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Proc Natl Acad Sci U S A. 2018 Apr 3;115(14):E3155-E3162. doi: 10.1073/pnas.1704639115. Epub 2018 Mar 19.

Dimeric sorting code for concentrative cargo selection by the COPII coat.

Nie C1,2, Wang H3, Wang R3, Ginsburg D4,5,6,7,8, Chen XW9,2,3.

Author information

1
State Key Laboratory of Membrane Biology, Peking University, Beijing 100871, China.
2
Institute of Molecular Medicine, Peking University, Beijing 100871, China.
3
Center for Life Sciences, Peking University, Beijing 100871, China.
4
Life Sciences Institute, University of Michigan, Ann Arbor, MI 48109; ginsburg@umich.edu xiaowei_chen@pku.edu.cn.
5
Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, MI 48109.
6
Department of Human Genetics, University of Michigan Medical School, Ann Arbor, MI 48109.
7
Department of Pediatrics, University of Michigan Medical School, Ann Arbor, MI 48109.
8
Howard Hughes Medical Institute, University of Michigan, Ann Arbor, MI 48109.
9
State Key Laboratory of Membrane Biology, Peking University, Beijing 100871, China; ginsburg@umich.edu xiaowei_chen@pku.edu.cn.

Abstract

The flow of cargo vesicles along the secretory pathway requires concerted action among various regulators. The COPII complex, assembled by the activated SAR1 GTPases on the surface of the endoplasmic reticulum, orchestrates protein interactions to package cargos and generate transport vesicles en route to the Golgi. The dynamic nature of COPII, however, hinders analysis with conventional biochemical assays. Here we apply proximity-dependent biotinylation labeling to capture the dynamics of COPII transport in cells. When SAR1B was fused with a promiscuous biotin ligase, BirA*, the fusion protein SAR1B-BirA* biotinylates and thus enables the capture of COPII machinery and cargos in a GTP-dependent manner. Biochemical and pulse-chase imaging experiments demonstrate that the COPII coat undergoes a dynamic cycle of engagement-disengagement with the transmembrane cargo receptor LMAN1/ERGIC53. LMAN1 undergoes a process of concentrative sorting by the COPII coat, via a dimeric sorting code generated by oligomerization of the cargo receptor. Similar oligomerization events have been observed with other COPII sorting signals, suggesting that dimeric/multimeric sorting codes may serve as a general mechanism to generate selectivity of cargo sorting.

KEYWORDS:

COPII; LMAN1; cargo receptor; cargo sorting; proximity-dependent biotinylation

PMID:
29555761
PMCID:
PMC5889621
DOI:
10.1073/pnas.1704639115
[Indexed for MEDLINE]
Free PMC Article

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