Uncoupled packaging of amyloid precursor protein and presenilin 1 into coat protein complex II vesicles.
Mutant forms of presenilin (PS) 1 and 2 and amyloid precursor protein (APP) lead to familial Alzheimer's disease. Several reports indicate that PS may modulate APP export from the endoplasmic reticulum (ER). To develop a test of this possibility, we reconstituted the capture of APP and PS1 in COPII (coat protein complex II) vesicles formed from ER membranes in permeabilized cultured cells. The recombinant forms of mammalian COPII proteins were active in a reaction that measures coat subunit assembly and coated vesicle budding on chemically defined synthetic liposomes. However, the recombinant COPII proteins were not active in cargo capture and vesicle budding from microsomal membranes. In contrast, rat liver cytosol was active in stimulating the sorting and packaging of APP, PS1, and p58 (an itinerant ER to Golgi marker protein) into transport vesicles from donor ER membranes. Budding was stimulated in dilute cytosol by the addition of recombinant COPII proteins. Fractionation of the cytosol suggested one or more additional proteins other than the COPII subunits may be essential for cargo selection or vesicle formation from the mammalian ER membrane. The recombinant Sec24C specifically recognized the APP C-terminal region for packaging. Titration of Sarla distinguished the packaging requirements of APP and PS1. Furthermore, APP packaging was not affected by deletion of PS1 or PS1 and 2, suggesting APP and PS1 trafficking from the ER are normally uncoupled.
Kim
Jinoh
J
Department of Molecular and Cell Biology, Howard Hughes Medical Institute, University of California, Berkeley, Berkeley, California 94720, USA.
Hamamoto
Susan
S
Ravazzola
Mariella
M
Orci
Lelio
L
Schekman
Randy
R
D000255:Adenosine Triphosphate / Q000737:chemistry; D016564:Amyloid beta-Protein Precursor / Q000737:chemistry* / Q000378:metabolism*; D000818:Animals; D016466:CHO Cells; D022181:COP-Coated Vesicles / Q000737:chemistry*; D006224:Cricetinae; D003600:Cytosol / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D004721:Endoplasmic Reticulum / Q000378:metabolism; D005347:Fibroblasts / Q000378:metabolism; D005982:Glutathione Transferase / Q000378:metabolism; D006801:Humans; D007425:Intracellular Membranes / Q000378:metabolism; D008081:Liposomes / Q000737:chemistry / Q000378:metabolism; D008099:Liver / Q000378:metabolism; D008565:Membrane Proteins / Q000737:chemistry* / Q000378:metabolism; D008862:Microsomes, Liver / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D053764:Presenilin-1; D017434:Protein Structure, Tertiary; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry; D013347:Subcellular Fractions / Q000378:metabolism
D016564:Amyloid beta-Protein Precursor; D008081:Liposomes; D008565:Membrane Proteins; C508853:PSEN1 protein, human; D053764:Presenilin-1; D011994:Recombinant Proteins; D000255:Adenosine Triphosphate; D005982:Glutathione Transferase