The cytosolic coat-protein complex COP-I interacts with cytoplasmic ‘retrieval’ signals present in membrane proteins that cycle between the endoplasmic reticulum (ER) and the Golgi complex, and is required for both anterograde and retrograde transport in the secretory pathway. Here we study the role of COP-I in Golgi-to-ER transport of several distinct marker molecules. Microinjection of anti-COP-I antibodies inhibits retrieval of the lectin-like molecule ERGIC-53 and of the KDEL receptor from the Golgi to the ER. Transport to the ER of protein toxins, which contain a sequence that is recognized by the KDEL receptor, is also inhibited. In contrast, microinjection of anti-COP-I antibodies or expression of a GTP-restricted Arf-1 mutant does not interfere with Golgi-to-ER transport of Shiga toxin/Shiga-like toxin-1 or with the apparent recycling to the ER of Golgi-resident glycosylation enzymes. Overexpression of a GDP-restricted mutant of Rab6 blocks transport to the ER of Shiga toxin/Shiga-like toxin-1 and glycosylation enzymes, but not of ERGIC-53, the KDEL receptor or KDEL-containing toxins. These data indicate the existence of at least two distinct pathways for Golgi-to-ER transport, one COP-I dependent and the other COP-I independent. The COP-I-independent pathway is specifically regulated by Rab6 and is used by Golgi glycosylation enzymes and Shiga toxin/Shiga-like toxin-1.
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We thank Leica Lasertechnik (Heidelberg, Germany), C. Zeiss (Jena, Germany), T.I.L.L. Photonics (Martinsried, Germany), Olympus Optical Co. (Hamburg, Germany), Eppendorf (Hamburg, Germany), Hamamatsu Photonics (Hamamatsu City, Japan) and Improvision (Coventry, UK) for their continued support of the Advanced Light Microscopy Facility (EMBL, Heidelberg, Germany). We also thank the following people for providing reagents: H.-P. Hauri for monoclonal antibodies to ERGIC-53 and p63; B. Luen Tang and W. Hong for monoclonal antibodies to the KDEL receptor; M. Lowe for polyclonal antibodies to GM130; the Zerial laboratory for anti-EEA1 human sera; S. Fuller for monoclonal antibody to PDI; and W. Balch for the cDNA encoding Sar1dn. This work was supported in part by grants to the individual laboratories and by a grant to B.S. from the Fogarty International Center, US NIH, while he was on sabbatical in T.N.’s laboratory. A.G. was financed in part by an ICRF postdoctoral fellowship.
Correspondence and requests for materials should be addressed to T.N. or R.P.
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