Studies on the Mechanisms of Autophagy: Maturation of the Autophagic Vacuole

• Published in: The Journal of cell biology Vol. 110; no. 6; pp. 1935 - 1945
• Main Author: Dunn, W. A.
• Format: Journal Article
• Language: English
• Published: New York, NY Rockefeller University Press 01.06.1990; The Rockefeller University Press
• Subjects: Animals; Antibodies; autophagy; Autophagy - physiology; Biological and medical sciences; Cell membranes; Cell metabolism, cell oxidation; Cell physiology; endoplasmic reticulum; Enzymes; Fundamental and applied biological sciences. Psychology; Hepatocytes; Hydrolases - metabolism; Immunohistochemistry; Liver; Liver - cytology; Liver - metabolism; Liver - ultrastructure; Lysosomes; Membrane proteins; Membrane Proteins - metabolism; Microscopy, Electron; Molecular and cellular biology; P branes; Phagocytosis - physiology; Proteins - metabolism; Rats; Rats, Inbred Strains; Receptor, IGF Type 2; Receptors; Receptors, Cell Surface - physiology; Vacuoles; Vacuoles - metabolism; Vacuoles - physiology; Vacuoles - ultrastructure; Immunogold technique; Membrane protein; Molecular processing; Hydrolase; Rat; Enzyme; Liver; Rodentia; Lysosome; Electron microscopy; Microsome; Immunoperoxidase technique; Characterization; Vertebrata; Mammalia; Immunoblotting assay; Carbohydrate; Biological receptor
• ISSN: 0021-9525; 1540-8140
• DOI: 10.1083/jcb.110.6.1935

Data presented in the accompanying paper suggests nascent autophagic vacuoles are formed from RER (Dunn, W. A. 1990. J. Cell Biol. 110:1923-1933). In the present report, the maturation of newly formed or nascent autophagic vacuoles into degradative vacuoles was examined using morphological and biochemical methods combined with immunological probes. Within 15 min of formation, autophagic vacuoles acquired acid hydrolases and lysosomal membrane proteins, thus becoming degradative vacuoles. A previously undescribed type of autophagic vacuole was also identified having characteristics of both nascent and degradative vacuoles, but was different from lysosomes. This intermediate compartment contained only small amounts of cathepsin L in comparison to lysosomes and was bound by a double membrane, typical of nascent vacuoles. However, unlike nascent vacuoles yet comparable to degradative vacuoles, these vacuoles were acidic and contained the lysosomal membrane protein, lgp120, at the outer limiting membrane. The results were consistent with the stepwise acquisition of lysosomal membrane proteins and hydrolases. The presence of mannose-6-phosphate receptor in autophagic vacuoles suggested a possible role of this receptor in the delivery of newly synthesized hydrolases from the Golgi apparatus. However, tunicamycin had no significant effect on the amount of mature acid hydrolases present in a preparation of autophagic vacuoles isolated from a metrizamide gradient. Combined, the results suggested nascent autophagic vacuoles mature into degradative vacuoles in a stepwise fashion: (a) acquisition of lysosomal membrane proteins by fusing with a vesicle deficient in hydrolytic enzymes (e.g., prelysosome); (b) vacuole acidification; and (c) acquisition of hydrolases by fusing with preexisting lysosomes or Golgi apparatus-derived vesicles.