FUSION OF LYSOSOMES WITH LATE ENDOSOMES PRODUCES A HYBRID ORGANELLE OF INTERMEDIATE DENSITY AND IS NSF DEPENDENT

Using a cell-free content mixing assay containing rat liver endosomes and lysosomes in the presence of pig brain cytosol, we demonstrated th at after incubation at 37 degrees C, late endosome-lysosome hybrid org anelles were formed, which could be isolated by density gradient centr ifugation. ImmunoEM showed that the hybrids contained both an endocyto sed marker and a lysosomal enzyme. Formation of the hybrid organelles appeared not to require vesicular transport between late endosomes and lysosomes but occurred as a result of direct fusion. Hybrid organelle s with similar properties were isolated directly from rat liver homoge nates and thus were not an artifact of cell-free incubations. Direct f usion between late endosomes and lysosomes was an N-ethylmaleimide-sen sitive factor-dependent event and was inhibited by GDP-dissociation in hibitor, indicating a requirement for a rab protein, We suggest that i n cells, delivery of endocytosed ligands to an organelle where proteol ytic digestion occurs is mediated by direct fusion of late endosomes w ith lysosomes. The consequences of this fusion to the maintenance and function of lysosomes are discussed.