Vol. 151, No. 1, 1988 BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS February 29, 1988 Pages 40-47 PRELYSOSOMAL CONVERGENCEOF AUTOPHAGIC AND ENDOCYTIC PATHWAYS Paul B. Gordon and Per 0. Seglen Department of Tissue Culture, Institute for Cancer Research, The Norwegian Radium Hospital, Montebello, 0310 Oslo 3, Norway Received January 4, 1988 [14C]Lactose, electroinjected into the cytosol of isolated rat hepatocytes, was sequestered by autophagy, transferred to lysosomes and eventually hydrolysed. Asparagine prevented the fusion between prelysosomal autophagic vacuoles and lysosomes, and caused lactose to accumulate in the former. However, if the hepatocytes were simultaneously allowed to endocytose added B-galactosidase, no lactose accumulation occurred. These results suggest that autophagically sequestered lactose and endocytosed S-galactosidase were delivered to the same prelysosomal vacuole, where the lactose was hydrolysed by the enzyme. The name amphisome is suggested for this new functional compartment, common to the autophagic and endocytic pathways. 0 1988 Academic Press, Inc. Electroinjection of radioactive sugars into isolated hepatocytes has proved to be a useful technique for investigation and characterization of the autophagic-lysosomal pathway (1,2). Inert sugars like [14C]sucrose and C3H]raffinose are sequestered autophagically along with proteins and other cytoplasmic components, and can be used as probes of autophagic sequestration, the first step in the pathway (3,4). [14C1Lactose, on the other hand, is rapidly hydrolysed by lysosomal 6-galactosidase once it enters the lysosomes and can, therefore, be used to investigate post-sequestrational events (5). In previous work, we showed that autophagically sequestered sucrose, accumulating in lysosomes, could be hydrolysed upon endocytosis of added invertase, a sucrose-cleaving enzyme (2). Pre-accumulated lysosomal sucrose was completely hydrolysed even in the absence of ongoing autophagy, indicating that invertase- loaded endosomes could fuse directly with all lysosomes that had received autophagic material (6). Convergence of the autophagic 0006-291X/88 $1.50 Copyright 0 I988 by Academic Press, Inc. All rights of reproduction in any form reserved. 40