INHIBITION OF ENDOSOME FUSION BY PHOSPHOLIPASE-A(2) (PLA2) INHIBITORS POINTS TO A ROLE FOR PLA2 IN ENDOCYTOSIS

Fusion of intracellular membrane-bound compartments is a common step in the transport of macromolecules along the endocytic and secretory pathways. A large number of factors active in the fusion process or its regulation have been identified; however, the actual sequence of events leading to membrane fusion is still unknown. In this study, we have assessed a possible role for PLA2 in endosome fusion by using an in vitro reconstitution assay and by examining endocytosis in intact cells. Several PLA2 inhibitors blocked endosome fusion in a broken-cell preparation. Inhibition was reversed by addition of arachidonic acid. At the electron microscope level, endosome clusters were observed even in the presence of inhibitors; however, actual fusion between endosomes was largely reduced. Fusion frequency increased upon the addition of arachidonic acid. A membrane-permeable PLA2 inhibitor blocked mixing of ligands internalized sequentially but did not affect internalization. The results indicate that vesicle fusion along the endocytic pathway requires a PLA2 activity. The effect of this activity would be, at least in part, mediated by arachidonic acid release.