Luciano Dadamio - Academia.edu (original) (raw)

Neuron, 2005

gates of amyloid β-peptides (Aβ) (Haass and Selkoe, and Frédéric Checler 1, * 1993). Aβ is a 40-4... more gates of amyloid β-peptides (Aβ) (Haass and Selkoe, and Frédéric Checler 1, * 1993). Aβ is a 40-42 amino acid peptide that is gener-1 Institut de Pharmacologie Moléculaire et Cellulaire ated from the β-Amyloid Precursor Protein (βAPP) by Centre National de la Recherche Scientifique two sequential cleavages. The first of these cleavages UMR6097 CNRS/UNSA occurs in the extracellular domain and is mediated by Valbonne 06560 a membrane-bound aspartyl protease termed β-secre-France tase (Vassar and Citron, 2000). The second set of cleav-2 Centre for Research in Neurodegenerative Diseases ages occurs at residues 40-42 (termed γ-site) and at Department of Medicine residues 48-52 (termed ⑀-site) within the transmem-University of Toronto and University Health Network brane domain of the βAPP stub generated by β-secre-Toronto Western Hospital Research Institute tase. The γ-site cleavage generates Aβ, while the con-6 Queen's Park Crescent current ⑀-site cleavage generates a cytosolic stub Toronto, Ontario M5S 3H2 referred to as ICD (Passer et al., 2000) or AICD (βAPP Canada IntraCellular Domain). The exact role of AICD remains 3 Institute for Pharmacy and Molecular Biotechnology unclear. University of Heidelberg Both the γand the ⑀-site cleavages are mediated by 69120 Heidelberg presenilin (PS)-independent and dependent proteases Germany (De Strooper et al., 1998; Armogida et al., 2001). The 4 Albert Einstein College of Medicine presenilin-dependent γ-secretase and ⑀-site proteolytic New York, New York activities (which are often generically collectively termed 5 Dipartimento di Biochimica e Biotecnologie Mediche γ-secretase) are dependent upon a multimeric complex Universita' Degli Studi di Napoli Federico II of at least four different membrane proteins including Napoli Presenilin 1(PS1) or Presenilin 2 (PS2), nicastrin, Aph-1, Italy and Pen-2 (Yu et al., 2000; Francis et al., 2002). In these 6 Center for Neurologic Diseases complexes, the presenilins have been proposed as a Harvard Medical School novel type of transmembrane aspartyl protease bearing Boston, Massachusetts the catalytic core of the γ-secretase (Wolfe et al., 1999). This novel type of intramembranous proteolysis apparently governs the function of βAPP and several Type I transmembrane proteins including Notch, cadherins, Summary ErbB-4, CD44, or p75 NTR . Many of these proteins are involved in a variety of vital cellular functions such as Amyloid ␤-peptide (A␤), which plays a central role in intracellular signaling in development and adulthood, Alzheimer's disease, is generated by presenilincell adhesion, cell growth and proliferation, and kinase dependent ␥-secretase cleavage of ␤-amyloid precuractivities (for review see Sisodia and St. George-Hyssor protein (␤APP). We report that the presenilins lop, 2002; Pollack and Lewis, 2005). Thus, γ-secretase (PS1 and PS2) also regulate A␤ degradation. Presenicleavage of Notch releases an intracellular fragment lin-deficient cells fail to degrade A␤ and have drastic called NICD (Notch IntraCellular Domain), which acts reductions in the transcription, expression, and activas a transcription factor mediating signal transduction ity of neprilysin, a key A␤-degrading enzyme. Nepriin the Notch-Delta pathway, a critical intercellular siglysin activity and expression are also lowered by naling mechanism, during both embryonic develop-␥-secretase inhibitors and by PS1/PS2 deficiency in ment and adulthood (Kopan et al., 1996; Shen et al., mouse brain. Neprilysin activity is restored by tran-1997; De Strooper et al., 1998; Kopan and Goate, 2000). sient expression of PS1 or PS2 and by expression Under normal conditions, Aβ occurs as a soluble of the amyloid intracellular domain (AICD), which is fragment, the concentration of which is normally tightly cogenerated with A␤, during ␥-secretase cleavage of controlled below the threshold for its self-aggregation ␤APP. Neprilysin gene promoters are transactivated into β sheet fibrils (Burdick et al., 1992). Aβ is actively by AICDs from APP-like proteins (APP, APLP1, and degraded by several enzymes including