Roles for α2p24 and COPI in endoplasmic reticulum cargo exit site formation

被引:78
作者
Lavoie, C
Paiement, J
Dominguez, M
Roy, L
Dahan, S
Gushue, JN
Bergeron, JJM
机构
[1] Univ Montreal, Fac Med, Dept Pathol & Biol Cellulaire, Montreal, PQ H3C 3J7, Canada
[2] McGill Univ, Dept Anat & Cell Biol, Montreal, PQ H3A 2B2, Canada
关键词
cell-free assembly; transitional endoplasmic reticulum; endoplasmic reticulum cargo exit sites; alpha(2)p24; COPI;
D O I
10.1083/jcb.146.2.285
中图分类号
Q2 [细胞生物学];
学科分类号
071009 ; 090102 ;
摘要
A two-step reconstitution system for the generation of ER cargo exit sites from starting ER-derived low density microsomes (LDMs; 1.17 g/cc) is described. The first step is mediated by the hydrolysis of Mg(2+)ATP and Mg(2+)GTP, leading to the formation of a transitional ER (tER) with the soluble cargo albumin, transferrin, and the ER-to-Golgi recycling membrane proteins alpha 2p24 and p58 (ERGIC-53, ER-Golgi intermediate compartment protein) enriched therein. Upon further incubation (step two) with cytosol and mixed nucleotides, interconnecting smooth ER tubules within tER transforms into vesicular tubular clusters (VTCs). The cytosolic domain of alpha(2)p24 and cytosolic COPI coatomer affect VTC formation. This is deduced from the effect of antibodies to the COOH-terminal tail of alpha(2)p24, but not of antibodies to the COOH-terminal tail of calnexin on this reconstitution, as well as the demonstrated recruitment of COPI coatomer to VTCs, its augmentation by GTP gamma S, inhibition by Brefeldin A (BFA), or depletion of beta-COP from cytosol. Therefore, the p24 family member, alpha(2)p24, and its cytosolic coat ligand, COPI coatomer, play a role in the de novo formation of VTCs and the generation of ER cargo exit sites.
引用
收藏
页码:285 / 299
页数:15
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