Differential effects of a GTP-restricted mutant of Sar1p on segregation of cargo during export from the endoplasmic reticulum.

Export of cargo from the endoplasmic reticulum (ER) is the first membrane trafficking step in the secretory pathway. To date, all cargo proteins appear to use a common set of machinery for the initial stages of export, namely the COPII coat complex. Recent data from both yeast and mammalian systems have emerged suggesting that specific cargoes could be sorted from one another at the point of exit from the endoplasmic reticulum or immediately afterwards. Here, we have examined the mechanisms used for export of different types of cargo molecule from the endoplasmic reticulum. All cargoes examined utilise the COPII machinery, but specific differences are seen in the accumulation of cargo into ER-derived pre-budding complexes following expression of a GTP-restricted mutant of the Sar1p GTPase. Glycosylphosphatidylinositol (GPI)-anchored GFP is seen to be restricted to the ER under these conditions whereas other cargoes, including ts045-G and lumFP accumulate in pre-budding complexes. Following exit, GPI-FP, lumFP and ts045-G-FP all travel to the Golgi in the same vesicular tubular clusters (VTCs). These data show a differential requirement for efficient GTP hydrolysis by the Sar1p GTPase in export of cargo from the ER.