COPI‐coated vesicles mediate retrograde membrane traffic from the cis‐Golgi to the endoplasmic reticulum (ER) in all eukaryotic cells. However, it is still unknown whether COPI vesicles fuse everywhere or at specific sites with the ER membrane. Taking advantage of the circumstance that the vesicles still carry their coat when they arrive at the ER, we have visualized active ER arrival sites (ERAS) by monitoring contact between COPI coat components and the ER‐resident Dsl tethering complex using bimolecular fluorescence complementation (BiFC). ERAS form punctate structures near Golgi compartments, clearly distinct from ER exit sites. Furthermore, ERAS are highly polarized in an actin and myosin V‐dependent manner and are localized near hotspots of plasma membrane expansion. Genetic experiments suggest that the COPI•Dsl BiFC complexes recapitulate the physiological interaction between COPI and the Dsl complex and that COPI vesicles are mistargeted in dsl1 mutants. We conclude that the Dsl complex functions in confining COPI vesicle fusion sites.
The ER‐resident Dsl tethering complex interacts with COPI vesicles at spatially distinct ER arrival sites to prevent their mistargeting to adjacent ER exit sites, thus allowing to confine retrograde membrane transport vesicle fusion sites.
BiFC data reveal that coated COPI vesicles interact with the ER‐resident Dsl complex at discrete subdomains of the ER (ER arrival sites, ERAS).
ERAS and ER exit sites (ERES) share an interface at the ER and are juxtaposed to the Golgi, possibly marking an interaction hub of the early secretory pathway.
The Dsl complex regulates trafficking by preventing COPI vesicles from approaching adjacent ERES.
Growing cells feature a dominant polarized ERAS along with markers of secretion at the site of outgrowth, pointing towards a polarized focus of early trafficking activity.
Non‐growing cells show a disperse ERES‐like ERAS pattern.
The EMBO Journal (2016) 35: 1935–1955
- Received August 21, 2015.
- Revision received June 17, 2016.
- Accepted June 21, 2016.
- © 2016 The Authors