Retrograde transport from the pre-Golgi intermediate compartment and the Golgi complex is affected by the vacuolar H+-ATPase inhibitor bafilomycin A1

The effect of the vacuolar H+-ATPase inhibitor bafilomycin A1 (Baf A1) on t he localization of pre-Golgi intermediate compartment (IC) and Golgi marker proteins was used to study the role of acidification in the function of ea rly secretory compartments. Baf A1 inhibited both brefeldin A- and nocodazo le-induced retrograde transport of Golgi proteins to the endoplasmic reticu lum (ER), whereas anterograde ER-to-Golgi transport remained largely unaffe cted. Furthermore, p58/ERGIC-53, which normally cycles between the ER, IC, and cis-Golgi, was arrested in pre-Golgi tubules and vacuoles, and the numb er of p58-positive similar to 80-nm Golgi (coatomer protein I) vesicles was reduced, suggesting that the drug inhibits the retrieval of the protein fr om post-ER compartments. In parallel, redistribution of beta-coatomer prote in from the Golgi to peripheral pre-Golgi structures took place. The small GTPase rab1p was detected in short pre-Golgi tubules in control cells and w as efficiently recruited to the tubules accumulating in the presence of Baf A1. In contrast, these tubules showed no enrichment of newly synthesized, anterogradely transported proteins, indicating that they participate in ret rograde transport. These results suggest that the pre-Golgi structures cont ain an active H+-ATPase that regulates retrograde transport at the ER-Golgi boundary. Interestingly, although Baf A1 had distinct effects on periphera l pre-Golgi structures, only more central, p58-containing elements accumula ted detectable amounts of 3-(2,4-dinitroanilino)-3'-amino-N-methyl-dipropyl amine (DAMP), a marker for acidic compartments, raising the possibility tha t the lumenal pH of the pre-Golgi structures gradually changes in parallel with their translocation to the Golgi region.