Study on the putative contribution of caspases and the proteasome to the degradation of Aph-1a and Pen-2.
The presenilin-dependent gamma-secretase complex is mainly composed of four distinct proteins, namely presenilin 1 or presenilin 2, nicastrin, anterior pharynx defective-1 (Aph-1) and presenilin enhancer (Pen-2). The mechanisms by which the complex is assembled, how its stoichiometry is controlled and how its catalytic activity is regulated are poorly understood. Recent studies indicated that Aph-1 and Pen-2 undergo proteolysis by the proteasome. We have examined the susceptibility of endogenous and overexpressed Aph-1a and Pen-2 to proteolysis by endogenous and purified proteasome as well as by recombinant caspases. We show that endogenous Aph-1a and Pen-2 resist proteolysis by caspases and by the proteasome. Furthermore, we show that unexpected interference of proteasome inhibitors with the cmv promoter region driving expression of Aph-1a and Pen-2 led to artifactual enhancement of overexpressed Aph-1a and Pen-2-like immunoreactivities but that these proteins also resist to in vitro degradation by endogenous and purified proteasome.
Dunys
Julie
J
Institut de Pharmacologie Moléculaire et Cellulaire, UMR6097 CNRS/UNSA, Equipe labellisée Fondation pour la Recherche Médicale, Valbonne, France.
Kawarai
Toshitaka
T
Giaime
Emilie
E
Wilk
Sherwin
S
Herrant
M
M
Auberger
P
P
St George-Hyslop
Peter
P
Alves da Costa
Cristine
C
Checler
Frédéric
F
D053829:Amyloid Precursor Protein Secretases / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D020169:Caspases / Q000378:metabolism*; D002417:Cattle; D002460:Cell Line; D004305:Dose-Response Relationship, Drug; D010450:Endopeptidases; D015870:Gene Expression / Q000187:drug effects; D007976:Leupeptins / Q000494:pharmacology; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D009154:Mutation / Q000502:physiology; D009842:Oligopeptides / Q000494:pharmacology; D010447:Peptide Hydrolases / Q000235:genetics / Q000378:metabolism*; D011480:Protease Inhibitors / Q000494:pharmacology; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism*; D014162:Transfection / Q000379:methods
D007976:Leupeptins; D008565:Membrane Proteins; D009842:Oligopeptides; C469349:PSENEN protein, human; D011480:Protease Inhibitors; C090535:benzyloxycarbonyl-isoleucyl-glutamyl(O-tert-butyl)-alanyl-leucinal; C475154:APH1A protein, human; D053829:Amyloid Precursor Protein Secretases; D010450:Endopeptidases; D010447:Peptide Hydrolases; D020169:Caspases; D046988:Proteasome Endopeptidase Complex; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde