A phorbol ester-binding protein is required downstream of Rab5 is endosomefusion

Previous observations indicate that a zinc and phorbol ester binding factor is necessary for endosome fusion, To further characterize the role of this factor in the process, we used an in vitro endosome fusion assay supplemen ted with recombinant Rab5 proteins. Both zinc depletion and addition of cal phostin C, an inhibitor of protein kinase C, inhibited endosome fusion in t he presence of active Rab5. Addition of the phorbol ester PMA (phorbol 12-m yristate 13-acetate) reversed the inhibition of endosome fusion caused by a Rab5 negative mutant. Moreover, PMA stimulated fusion in the presence of R ab5 immunodepleted cytosol. These results suggest that the phorbol ester bi nding protein is acting downstream of Rab5 in endosome fusion, (C) 1998 Fed eration of European Biochemical Societies.