COPII-coated vesicle formation reconstituted with purified coat proteins and chemically defined liposomes.
COPII vesicle formation requires only three coat assembly subunits: Sar1p, Sec13/31p, and Sec23/24p. PI 4-phosphate or PI 4,5-bisphosphate is required for the binding of these proteins to liposomes. The GTP-bound form of Sar1p recruits Sec23/24p to the liposomes as well as to the ER membranes, and this Sar1p-Sec23/24p complex is required for the binding of Sec13/31p. Ultrastructural analysis shows that the binding of COPII coat proteins to liposomes results in coated patches, coated buds, and coated vesicles of 50-90 nm in diameter. Budding proceeds without rupture of the donor liposome or vesicle product. These observations suggest that the assembly of the COPII coat on the ER occurs by a sequential binding of coat proteins to specific lipids and that this assembly promotes the budding of COPII-coated vesicles.
Matsuoka
K
K
Department of Molecular and Cell Biology, Howard Hughes Medical Institute, University of California, Berkeley 94720, USA.
Orci
L
L
Amherdt
M
M
Bednarek
S Y
SY
Hamamoto
S
S
Schekman
R
R
Yeung
T
T
D022181:COP-Coated Vesicles; D018699:Coated Vesicles / Q000378:metabolism* / Q000648:ultrastructure; D004721:Endoplasmic Reticulum / Q000378:metabolism; D005656:Fungal Proteins / Q000302:isolation & purification / Q000378:metabolism*; D019204:GTP-Binding Proteins / Q000302:isolation & purification / Q000378:metabolism*; D020690:GTPase-Activating Proteins; D006157:Guanosine Monophosphate / Q000031:analogs & derivatives; D008051:Lipid Bilayers; D008081:Liposomes / Q000737:chemistry / Q000378:metabolism*; D008565:Membrane Proteins / Q000302:isolation & purification / Q000378:metabolism*; D008861:Microsomes / Q000378:metabolism; D020559:Monomeric GTP-Binding Proteins; D028861:Nuclear Pore Complex Proteins; D010712:Phosphatidic Acids; D010743:Phospholipids; D011510:Proteolipids / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D033921:Vesicular Transport Proteins
D005656:Fungal Proteins; D020690:GTPase-Activating Proteins; D008051:Lipid Bilayers; D008081:Liposomes; D008565:Membrane Proteins; D028861:Nuclear Pore Complex Proteins; D010712:Phosphatidic Acids; D010743:Phospholipids; D011510:Proteolipids; C079620:SEC13 protein, S cerevisiae; C079037:SEC23 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D033921:Vesicular Transport Proteins; C020485:proteoliposomes; D006157:Guanosine Monophosphate; D019204:GTP-Binding Proteins; D020559:Monomeric GTP-Binding Proteins; C077395:SAR1 protein, S cerevisiae