function as a "substrate receptor" within the γ-secretase complex.A lzheimer's disease (AD), a progressive neurodegenerative disease, is the most prevalent cause of dementia in humans. The principal neuropathological hallmark of AD is the presence of senile plaques composed of dystrophic neurites surrounding extracellular aggregates of Aβ peptides (1). Aβ peptides are liberated from amyloid precursor proteins (APP) by the concerted action of β-site APP cleaving enzyme 1 and γ-secretase (2, 3). γ-Secretase is a macromolecular complex consisting of presenilin 1 or presenilin 2 (PS1 or PS2), anterior-pharynx-defective 1 (APH-1), nicastrin (NCT), and presenilin enhancer 2 (PEN-2) that catalyzes intramembranous proteolysis of several membrane-tethered substrates (4). Evidence has emerged to reveal the functions of each subunit: PS is the catalytic subunit (5); APH-1 serves as a scaffold for the complex assembly (6); NCT seems to be responsible for substrate binding (7); and PEN-2 promotes endoproteolytic cleavage of PS and "activation" of the enzyme complex (5, 8). Although γ-secretase cleaves a multiplicity of substrates at heterogeneous sites within individual transmembrane domains, the molecular mechanism(s) underlying substrate recognition and processing remain elusive.NCT, a 709-aa type I transmembrane glycoprotein with a large, heavily glycosylated ectodomain (ECD), was first identified as a PS-inter...

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supporting

confidence: 89%

Identification of a tetratricopeptide repeat-like domain in the nicastrin subunit of γ-secretase using synthetic antibodies

Zhang¹,

Hoey

Lin³

et al. 2012

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

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“…In HsNCT, there are 16 potential N-glycosylation sites. The glycosylation of HsNCT is reported to be important for the formation and stabilization of the γ-secretase complex, although glycosylation may not be essential for the protease activity of γ-secretase (32,33). Consistent with this conclusion, the potential N-glycosylation sites of nicastrin are not conserved across different organisms.…”

Section: Discussionmentioning

confidence: 88%

Crystal structure of the γ-secretase component nicastrin

Xie

Yan

Zhou

et al. 2014

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

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γ-Secretase is an intramembrane protease responsible for the generation of amyloid-β (Aβ) peptides. Aberrant accumulation of Aβ leads to the formation of amyloid plaques in the brain of patients with Alzheimer's disease. Nicastrin is the putative substrate-recruiting component of the γ-secretase complex. No atomicresolution structure had been identified on γ-secretase or any of its four components, hindering mechanistic understanding of γ-secretase function. Here we report the crystal structure of nicastrin from Dictyostelium purpureum at 1.95-Å resolution. The extracellular domain of nicastrin contains a large lobe and a small lobe. The large lobe of nicastrin, thought to be responsible for substrate recognition, associates with the small lobe through a hydrophobic pivot at the center. The putative substrate-binding pocket is shielded from the small lobe by a lid, which blocks substrate entry. These structural features suggest a working model of nicastrin function. Analysis of nicastrin structure provides insights into the assembly and architecture of the γ-secretase complex.

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“…Studies have shown that PEN-2 and APH-1 may be involved in coordinately regulating proteolytic processing of PS1 (34,37). Nicastrin has been shown to form a complex with PS1 (38), and its presence is required both for the cleavage of PS1 into its active C-and N-terminal fragments and for its transport beyond the ER (39,40). Likewise, in the absence of PS1, NCT is not transported suggesting that interactions between these proteins are needed for them to either fold correctly, making it possible to exit the ER, or perhaps to mask an ER retention signal (27)(28)(29).…”

Section: Discussionmentioning

confidence: 99%

The Transmembrane Domain Region of Nicastrin Mediates Direct Interactions with APH-1 and the γ-Secretase Complex

Morais

Crystal

Pijak

et al. 2003

Journal of Biological Chemistry

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Nicastrin (NCT) is a type I integral membrane protein that is one of the four essential components of the ␥-secretase complex, a protein assembly that catalyzes the intramembranous cleavage of the amyloid precursor protein and Notch. Other ␥-secretase components include presenilin-1 (PS1), APH-1, and PEN-2, all of which span the membrane multiple times. The mechanism by which NCT associates with the ␥-secretase complex and regulates its activity is unclear. To avoid the misfolding phenotype often associated with introducing deletions or mutations into heavily glycosylated and disulfidebonded proteins such as NCT, we produced chimeras between human (hNCT) and Caenorhabditis elegans NCT (ceNCT). Although ceNCT did not associate with human ␥-secretase components, all of the ceNCT/hNCT chimeras interacted with ␥-secretase components from human, C. elegans, or both, indicating that they folded correctly. A region at the C-terminal end of hNCT, encompassing the last 50 residues of its ectodomain, the transmembrane domain, and the cytoplasmic domain was important for mediating interactions with human PS1, APH-1, and PEN-2. This finding is consistent with the fact that the bulk of the ␥-secretase complex proteins resides within the membrane, with relatively small extramembranous domains. Finally, hNCT associated with hAPH-1 in the absence of PS, consistent with NCT and APH-1 forming a subcomplex prior to association with PS1 and PEN-2 and indicating that the interactions between NCT with PS1 may be indirect or stabilized by the presence of APH-1.

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γ-Secretase activity requires the presenilin-dependent trafficking of nicastrin through the Golgi apparatus but not its complex glycosylation

Cited by 176 publications

References 57 publications

Identification of a tetratricopeptide repeat-like domain in the nicastrin subunit of γ-secretase using synthetic antibodies

Identification of a tetratricopeptide repeat-like domain in the nicastrin subunit of γ-secretase using synthetic antibodies

Crystal structure of the γ-secretase component nicastrin

The Transmembrane Domain Region of Nicastrin Mediates Direct Interactions with APH-1 and the γ-Secretase Complex

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