Deubiquitination complex platform: A plausible mechanism for regulating the substrate specificity of deubiquitinating enzymes

Deubiquitinating enzymes (DUBs) or deubiquitinases facilitate the escape of multiple proteins from ubiquitin‒proteasome degradation and are critical for regulating protein expression levels in vivo. Therefore, dissecting the underlying mechanism of DUB recognition is needed to advance the development of drugs related to DUB signaling pathways. To data, extensive studies on the ubiquitin chain specificity of DUBs have been reported, but substrate protein recognition is still not clearly understood. As a breakthrough, the scaffolding role may be significant to substrate protein selectivity. From this perspective, we systematically characterized the scaffolding proteins and complexes contributing to DUB substrate selectivity. Furthermore, we proposed a deubiquitination complex platform (DCP) as a potentially generic mechanism for DUB substrate recognition based on known examples, which might fill the gaps in the understanding of DUB substrate specificity.

Research Progress of the Roles of Ubiquitination/Deubiquitination in Androgen Receptor Abnormalities and Prostate Cancer / 中国医学科学院学报

Ubiquitin is a small molecule protein consisting of 76 amino acids,widely found in eukaryotic cells. The process by which ubiquitin binding to a specific protein is called ubiquitination. Deubiquitination is the reversed process of ubiquitination. Ubiquitination stimulates downstream signal,including complex assembly,protein conformation and activity changes,proteolysis,autophagy,guilt,chromatin remodeling,and DNA repair. More than 80% of eukaryotic protein degradation is mediated by the ubiquitination system,and ubiquitin-dependent proteolysis is an extremely complex process involving many biomolecular processes. By regulating protein homeostasis,ubiquitination can also regulate a variety of biological processes including cell cycle,cell proliferation,and apoptosis,which are closely related to tumorigenesis and progression. Many abnormalities of androgen receptor (AR) including AR gene amplification,mutation,shear mutation,and AR activity enhancement are closely related to prostate cancer progression. In particular,prostate cancer progression is regulated by the ubiquitination/deubiquitination processes. This article summarizes the recent research advances in the roles of ubiquitination/deubiquitination in AR abnormalities and prostate cancer.

Discovery of 1,2,3triazolo4,5-pyrimidine derivatives as highly potent, selective, and cellularly active USP28 inhibitors

Ubiquitin specific peptidase 28 (USP28) is closely associated to the occurrence and development of various malignancies, and thus has been validated as a promising therapeutic target for cancer therapy. To date, only few USP28 inhibitors with moderate inhibitory activity have been reported, highly potent and selective USP28 inhibitors with new chemotypes remain to be discovered for pathologically investigating the roles of deubiquitinase. In this current study, we reported the synthesis and biological evaluation of new [1,2,3]triazolo[4,5-]pyrimidine derivatives as potent USP28 inhibitors. Especially, compound potently inhibited USP28 (IC = 1.10 ± 0.02 μmol/L,  = 40 nmol/L), showing selectivity over USP7 and LSD1 (IC > 100 μmol/L). Compound was cellularly engaged to USP28 in gastric cancer cells. Compound reversibly bound to USP28 and directly affected its protein levels, thus inhibiting the proliferation, cell cycle at S phase, and epithelial-mesenchymal transition (EMT) progression in gastric cancer cell lines. Docking studies were performed to rationalize the potency of compound . Collectively, compound could serve as a new tool compound for the development of new USP28 inhibitors for exploring the roles of deubiquitinase in cancers.

The role of ubiquitination and deubiquitination in the regulation of cell junctions

Maintenance of cell junctions plays a crucial role in the regulation of cellular functions including cell proliferation, permeability, and cell death. Disruption of cell junctions is implicated in a variety of human disorders, such as inflammatory diseases and cancers. Understanding molecular regulation of cell junctions is important for development of therapeutic strategies for intervention of human diseases. Ubiquitination is an important type of post-translational modification that primarily regulates endogenous protein stability, receptor internalization, enzyme activity, and protein-protein interactions. Ubiquitination is tightly regulated by ubiquitin E3 ligases and can be reversed by deubiquitinating enzymes. Recent studies have been focusing on investigating the effect of protein stability in the regulation of cell-cell junctions. Ubiquitination and degradation of cadherins, claudins, and their interacting proteins are implicated in epithelial and endothelial barrier disruption. Recent studies have revealed that ubiquitination is involved in regulation of Rho GTPases' biological activities. Taken together these studies, ubiquitination plays a critical role in modulating cell junctions and motility. In this review, we will discuss the effects of ubiquitination and deubiquitination on protein stability and expression of key proteins in the cell-cell junctions, including junction proteins, their interacting proteins, and small Rho GTPases. We provide an overview of protein stability in modulation of epithelial and endothelial barrier integrity and introduce potential future search directions to better understand the effects of ubiquitination on human disorders caused by dysfunction of cell junctions.

Research advance on BAP1 and its function in tumor suppression / 上海交通大学学报（医学版）

BAP1 is a member of the ubiquitin C-terminal hydrolases (UCH) subfamily of deubiquitylases with basic function of removing monoubiquitin or ubiquitin chains from the specific substrate proteins. As well, it is a key factor in regulating gene expression, cell cycle, cell differentiation,cell apoptosis and DNA damage response, dependent or independent of its deubiquitination function. Evidences haverevealed that mutation or downregulation of BAP1 can prominently increase the occurrence of cancers, including uveal melanoma, mesothelioma, renal cell carcinoma, breast cancer and lung cancer. Currently, the tumor spectrum and the pathogenic mechanism on BAP1 have not been illustrated clearly, and need to be further researched,which might bring a new opportunity in treatment of cancers.

