DEGRADATION OF ORNITHINE DECARBOXYLASE BY THE MAMMALIAN AND YEAST 26SPROTEASOME COMPLEXES REQUIRES ALL THE COMPONENTS OF THE PROTEASE

Ornithine decarboxylase (ODC), a key enzyme in the biosynthesis of pol yamines, is an extremely short-lived protein. This attribute is import ant for the regulation of the activity of the enzyme and implies that the mechanisms involved in its degradation play an important role in t he control of the cellular processes in which the enzyme is involved. Recently, it has been shown that ODC is degraded by the 26S proteasome complex in a process that requires antizyme, but not ubiquitin. With one reported exception, ODC, the 26S complex recognizes and degrades s pecifically ubiquitinated proteins. Their unconjugated counterparts ar e not targeted. The 26S complex is composed of a core catalytic unit, the 20S proteasome complex, and two additional, and apparently distinc t, subcomplexes. The two additional subcomplexes are regulatory subuni ts that are required in order to confer specificity and control. In th is study, we demonstrate that, like the degradation of ubiquitin-conju gated proteins, ubiquitin-independent degradation of ODC also requires prior assembly of the mammalian 26S proteasome from all the three sub units, the 20S proteasome and the two subcomplexes. The combination of any two subunits does not support generation of a proteolytically act ive complex. This is also true for the yeast 26S complex. Like the mam malian 20S proteasome, the yeast 20S complex can cleave short peptides in an ATP-independent mode, but cannot degrade ODC or ubiquitin-conju gated proteins. These proteins are degraded only following addition of the regulatory subunits and generation of the high-molecular-mass 26S complex. In a distinct, but related, set of experiments, we demonstra te that the degradation of ODC by the assembled 26S proteasome in vitr o reproduces faithfully proteolysis of the enzyme in the intact cell. Namely, (a) a C-terminal-deleted mouse ODC and trypanosome ODC are sta ble both in vitro and in vivo, and (b) like proteolysis in the intact cell, degradation in the reconstituted cell-free system is also depend ent upon the addition of antizyme.