Mechanism of the cleavage specificity of Alzheimer’s disease γ-secretase identified by phenylalanine-scanning mutagenesis of the transmembrane domain of the amyloid precursor protein

Lichtenthaler, Stefan F.; Wang, Rong; Grimm, Heike S.; Uljon, Sacha N.; Masters, Colin L.; Beyreuther, Konrad

doi:10.1073/pnas.96.6.3053

Cited by 232 publications

(234 citation statements)

References 38 publications

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“…We show that the V 3 G mutation at P1Ј of syndecan 3 even favors ␥-secretase cleavage of syndecan 3 and conclude that valine at P1Ј is neither necessary nor necessarily in favor of processing by ␥-secretase. This result is in agreement with previous studies on APP, showing loose sequence specificity for ␥-cleavage, which seems to be determined by structural properties of the transmembrane domain such as length or ␣-helix content (45)(46)(47).…”

Section: Fig 4 Luciferase Assay To Quantify Sicd Generation In Hek2supporting

confidence: 83%

Syndecan 3 Intramembrane Proteolysis Is Presenilin/γ-Secretase-dependent and Modulates Cytosolic Signaling

Schulz

Annaert

Vandekerckhove

et al. 2003

Journal of Biological Chemistry

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The syndecans play critical roles in several signal transduction pathways. The core proteins of these heparan sulfate proteoglycans are characterized by highly conserved transmembrane and intracellular domains which are required for signaling across the membrane and for interaction with cytosolic proteins. However, regulatory mechanisms controlling these functions remain largely unknown. Here we show that, upon ligandinduced primary proteolytic cleavage within the ectodomain, the intracellular domain of syndecan 3 is released by regulated intramembrane proteolysis. The cleavage is mediated by presenilin/␥-secretase complex and negatively regulates the plasma membrane targeting of the transcriptional cofactor CASK.Heparan sulfate proteoglycans (HSPG) 1 are present on the surface of all adherent cells. Binding to a large number of ligands depends upon highly charged heparan sulfate chains, linear polysaccharide chains that can be modified by sequential enzymatic modifications. These modifications lead to enormous structural diversity (for review see Ref. 1). Although numerous functions for the heparan sulfate moiety of proteoglycans have been shown in vivo and in vitro, the roles of the core proteins remain largely elusive (2, 3). The syndecan family of HSPG, as opposed to the GPI-anchored glypicans or matrix-associated HSPG, contains a transmembrane and cytosolic domain that is highly conserved across homologues and species, suggesting an important role for this part of the protein (4). Indeed, a role in FGF signaling, neurite outgrowth, dendritic spine morphogenesis, and left-right axis formation has been demonstrated (5-8), but the regulatory mechanisms are incompletely understood. The cytosolic domain can be subdivided into a membrane-proximal C1 and a C-terminal C2 conserved region that is nearly identical in all syndecans and has a more variable V region in the middle. The most conserved residues are serines and tyrosines that at least partially undergo phosphorylation, a basic motif that binds nonreceptor tyrosine kinases and cytoskeletal proteins, and the C terminus that is required for interaction with PDZ proteins such as syntenin, CASK, synectin, or synbindin (for review see Ref. 4).The heparan sulfate-bearing syndecan ectodomain has been shown to undergo ligand-activated or stress-induced proteolytic cleavage and shedding (9 -11), but the fate of the Cterminal fragment (CTF) that remains associated with the membrane is unknown. We surmised that the remaining CTF is subject to further proteolytic degradation, ectodomain-shedding being a first step in an intracellular signaling cascade. Because regulated intramembrane proteolysis, a novel mechanism of signal transduction across the membrane (12), has been demonstrated to depend upon prior ectodomain cleavage of type I transmembrane protein substrates (13)(14)(15)(16)(17)(18)(19)(20)(21)(22) and is mediated by presenilins (PS) as part of a larger so-called ␥-secretase complex (23), we investigated whether syndecan 3 undergoes PS-dependent cleavage. EXPERI...

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Section: Fig 4 Luciferase Assay To Quantify Sicd Generation In Hek2supporting

confidence: 83%

Syndecan 3 Intramembrane Proteolysis Is Presenilin/γ-Secretase-dependent and Modulates Cytosolic Signaling

Schulz

Annaert

Vandekerckhove

et al. 2003

Journal of Biological Chemistry

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“…Mutations into the N-terminal 16 residues of C99 were not introduced, because C83-which lacks the N-terminal 16 residues of C99 because of cleavage by ␣-secretase-is cleaved by ␥-secretase at the same site as C99 (15)(16)(17), and, thus, the N-terminal 16 residues of C99 do not influence the ␥-cleavage site. With four mutants, we analyzed possible sequence elements in the ectodomain and the cytoplasmic domain of C99 ( Fig.…”

Section: Resultsmentioning

confidence: 99%

“…COS7 cells were cultured according to the protocol described previously (17). Cell culture media were obtained from Sigma.…”

Section: Methodsmentioning

confidence: 99%

“…The ␥-cleavage seems to occur with broad substrate specificity because A␤ is still generated when residues within the TMD of C99 are mutated (15,(17)(18)(19)(20)(21). In contrast, the specific cleavage site used by ␥-secretase (e.g., cleavage after residue 40 or 42) and thus the C terminus of A␤ strongly depends on the amino acid composition of the TMD (15,17,18,20,21).The proteolytic processing of APP is similar to the processing of the cell surface receptor Notch (for an overview, see ref. 8).…”

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confidence: 99%

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The intramembrane cleavage site of the amyloid precursor protein depends on the length of its transmembrane domain

