CHARACTERISTICS OF ENDOPLASMIC RETICULUM-DERIVED TRANSPORT VESICLES

被引:100
作者
REXACH, MF
LATTERICH, M
SCHEKMAN, RW
机构
[1] UNIV CALIF BERKELEY, DEPT MOLEC & CELL BIOL, BERKELEY, CA 94720 USA
[2] UNIV CALIF BERKELEY, HOWARD HUGHES MED INST, BERKELEY, CA 94720 USA
关键词
D O I
10.1083/jcb.126.5.1133
中图分类号
Q2 [细胞生物学];
学科分类号
071009 ; 090102 ;
摘要
We have isolated vesicles that mediate protein transport from the ER to Golgi membranes in perforated yeast. These vesicles, which form de novo during in vitro incubations, carry lumenal and membrane proteins that include core-glycosylated pro-alpha-factor, Bet1, Sec22, and Bos1, but not ER-resident Kar2 or Sec61 proteins. Thus, lumenal and membrane proteins in the ER are sorted prior to transport vesicle scission. Inhibition of Yptlp-function, which prevents newly formed vesicles from docking to cis-Golgi membranes, was used to block transport. Vesicles that accumulate are competent for fusion with cis-Golgi membranes, but not with ER membranes, and thus are functionally committed to vectorial transport. A 900-fold enrichment was developed using differential centrifugation and a series of velocity and equilibrium density gradients. Electron microscopic analysis shows a uniform population of 60 nm vesicles that lack peripheral protein coats. Quantitative Western blot analysis indicates that protein markers of cytosol and cellular membranes are depleted throughout the purification, whereas the synaptobrevin-like Bet1, Sec22, and Bos1 proteins are highly enriched. Uncoated ER-derived transport vesicles (ERV) contain twelve major proteins that associate tightly with the membrane. The ERV proteins may represent abundant cargo and additional targeting molecules.
引用
收藏
页码:1133 / 1148
页数:16
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