Role of bestatin-Sensitive Exopeptidases in the Intracellular Degradation of Hepatic Proteins

Abstract

Injection of bestatin into intact mice produces accumulation of di- and tripeptide intermediates in the degradation of short- and long-lived hepatic proteins, whereas lysosomal breakdown of endocytosed plasma asialoglycoproteins is not affected. The majority of the peptides are found in the liver cytosol, but a minor portion appears in a sedimentable fraction containing mitochondria and lysosomes (Botbol, V., and Scornik, O. A. (1983) J. Biol. Chem. 258, 1942-1949). We now report that (a) the primary location of the intermediates is the cytosol. The particulate fraction represents cytosolic peptides trapped within mitochondria, as evidenced by sedimentation equilibrium in sucrose gradients after loading lysosomes with Triton WR1339 and by the sensitivity of the particles to lysis by digitonin. (b) In isolated hepatocytes, where we can measure simultaneously protein breakdown and bestatin-induced peptides, the accumulation of intermediates parallels protein degradation of analog-containing, short- and long-lived proteins, even after stimulation of the latter by amino acid deprivation. These observations are consistent with the hypothesis that bestatin inhibits cytosolic exopeptidases that complete the intracellular breakdown to amino acids of the major classes of hepatic proteins. The role of cytosolic exopeptidases is expected in the rapid degradation of abnormal proteins, a demonstrated cytosolic process. In stimulated degradation of long-lived proteins, the importance of cytosolic exopeptidases implies either that this process is largely cytosolic or, more likely, that peptides escape from autophagic organelles.