A Di-acidic (DXE) Code Directs Concentration of Cargo during Export from the Endoplasmic Reticulum*
- Noriyuki Nishimuraद,
- Sergei Bannykh‡§,
- Sarah Slabough‡,
- Jeanne Matteson‡,
- Yoram Altschuler‖,
- Klaus Hahn‡ and
- William E. Balch‡§**
- From the Departments of ‡Cell and§Molecular Biology, The Scripps Research Institute, La Jolla, California 92037 and the ‖Department of Anatomy, University of California, San Francisco, California 941143
Abstract
Efficient export of vesicular stomatitis virus glycoprotein (VSV-G), a type I transmembrane protein, from the endoplasmic reticulum requires a di-acidic code (DXE) located in the cytosolic carboxyl-terminal tail (Nishimura, N., and Balch, W. E. (1997) Science 277, 556–558). Mutation of the DXE code by mutation to AXA did not prevent VSV-G recruitment to pre-budding complexes formed in the presence of the activated form of the Sar1 and the Sec23/24 complex, components of the COPII budding machinery. However, the signal was required at a subsequent concentration step preceding vesicle fission. By using green fluorescence protein-tagged VSV-G to image movement in a single cell, we found that VSV-G lacking the DXE code fails to be concentrated into COPII vesicles. As a result, the normal 5–10-fold increase in the steady-state concentration of VSV-G in downstream pre-Golgi intermediates and Golgi compartments was lost. These results demonstrate for the first time that inactivation of the DXE signal uncouples early cargo selection steps from concentration into COPII vesicles. We propose that two sequential steps are required for efficient export from the endoplasmic reticulum.
Footnotes
-
↵* This work was supported by National Institutes of Health Grant GM 42336 (to W. E. B.) and by Core C of the National Cancer Institute Grant CA 58689.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
-
↵¶ Senior Postdoctoral Fellow of the American Cancer Society.
-
↵** To whom correspondence should be addressed: Depts. of Cell and Molecular Biology, The Scripps Research Institute, 10550 N. Torrey Pines Rd., La Jolla, CA 92037. Tel: 619-784-2310; Fax: 619-784-9126; E-mail: webalch{at}scripps.edu.
-
↵2 The on-line address is as follows:www.scripps.edu/cb/balch/dxepaper. (User, balch) (password, ER).
- Abbreviations:
- ER
-
endoplasmic reticulum
- VSV-G
-
vesicular stomatitis virus glycoprotein
- endo H
-
endoglycosidase H
- BHK
-
baby hamster kidney
- NRK
-
normal rat kidney
- GFP
-
green fluorescent protein
- GST
-
glutathioneS-transferase
- HRP
-
horseradish peroxidase
- HA
-
hemagglutinin
- PBS
-
phosphate-buffered saline
- GS beads
-
glutathione-Sepharose beads
- BSA
-
bovine serum albumin
- Syn5
-
syntaxin 5
- SFV
-
Semliki forest virus
- TGN
-
trans Golgi network
-
- Received February 2, 1999.
- Revision received March 24, 1999.
- The American Society for Biochemistry and Molecular Biology, Inc.