Correlation between ubiquitin specific peptidase 7 expression and male infertility / 第二军医大学学报

Objective To investigate the expression of ubiquitin specific peptidase 7 (USP7) in the sperm of infertile man suffering asthenozoospermia or oligoasthenozoospermia, and to explore its role in the pathogenesis of male infertility. Methods The semen samples were obtained from 120 donors attending the Fertility Clinic, the Centre of Reproductive Medicine, the Second Affiliated Hospital, Chongqing Medical University. The sperm specimens were divided into normozoospermia group (n=57), asthenozoospermia group (n =37) or oligoasthenozoospermia group (n =26) after semen analysis according to 2010 WHO standards. The localization and expression of USP7 were examined using immunofluorescence and Western blotting analysis in the three groups. Then the correlation between USP7 expression and sperm parameterswas analyzed. Results USP7 mainly localized in the middle part and main part of thesperm tail; the USP expression was significantly lower in asthenozoospermia group(0. 86 ± 0. 53) and oligoasthenozoospermia group (0. 62 ± 0. 43) compared with normozoospermia group (1. 63 ± 0. 76, P<0. 01). And USP7 expression was found positively correlated with the ratio of sperm moving forward(r = 0. 431 2, P =0. 008 7), total motility rate(r=0. 443 8, P =0. 006 7), and sperm concentration(r = 0. 455 0, P =0. 005 3). Conclusion The lowered expression of USP7 is related to the reduction of human sperm motility and concentration, indicating that USP7 expressionmay serve as an indicator for sperm quality evaluation in infertile men.

USP inhibitors and screening methods / 中国生化药物杂志

The ubiquitin-specific protease ( USP) inhibitors influence many crucial cellular activities and some immune processes, such as anti-inflammatory, anti-infection and anti-tumor by silencing the functions of USP.The main USP inhibitors, which potency and specificity are underlined and current methods for detecting and identifying USP inhibitors are discussed of in this review.

Conservation and developmental expression of ubiquitin isopeptidases in Schistosoma mansoni

Several genes related to the ubiquitin (Ub)-proteasome pathway, including those coding for proteasome subunits and conjugation enzymes, are differentially expressed during the Schistosoma mansoni life cycle. Although deubiquitinating enzymes have been reported to be negative regulators of protein ubiquitination and shown to play an important role in Ub-dependent processes, little is known about their role in S. mansoni . In this study, we analysed the Ub carboxyl-terminal hydrolase (UCHs) proteins found in the database of the parasite’s genome. An in silico ana- lysis (GeneDB and MEROPS) identified three different UCH family members in the genome, Sm UCH-L3, Sm UCH-L5 and Sm BAP-1 and a phylogenetic analysis confirmed the evolutionary conservation of the proteins. We performed quantitative reverse transcription-polymerase chain reaction and observed a differential expression profile for all of the investigated transcripts between the cercariae and adult worm stages. These results were corroborated by low rates of Z-Arg-Leu-Arg-Gly-Gly-AMC hydrolysis in a crude extract obtained from cercariae in parallel with high Ub conjugate levels in the same extracts. We suggest that the accumulation of ubiquitinated proteins in the cercaria and early schistosomulum stages is related to a decrease in 26S proteasome activity. Taken together, our data suggest that UCH family members contribute to regulating the activity of the Ub-proteasome system during the life cycle of this parasite.

Advances in the study of deubiquitinating enzymes in cancer research / 中国癌症杂志

Deubiquitinating enzymes, reversing protein ubiquitination, most of the researches focused on the ifeld of molecular biology. However, it have not yet attracted enough attention in translational medicine research. In fact, target proteins of deubiquitinating enzymes affect the tumor progression through various ways, for example, cell apoptosis and autophagy, the link between inlfammation and cancer, tumor hypoxia, signal transduction, cell cycle regulation and DNA damage. This paper reviewed the research progress on the relations between deubiquitinating enzymes and the correlated factors of tumor.

The Molecular Biologic Study about the Role of Deubiquitination Gene in Bladder Tumor Progression / 대한비뇨기과학회지

PURPOSE: Ubiqutin-proteasome-mediated proteolysis is an important pathway of nonlysosomal protein degradation, which controls the timed destruction of cellular regulatory proteins of the p21, p27, p53 gene (p53), Retinoblastoma gene (Rb), E2F-1, E2F-4 and cyclin etc. The deubiqutination (DUB) gene prevents ubiquitin-proteasome- mediated proteolysis by removal of ubiquitin from ubiqutinated proteins. We investigated the role of the DUB gene (DFFRY or DFFRX) in the progression of a bladder tumor associated with Rb and p53, which follows ubiquitin-proteasome-mediated proteolysis. MATERIALS AND METHODS: RT4 (bladder pailloma cell line), 5637 (superficial bladder transitional cell carcinoma cell line), and HT1376 (invasive bladder transitional cell carcinoma cell line) were cultured. mRNA was extracted from each cell line by the Poly-A tract mRNA isolation system. Relative quantitative RT-PCR (reverse transcription-polymerase chain reaction) for the DUB gene (DFFRY or DFFRX) was performed. Northern blotting for the DUB gene, p53 and Rb were performed. Proteins were harvested at 80% confluence from each cell line. Western blotting for the p53 and Rb were performed. RESULTS: The relative quantitative RT-PCR for the DUB gene showed it to be expressed most strongly in RT4, and most weakly in HT1376, which were identical to the findings of the Northern blotting. The p53 was expressed at similar levels on the Northern blots of all three cell lines, whereas on Western blotting, it was expressed most strongly in 5637, most weakly in RT4, and moderately in HT1376. The Rb was expressed only in RT4 on Northern and Western blotting. CONCLUSIONS: The DUB gene might be related to the inhibition of the progression of bladder tumors associated with Rb and p53, which follows ubiquitin-proteasome- mediated proteolysis.