Degradation of ornithine decarboxylase in reticulocyte lysate is ATP-dependent but ubiquitin-independent.
Reticulocyte lysate contains all the components of the ubiquitin-dependent proteolytic system. Several proteins are degraded in reticulocyte lysate in a ubiquitin-dependent manner. However, none of the proteins studied has a short intracellular half-life. We have investigated the degradation of ornithine decarboxylase (ODC), one of the most labile proteins in mammalian cells. ODC is efficiently degraded in reticulocyte lysate depleted of the ubiquitin activating enzyme, E1, in fraction II of reticulocyte lysate completely lacking ubiquitin, and in fraction II depleted of the entire complex of enzymes responsible for the ligation of ubiquitin to target proteins. The degradation of ODC is ATP dependent. Therefore, our results demonstrate that in addition to the ubiquitin-dependent proteolytic pathway, reticulocyte lysate contains at least one additional ATP-dependent proteolytic pathway. In vitro synthesized ODC served as a substrate in the present degradation study. Its successful utilization establishes a general strategy for investigating the degradation of short-lived proteins (for which a corresponding cDNA is available), that constitute a very small fraction of cellular proteins and for which purification is difficult or impossible. In contrast to ODC synthesized in vitro, that isolated from cells was not degraded by the reticulocyte lysate degradation system, suggesting that post-translational modifications may be involved in regulating ODC degradation.
Bercovich
Z
Z
Department of Virology, Weizmann Institute of Science, Rehovot, Israel.
Rosenberg-Hasson
Y
Y
Ciechanover
A
A
Kahana
C
C
D000255:Adenosine Triphosphate / Q000097:blood*; D000818:Animals; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics; D006418:Heme / Q000494:pharmacology; D007460:Iodoacetamide / Q000494:pharmacology; D007700:Kinetics; D051379:Mice; D009113:Muramidase / Q000378:metabolism; D009955:Ornithine Decarboxylase / Q000097:blood* / Q000235:genetics; D010618:Phenanthrolines / Q000494:pharmacology; D011817:Rabbits; D012156:Reticulocytes / Q000378:metabolism*; D014452:Ubiquitins / Q000494:pharmacology*
D010618:Phenanthrolines; D014452:Ubiquitins; D006418:Heme; D000255:Adenosine Triphosphate; D004247:DNA; D009113:Muramidase; D009955:Ornithine Decarboxylase; C025205:1,10-phenanthroline; D007460:Iodoacetamide