Identification of phospholipid scramblase 1 as a novel interacting molecule with beta -secretase (beta -site amyloid precursor protein (APP) cleaving enzyme (BACE)).
beta-Site amyloid precursor protein (APP)-cleaving enzyme (BACE) is an integral membrane aspartic proteinase responsible for beta-site processing of APP, and its cytoplasmic region composed of 24 amino acid residues has been shown to be involved in the endosomal localization of BACE. With the yeast two-hybrid screening, we found that the cytoplasmic domain of phospholipid scramblase 1 (PLSCR1), a type II integral membrane protein, interacts with the cytoplasmic region of BACE. In cultured cells, BACE and PLSCR1 were colocalized in the Golgi area and in endosomal compartments, whereas they were co-redistributed in late endosome-derived multivesicular bodies when treated with U18666A, suggesting that both proteins share a common trafficking pathway in cells. Co-immunoprecipitation analysis showed that both proteins form a protein complex at an endogenous expression level in the human neuroblastoma SH-SY5Ycells, and the dileucine residue of the BACE tail is also revealed to be essential for the physical interaction with PLSCR1 in vitro and in vivo. Moreover, both BACE and PLSCR1 were localized in a low buoyant lipid microdomain in SH-SY5Y cells. The dileucine-defective BACE mutant was also fractionated into the lipid microdomain, but much less stably than wild-type BACE. Taken together, our current study suggests the functional involvement of PLSCR1 in the intracellular distribution of BACE and/or recruitment of BACE into the detergent-insoluble lipid raft.
Kametaka
Satoshi
S
Department of Cell Biology and Neuroscience A1, Osaka University Graduate School of Medicine, Yamadaoka 2-2, Suita, Osaka 565-0871, Japan.
Shibata
Masahiro
M
Moroe
Kimiho
K
Kanamori
Shiro
S
Ohsawa
Yoshiyuki
Y
Waguri
Satoshi
S
Sims
Peter J
PJ
Emoto
Kazuo
K
Umeda
Masato
M
Uchiyama
Yasuo
Y
D053829:Amyloid Precursor Protein Secretases; D016282:Aspartic Acid Endopeptidases / Q000737:chemistry / Q000378:metabolism*; D001665:Binding Sites; D002352:Carrier Proteins / Q000737:chemistry / Q000378:metabolism*; D010450:Endopeptidases; D011992:Endosomes / Q000737:chemistry; D006056:Golgi Apparatus / Q000737:chemistry; D006801:Humans; D021962:Membrane Microdomains; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism*; D049294:Phospholipid Transfer Proteins; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D014407:Tumor Cells, Cultured; D020798:Two-Hybrid System Techniques
D002352:Carrier Proteins; D008565:Membrane Proteins; D049294:Phospholipid Transfer Proteins; D053829:Amyloid Precursor Protein Secretases; D010450:Endopeptidases; D016282:Aspartic Acid Endopeptidases; C509280:BACE1 protein, human