Oncogene database

7375

USP4

ubiquitin specific peptidase 4 (proto-oncogene);USP4;ubiquitin specific peptidase 4 (proto-oncogene)

deubiquitinating enzyme 4|ubiquitin carboxyl-terminal esterase 4|ubiquitin carboxyl-terminal hydrolase 4|ubiquitin specific protease 4 (proto-oncogene)|ubiquitin thioesterase 4|ubiquitin thiolesterase 4|ubiquitin-specific processing protease 4|ubiquitous

3p21.3

protein-coding

We have identified a novel gene, USP15, encoding a human ubiquitin-specific protease (USP). The USP15 protein consists of 952 amino acids with a predicted molecular mass of 109.2 kDa and contains the highly conserved Cys and His boxes present in ALL members of the UBP family of deubiquitinating enzymes. USP15 shares 60.5% sequence identity and 76% sequence similarity with the human homolog (UNP/Unph/USP4) of the mouse Unp proto-oncogene. Recombinant USP15 demonstrated ubiquitin-specific protease activity against engineered linear fusions of ubiquitin to beta-galactosidase and glutathione S-transferase. USP15 can also cleave the ubiquitin-proline bond, a property previously unique to Unp/UNP. Chromosomal mapping by fluorescence in situ hybridization and radiation hybrid analyses localized the USP15 gene to chromosome band 12q14, a different location than that of UNP (3p21.3). Analysis of expressed sequence tag databases reveals evidence of alternate polyadenylation sites in the USP15 gene and also indicates that the gene may possess an exon/intron structure similar to that of the Unp gene, suggesting they have descended from a common ancestor. A systematic nomenclature for the human USPs is proposed.

The ubiquitin-specific proteases (Ubps) are a family of largely dissimilar enzymes with two major conserved sequence regions, containing either a conserved cysteine residue or two conserved histidine residues, respectively. The murine Unp oncoprotein and its human homologue, Unph, both contain regions similar to the conserved Cys and His boxes common to ALL the Ubps. In this study we show that Unp and Unph are active deubiquitinating enzymes, being able to cleave ubiquitin from both natural and engineered linear ubiquitin-protein fusions, including the polyubiquitin precursor. mutation of the conserved Unp Cys and His residues abolishes this activity, and identifies the likely His residue in the catalytic triad. Unp is tumorigenic when overexpressed in mice, leading to the suggestion that Unp may play a role in the regulation of ubiquitin-dependent protein degradation. We have demonstrated here that the high-level expression of Unp in yeast does not disrupt the degradation of the N-end rule substrate Tyr-beta-galactosidase (betagal), the non-N-end rule substrate ubiquitin-Pro-betagal, or the degradation of abnormal, canavanine-containing proteins. These data suggest that Unp is not a general modulator of ubiquitin-dependent proteolysis. However, Unp may have a role in the regulation of the degradation of a specific, as yet undescribed, substrate(s).

The oncogenic deubiquitylating enzyme (DUB) Unp/Usp4, which binds to the retinoblastoma family of tumor suppressor proteins, was originally described as a nuclear protein. However, more recent studies have shown it to be cytoplasmic. In addition, analysis of its subcellular localization has been complicated by the existence of the paralog Usp15. In this study, we resolved this controversy by investigating the localization of exogenously expressed Usp4 (using red fluorescent protein-Usp4) and of endogenous Usp4 (using highly specific antibodies that can distinguish Usp4 from Usp15). We found that by inhibiting nuclear export with leptomycin B, both exogenous and endogenous Usp4 accumulate in the nucleus. Further, using a Rev-green fluorescent protein-based export assay, we confirmed the existence of a nuclear export signal ((133)VEVYLLELKL(142)) in Usp4. In addition, a functional nuclear import signal ((766)QPQKKKK(772)) was also identified, which was specifically recognized by importin alpha/beta. Finally, we show that the equilibrium of Usp4 subcellular localization varies between different cell types. Usp4 is thus the first DUB reported to have nucleocytoplasmic shuttling properties. The implications of this shuttling for its function as a DUB are discussed.

UnpEL (also known as Usp4 or Unph) is an oncogenic protein, because its expression with a strong promoter results in the tumorigenic transformation of NIH3T3 cells injected into nude mice. Although the structure of UnpEL is that of a deubiquitinating enzyme, neither its precise function in mammalian cells nor the mechanism of UnpEL-mediated tumorigenesis is known. Here, we show that UnpEL functions as a deubiquitinating enzyme in human HEK293T cells and its isopeptidase activity deconjugates ubiquitin specifically from a UnpEL-interacting protein Ro52. We further show that UnpEL translocates to the cytoplasmic rod-like structures and colocalizes with Ro52 when Ro52 is overexpressed in HEK293 cells. These results suggest that UnpEL colocalizes with the unubiquitinated form of Ro52 to the cytoplasmic rod-like structures, where it keeps Ro52 unubiquitinated. The continuous deubiquitination of Ro52 might be involved in tumorigenesis.

The murine Unp proto-oncogene encodes a nuclear ubiquitin protease whose overexpression leads to oncogenic transformation of NIH3T3 cells. We have cloned cDNAs originating from the human homolog of this gene, designated herein as Unph (Unp, human), and have used these cDNAs to map the gene to 3p21.3, a region frequently rearranged in human tumor cells. Unph mRNA levels are consistently elevated in small cell tumors and adenocarcinomas of the lung, suggesting a possible causative role for the gene in the neoplastic process.

The mouse Unp gene is related to TRE17, a human oncogene, but is not its mouse homolog. Unp is a ubiquitously expressed gene producing two related mRNAs. The protein product of Unp is localized in the nucleus. expression of Unp from a highly active promoter results in tumorigenic transformation of NIH3T3 cells injected into athymic mice. Unp therefore encodes a novel nuclear oncoprotein.