Identification and characterization of a mammalian protein interacting with 20S proteasome precursors

Abstract

The assembly of individual mammalian proteasome subunits into catalytically active 20S proteasome is not well understood. Herein, we report the identification and characterization of human and mouse homologues of the yeast proteasome maturating factor Ump1p. We delineate the region of hUMP1 implicated in the specific interaction with proteasome precursors and show that hUMP1 protein is absent from the mature form of the 20S proteasome. We also show that the transcript level of mammalian UMP1 is increased after IFN-␥ treatment and that mammalian UMP1 is functionally related to but not interchangeable with its yeast homologue. The proteasome is one of the major nonlysosomal proteases present in the cytosol and nucleus of cells. Through its catabolic functions, it is implicated in many cellular processes including the progression of cell cycle, the removal of misfolded proteins, apoptosis, and the production of peptides for presentation by MHC class I molecules (1-3). The proteasome is composed of a 20S barrel-shaped core particle, capped on each side by a 19S protein complex. Whereas the 20S proteasome can degrade short peptides, its association with the 19S caps forms the 26S proteasome, which is able to degrade multiubiquitylated protein substrates (4).The basic three-dimensional architecture of the 20S proteasome has been maintained from archaebacteria to humans and is characterized by four heptameric rings (5). The two outer rings are identical and are composed of ␣-subunits. The two inner rings, within which the catalytic centers are located, are also identical and are composed of ␤-subunits. Whereas the ␣-and ␤-rings of the archaebacterial 20S proteasome are formed of 7 identical ␣-subunits and 7 identical ␤-subunits, the more complex eukaryotic proteasomes contain at least 14 subunits, 7 distinct ␣-subunits, and 7 distinct ␤-subunits (6). In mammals, the existence of three additional ␤-subunits, which are expressed and incorporated into the 20S proteasome after IFN-␥ treatment, further increases the complexity of the proteasome composition. Contrary to the archaebacterial proteasome, in which all seven ␤-subunits are catalytically active, only three of the seven different ␤-subunits (␤1, ␤2, and ␤5) of the eukaryotic standard proteasome possess catalytic properties. In cells exposed to IFN-␥, these three catalytic subunits are replaced by three distinct ones (␤1i͞LMP2, ␤2i͞MECL-1, and ␤5i͞LMP7) resulting in the formation of a 20S proteasome termed immunoproteasome. The different cleavage specificity of the immunoproteasome compared with the standard proteasome has been shown to modulate the production of certain antigenic peptides presented by MHC class I molecules (2, 7).A host of sequential reactions is required to reach the complex quaternary structure of eukaryotic proteasomes. The active ␤-subunits carry N-terminal propeptides, which have been shown to play an important role, not only in the assembly and maturation of the 20S proteasome (8), but also to protect the N-terminal catalytic Thr of the matur...

Inhibition of Proteasome Activity Induces Concerted Expression of Proteasome Genes and de Novo Formation of Mammalian Proteasomes

“…In this context it is important to note that gene expression of mammalian Ump1/POMP-like factors appears to be differentially regulated as well. The mRNA levels of the human and mouse homologues are induced after treatment with interferon ␥, suggesting that these factors are somehow involved in immunoproteasome biogenesis (15)(16)(17). Here, we show that expression of mammalian POMP is also induced in response to proteasome inhibition, thereby facilitating the biogenesis pathway (Figs.…”

“…Ump1 is part of proteasome precursor complexes and is degraded upon completion of proteasome maturation (14). Mammalian Ump1 homologues have been identified in 16 S precursor complexes and are referred to as POMP (proteasome maturation protein), proteassemblin, or human/mouse UMP (8,(15)(16)(17).…”

RPN4 is a ligand, substrate, and transcriptional regulator of the 26S proteasome: A negative feedback circuit

“…One example is UMP1, a chaperone that is required for efficient assembly of the 20S proteasome and becomes its first substrate on the completion of assembly (22,53). Other examples are S. cerevisiae F-box proteins CDC4, GRR1, and MET30, which are short-lived substrate-recognition subunits of, respectively, SCF Cdc4 , SCF Grr1 , and SCF Met30 Ub ligases (54,55).…”

IFN-γ-induced immune adaptation of the proteasome system is an accelerated and transient response

“…Thus, degradation of Ump1p͞POMP signals the successful completion of the proteasome biogenesis program (17,20). Importantly, in mammalian cells, IFN-␥ was found to enhance POMP mRNA levels, suggesting that POMP may play an important role in i20S biogenesis (17,19).…”

“…A protein that is directly associated with proteasome precursor complexes is proteasome maturation protein (POMP), also named proteassemblin or human͞mouseUmp1 according to their yeast homologue Ump1 (17)(18)(19)(20). These factors are proposed to be involved in the coordinate processing of the ␤ subunits, thereby becoming the first substrate of the fully assembled and activated 20S proteasome.…”