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| GTP hydrolysis by arf-1 mediates sorting and concentration of Golgi resident enzymes into functional COPI vesicles |
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| Author(s): Lanoix J, Ouwendijk J, Lin CC, Stark A, Love HD, Ostermann J, Nilsson T |
| Source: EMBO JOURNAL Volume: 18 Issue: 18 Pages: 4935-4948 Published: SEP 15 1999 |
| Times Cited: 132 References: 66 |
| Abstract: Upon addition of GTP gamma S to in vitro budding reactions, COP I vesicles form but retain their coat, making them easy to isolate and analyze, We have developed an in vitro budding assay that reconstitutes the formation of COP I-derived vesicles under conditions where GTP hydrolysis can occur. Once formed, vesicles are uncoated and appear functional as they fuse readily with acceptor membranes. Electron microscopy shows a homogeneous population of uncoated vesicles that contain the medial/trans Golgi enzyme alpha 1,2-mannosidase II. Biochemical quantitation of vesicles reveals that resident Golgi enzymes are up to 10-fold more concentrated than in donor membranes, but vesicles formed in the presence of GTP gamma S show an average density of resident Golgi enzymes similar to that seen in donor membranes. We show that the sorting process is mediated by the small GTPase arf-l as addition of a dominant, hydrolysis-deficient arf-l (Q)71(L) mutant produced results similar to that of GTP gamma S, Strikingly, the average density of the anterograde cargo protein, polymeric IgA receptor, in COP I-derived vesicles was similar to that found in starting membranes and was independent of GTP hydrolysis. We conclude that hydrolysis of GTP bound to arf-l promotes selective segregation and concentration of Golgi resident enzymes into COP I vesicles. |
| Document Type: Article |
| Language: English |
| Reprint Address: Nilsson, T (reprint author), EMBL, Cell Biol Program, Meyerhofstr 1, D-69112 Heidelberg, Germany |
Addresses:
1. EMBL, Cell Biol Program, D-69112 Heidelberg, Germany
2. Vanderbilt Univ, Sch Med, Dept Biochem, Nashville, TN 37232 USA |
| Publisher: OXFORD UNIV PRESS, GREAT CLARENDON ST, OXFORD OX2 6DP, ENGLAND |
| Subject Category: Biochemistry & Molecular Biology; Cell Biology |
| IDS Number: 258TZ |
| ISSN: 0261-4189 |
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| | | | | | | | | Record from Web of Science® | | | | | | | | | |