Lichtenthaler

Beher

Grimm

et al. 2002

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

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Proteolytic processing of the amyloid precursor protein by ␤-secretase generates C99, which subsequently is cleaved by ␥-secretase, yielding the amyloid ␤ peptide (A␤). This ␥-cleavage occurs within the transmembrane domain (TMD) of C99 and is similar to the intramembrane cleavage of Notch. However, Notch and C99 differ in their site of intramembrane cleavage. The main ␥-cleavage of C99 occurs in the middle of the TMD, whereas the cleavage of Notch occurs close to the C-terminal end of the TMD, making it unclear whether both are cleaved by the same protease. To investigate whether ␥-cleavage always occurs in the middle of the TMD of C99 or may also occur at the end of the TMD, we generated C99-mutants with an altered length of the TMD and analyzed their ␥-cleavage in COS7 cells. The C terminus of A␤ and thus the site of ␥-cleavage were determined by using monoclonal antibodies and mass spectrometry. Compared with C99-wild type (wt), most mutants with an altered length of the TMD changed the cleavage site of ␥-secretase, whereas control mutants with mutations outside the TMD did not. Thus, the length of the whole TMD is a major determinant for the cleavage site of ␥-secretase. Moreover, the C99-mutants were not only cleaved at one site but at two sites within their TMD. One cleavage site was located around the middle of the TMD, regardless of its actual length. An additional cleavage occurred within the N-terminal half of their TMD and thus at the opposite side of the Notch cleavage site.R egulated intramembrane proteolysis of type I membrane proteins has been shown to play an important role in the pathogenesis of Alzheimer's disease and in cell differentiation (for a review, see ref. 1). The corresponding proteins involved in these biological processes are the amyloid precursor protein (APP) and Notch, respectively.The intramembrane proteolysis of APP leads to the generation of the amyloid ␤ peptide (A␤), which is deposited in the brains of patients with Alzheimer's disease (for a review, see ref.2). To undergo intramembrane proteolysis, APP is first proteolytically processed by one of the proteases termed ␣-and ␤-secretase (for a review, see ref.3). The ␣-secretase seems to be a member of the ADAM (a disintegrin and metalloprotease)-family of disintegrin-metalloproteases (4-6), whereas the recently identified ␤-secretase is a novel aspartyl protease called BACE (beta-site APP cleaving enzyme; for a review, see ref. 7). The ␣-and ␤-secretases cleave APP within its ectodomain at a short distance to the transmembrane domain (TMD), shearing off most of the ectodomain (secretory APP). Subsequently, the remaining membrane-bound C-terminal fragments C83 (through ␣-cleavage) and C99 (through ␤-cleavage) may undergo intramembrane proteolysis by a protease called ␥-secretase, which has not yet been unequivocally identified. However, the ␥-secretase cleavage depends on the presence of the polytopic membrane protein presenilin 1, which itself may be ␥-secretase (for reviews, see refs. 8 and 9). Because presenilin 1 is part...

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“…γ-Secretase is an intramembranous complex composed of presenilin-1 (PS1), PEN-2, nicastrin and APH-1, with PS1 constituting the active site [11]. Imprecise cleavage of C99 by γ-secretase produces Aβ variants, mainly Aβ40 and Aβ42 [12]. Conversely, cleavage of APP by α-secretase precludes the production www.cell-research.com | Cell Research Lin Teng et al 139…”

Section: Introductionmentioning

confidence: 99%

A GPCR/secretase complex regulates β- and γ-secretase specificity for Aβ production and contributes to AD pathogenesis

et al. 2010

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Dysregulation of β-site APP-cleaving enzyme (BACE) and/or γ-secretase leads to anomalous production of amyloid-β peptide (Aβ) and contributes to the etiology of Alzheimer's disease (AD). Since these secretases mediate proteolytic processing of numerous proteins, little success has been achieved to treat AD by secretase inhibitors because of inevitable undesired side effects. Thus, it is of importance to unravel the regulatory mechanisms of these secretases. Here, we show that δ-opioid receptor (DOR) promotes the processing of Aβ precursor protein (APP) by BACE1 and γ-secretase, but not that of Notch, N-cadherin or APLP. Further investigation reveals that DOR forms a complex with BACE1 and γ-secretase, and activation of DOR mediates the co-endocytic sorting of the secretases/ receptor complex for APP endoproteolysis. Dysfunction of the receptor retards the endocytosis of BACE1 and γ-secretase and thus the production of Aβ. Consistently, knockdown or antagonization of DOR reduces secretase activities and ameliorates Aβ pathology and Aβ-dependent behavioral deficits, but does not affect the processing of Notch, N-cadherin or APLP in AD model mice. Our study not only uncovers a molecular mechanism for the formation of a DOR/secretase complex that regulates the specificity of secretase for Aβ production but also suggests that intervention of either formation or trafficking of the GPCR/secretase complex could lead to a new strategy against AD, potentially with fewer side effects.

show abstract

Mechanism of the cleavage specificity of Alzheimer’s disease γ-secretase identified by phenylalanine-scanning mutagenesis of the transmembrane domain of the amyloid precursor protein

Cited by 232 publications

References 38 publications

Syndecan 3 Intramembrane Proteolysis Is Presenilin/γ-Secretase-dependent and Modulates Cytosolic Signaling

Syndecan 3 Intramembrane Proteolysis Is Presenilin/γ-Secretase-dependent and Modulates Cytosolic Signaling

The intramembrane cleavage site of the amyloid precursor protein depends on the length of its transmembrane domain

A GPCR/secretase complex regulates β- and γ-secretase specificity for Aβ production and contributes to AD pathogenesis

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