neprilysin APLP2), but not by A␤ or by the ␥-secretase cleavage (NEP), insulin-degrading enzyme (IDE), and endothelinproducts of Notch, N-or E-cadherins. The presenilinconverting enzyme (ECE) (Carson and Turner, 2002). Aldependent regulation of neprilysin, mediated by though these Aβ-degrading enzymes have been well AICDs, provides a physiological means to modulate characterized, very little is known about the regulatory A␤ levels with varying levels of ␥-secretase activity. mechanisms that govern their expression and/or activity. Nevertheless, under normal physiological circumstances, the balance between the rates of production *Correspondence: checler@ipmc.cnrs.fr Neuron 542 Neuron 552 CGACC3# raising a 295 bp probe, 5#GGAACCTCCCCCAAGTCC3#/ medicals, Orsay, France). For the preparation of nuclear extracts, R.S. (1998). The X11α protein slows cellular amyloid precursor pro-HEK293 cells were cultivated in 100 mm diameter dishes and trantein processing and reduces Aβ40 and Aβ42 secretion. J. Biol. siently transfected with 12 g cDNA of either empty pcDNA3 vector Chem. 273, 14761-14766. or a mix of myc-tagged-AICDC59 (6 g), TIP60 (3 g), and Fe65 (3 Burdick, D., Soreghan, B., Kwon, M., Kosmoski, J., Knauer, M., g) by means of DAC30 reactive as previously described. Forty-Henschen, A., Yates, J., Cotman, C., and Glabe, C. (1992). Assemeight hours after transfection, cells were harvested and nuclear exbly and aggregation properties of synthetic Alzheimer's A4/beta tracts were prepared according to the Current Protocols in Molecuamyloid peptide analogs. J. Biol. Chem. 267, 546-554. lar Biology (Ausubel et al., 2002). Binding reactions containing nuclear Cao, X., and Südhof, T.C. (2001). A transcriptively active complex extracts (10 g) were performed at 37°C, using the shorter 219 bp of APP with Fe65 and histone acetyltransferase tip60. Science 293, probe according to the manufacturer's directions. Then protein-115-120. DNA complexes were resolved by electrophoresis through 7% native polyacrylamide gels in buffer containing 5mM Tris (pH, 8.3) and Carson, J.A., and Turner, A.J. (2002). β-amyloid catabolism: roles 38 mM Glycine for 3 hr at 300V. Gels were dried and autoradiofor neprilysin (NEP) and other metallopeptidases. J. Neurochem. graphed on a BAS-1500 phosphorimager (Fujifilm, Tokyo, Japan). 81, 1-8. The specificity of the above described reactions were verified by Checler, F. (1993). Neuropeptide-degrading peptidases. In Methods Supershift gel assay corresponding to the preincubation of nuclear in Neurotransmitters and Neuropeptides Research. Part 2., T. Naextracts with either anti-myc (specific) or anti-rabbit (nonspecific) gatsu, H. Parvez, M. Naoi, and S. Parvez, eds. (Amsterdam: Elsevier antibodies before allowing the binding reactions and subsequent Science Publishers), pp. 375-418. incorporation of the labeled probe. Checler, F. (1999). Presenilins: Structural aspects and post-translational events. Mol. Neurobiol. 19, 255-265. Normal and Pathological Human Brain Tissues Chen, F., Gu, Y., Hasegawa, H., Ruan, X., Arawaka, S., Fraser, P., All brain samples correspond to frontal cortices. Samples from Westaway, D., Mount, H., and St. George-Hyslop, P. (2002). Presen-Rouen include a control brain (female, 74 years) and one Familial ilin 1 mutations activate γ-42-secretase but reciprocally inhibit AD brain (female, 51 years, Leu392Val-PS1 mutation). Samples ⑀-secretase cleavage of APP and S3-cleavage of Notch. J. Biol. from la Pitié Salpêtrière (Paris) correspond to two control brains Chem. 277, 36521-36526. (males, 72 and 55 years), two sporadic AD brains (females, 75 and De Strooper, B. (2003). Aph-1, Pen-2, and nicastrin with presenilin 82 years), and two Familial AD brains (female, 37 years, Leu235Progenerate an active γ-secretase complex. Neuron 38, 9-12. PS1 mutation; male, 44 years, Phe386Ser-PS1 mutation). De Strooper, B., Saftig, P., Craessaerts, K., Vanderstichele, H., Statistical Analysis Guhde, G., Von Figura, K., and Van Leuven, F. (1998). Deficiency of Statistical analyses were performed with PRISM Software (Graphpresenilin 1 inhibits the normal cleavage of amyloid precursor pro-Pad Software, San Diego) by using the unpaired Student's t test for tein. Nature 391, 387-390. pairwise comparisons.