title: Inhibition of pyruvate carboxylase degradation and total protein breakdown by lysosomotropic agents in 3T3-L1 cells.
abstract: 1. Exposure to [3H]biotin during the differentiation of 3T3-L1 cells to adipocytes selectively labelled pyruvate carboxylase (EC 6.4.1.1). A subsequent incubation of labelled cells permitted the measurement of the degradation rate constant of this mitochondrial enzyme. 2. In medium without serum, pyruvate carboxylase was degraded with a half-life of 64 h, considerably longer than that found for average cell protein. The long half-life is commensurate with the enzyme being catabolized when whole mitochondria are destroyed. 3. The breakdown of pyruvate carboxylase was inhibited to a greater extent than the breakdown of total cell protein by insulin, NH4Cl and inhibitors of lysosomal proteinases, suggesting that the enzyme is degraded by the autophagic lysosomal system of the cell. 4. The above evidence implies that whole mitochondria are degraded in lysosomes, a conclusion that agrees with earlier electron-microscopic evidence showing mitochondria within autophagic vacuoles. 5. A second degradative pathway must be invoked to account for the breakdown of mitochondrial proteins of short half-life.
mesh_terms: D000273:Adipose Tissue / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D000643:Ammonium Chloride / Q000494:pharmacology; D000818:Animals; D001710:Biotin / Q000378:metabolism; D001769:Blood; D002478:Cells, Cultured; D003513:Cycloheximide / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D007328:Insulin / Q000494:pharmacology; D008247:Lysosomes / Q000378:metabolism*; D051379:Mice; D011506:Proteins / Q000378:metabolism*; D011766:Pyruvate Carboxylase / Q000037:antagonists & inhibitors*
chemical_list: D007328:Insulin; D011506:Proteins; D000643:Ammonium Chloride; D001710:Biotin; D003513:Cycloheximide; D011766:Pyruvate Carboxylase