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© The Rockefeller University Press, 0021-9525/1997//789 $5.00
The Journal of Cell Biology, Volume 136, Number 4, , 1997 789-802

Article

COPI-independent Anterograde Transport: Cargo-selective ER to Golgi Protein Transport in Yeast COPI Mutants



Erin C. Gaynor and Scott D. Emr

Department of Biology, The Division of Cellular and Molecular Medicine, and The Howard Hughes Medical Institute, University of California, San Diego, La Jolla, California 92093-0668

The coatomer (COPI) complex mediates Golgi to ER recycling of membrane proteins containing a dilysine retrieval motif. However, COPI was initially characterized as an anterograde-acting coat complex. To investigate the direct and primary role(s) of COPI in ER/Golgi transport and in the secretory pathway in general, we used PCR-based mutagenesis to generate new temperature-conditional mutant alleles of one COPI gene in Saccharomyces cerevisiae, SEC21 ({gamma}-COP). Unexpectedly, all of the new sec21 ts mutants exhibited striking, cargo-selective ER to Golgi transport defects. In these mutants, several proteins (i.e., CPY and {alpha}-factor) were completely blocked in the ER at nonpermissive temperature; however, other proteins (i.e., invertase and HSP150) in these and other COPI mutants were secreted normally. Nearly identical cargo-specific ER to Golgi transport defects were also induced by Brefeldin A. In contrast, all proteins tested required COPII (ER to Golgi coat complex), Sec18p (NSF), and Sec22p (v-SNARE) for ER to Golgi transport. Together, these data suggest that COPI plays a critical but indirect role in anterograde transport, perhaps by directing retrieval of transport factors required for packaging of certain cargo into ER to Golgi COPII vesicles. Interestingly, CPY–invertase hybrid proteins, like invertase but unlike CPY, escaped the sec21 ts mutant ER block, suggesting that packaging into COPII vesicles may be mediated by cis-acting sorting determinants in the cargo proteins themselves. These hybrid proteins were efficiently targeted to the vacuole, indicating that COPI is also not directly required for regulated Golgi to vacuole transport. Additionally, the sec21 mutants exhibited early Golgi-specific glycosylation defects and structural aberrations in early but not late Golgi compartments at nonpermissive temperature. Together, these studies demonstrate that although COPI plays an important and most likely direct role both in Golgi–ER retrieval and in maintenance/function of the cis-Golgi, COPI does not appear to be directly required for anterograde transport through the secretory pathway.

Abbreviations used in this paper: ALP, alkaline phosphatase; ARF, ADP-ribosylation factor; BFA, Brefeldin A; CPS, carboxypeptidase S; CYP, carboxypeptidase Y; CV, coated vesicle; HA, hemagglutinin; ts, temperature sensitive.

Please address all correspondence to Scott Emr, Department of Biology, the Division of Cellular and Molecular Medicine, and the Howard Hughes Medical Institute, University of California, San Diego, La Jolla, CA 92093-0668. Tel.: (619) 534-6462; Fax.: (619) 534-6414.


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