Identification of novel γ-secretase-associated proteins in detergent-resistant membranes from brain.
In Alzheimer disease, oligomeric amyloid β-peptide (Aβ) species lead to synapse loss and neuronal death. γ-Secretase, the transmembrane protease complex that mediates the final catalytic step that liberates Aβ from its precursor protein (APP), has a multitude of substrates, and therapeutics aimed at reducing Aβ production should ideally be specific for APP cleavage. It has been shown that APP can be processed in lipid rafts, and γ-secretase-associated proteins can affect Aβ production. Here, we use a biotinylated inhibitor for affinity purification of γ-secretase and associated proteins and mass spectrometry for identification of the purified proteins, and we identify novel γ-secretase-associated proteins in detergent-resistant membranes from brain. Furthermore, we show by small interfering RNA-mediated knockdown of gene expression that a subset of the γ-secretase-associated proteins, in particular voltage-dependent anion channel 1 (VDAC1) and contactin-associated protein 1 (CNTNAP1), reduced Aβ production (Aβ40 and Aβ42) by around 70%, whereas knockdown of presenilin 1, one of the essential γ-secretase complex components, reduced Aβ production by 50%. Importantly, these proteins had a less pronounced effect on Notch processing. We conclude that VDAC1 and CNTNAP1 associate with γ-secretase in detergent-resistant membranes and affect APP processing and suggest that molecules that interfere with this interaction could be of therapeutic use for Alzheimer disease.
Karolinska Institutet Dainippon Sumitomo Pharma Alzheimer Center, KI Alzheimer Disease Research Center, Department of Neurobiology, Karolinska Institutet, Novum, Huddinge SE-141 57, Sweden. hurj@mskcc.org
Ji-Yeun
JY
Hur
Yasuhiro
Y
Teranishi
Takahiro
T
Kihara
Natsuko Goto
NG
Yamamoto
Mitsuhiro
M
Inoue
Waltteri
W
Hosia
Masakazu
M
Hashimoto
Bengt
B
Winblad
Susanne
S
Frykman
Lars O
LO
Tjernberg
D000595:Amino Acid Sequence; D053829:Amyloid Precursor Protein Secretases / Q000302:isolation & purification / Q000378:metabolism*; D016229:Amyloid beta-Peptides / Q000096:biosynthesis; D000818:Animals; D001921:Brain / Q000201:enzymology / Q000378:metabolism*; D015816:Cell Adhesion Molecules, Neuronal / Q000235:genetics / Q000378:metabolism*; D002846:Chromatography, Affinity; D055785:Gene Knockdown Techniques; D057809:HEK293 Cells; D006801:Humans; D008297:Male; D008562:Membrane Glycoproteins / Q000378:metabolism; D021962:Membrane Microdomains / Q000378:metabolism* / Q000648:ultrastructure; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry; D053764:Presenilin-1 / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D051381:Rats; D017207:Rats, Sprague-Dawley; D051880:Receptors, Notch / Q000378:metabolism; D050827:Syntaxin 1 / Q000737:chemistry / Q000378:metabolism; D053719:Tandem Mass Spectrometry; D050995:Voltage-Dependent Anion Channel 1 / Q000235:genetics / Q000378:metabolism*
D016229:Amyloid beta-Peptides; C491323:CNTNAP1 protein, human; D015816:Cell Adhesion Molecules, Neuronal; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; C508853:PSEN1 protein, human; D010446:Peptide Fragments; D053764:Presenilin-1; D051880:Receptors, Notch; D050827:Syntaxin 1; C100672:Tmed10 protein, rat; C494762:VDAC1 protein, human; C413604:nicastrin protein; D050995:Voltage-Dependent Anion Channel 1; D053829:Amyloid Precursor Protein Secretases