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OBJECTIVES@#To investigate the effects and mechanisms of deubiquitinating enzyme Josephin domain containing 2 (JOSD2) on susceptibility of non-small cell lung carcinoma (NSCLC) cells to anti-cancer drugs.@*METHODS@#The transcriptome expression and clinical data of NSCLC were downloaded from the Gene Expression Omnibus. Principal component analysis and limma analysis were used to investigate the deubiquitinating enzymes up-regulated in NSCLC tissues. Kaplan-Meier analysis was used to investigate the relationship between the expression of deubiquitinating enzymes and overall survival of NSCLC patients. Gene ontology enrichment and gene set enrichment analysis (GSEA) were used to analyze the activation of signaling pathways in NSCLC patients with high expression of JOSD2. Gene set variation analysis and Pearson correlation were used to investigate the correlation between JOSD2 expression levels and DNA damage response (DDR) pathway. Western blotting was performed to examine the expression levels of JOSD2 and proteins associated with the DDR pathway. Immunofluorescence was used to detect the localization of JOSD2. Sulforhodamine B staining was used to examine the sensitivity of JOSD2-knock-down NSCLC cells to DNA damaging drugs.@*RESULTS@#Compared with adjacent tissues, the expression level of JOSD2 was significantly up-regulated in NSCLC tissues (P<0.05), and was significantly correlated with the prognosis in NSCLC patients (P<0.05). Compared with the tissues with low expression of JOSD2, the DDR-related pathways were significantly upregulated in NSCLC tissues with high expression of JOSD2 (all P<0.05). In addition, the expression of JOSD2 was positively correlated with the activation of DDR-related pathways (all P<0.01). Compared with the control group, overexpression of JOSD2 significantly promoted the DDR in NSCLC cells. In addition, DNA damaging agents significantly increase the nuclear localization of JOSD2, whereas depletion of JOSD2 significantly enhanced the sensitivity of NSCLC cells to DNA damaging agents (all P<0.05).@*CONCLUSIONS@#Deubiquitinating enzyme JOSD2 may regulate the malignant progression of NSCLC by promoting DNA damage repair pathway, and depletion of JOSD2 significantly enhances the sensitivity of NSCLC cells to DNA damaging agents.
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Humanos , Carcinoma de Pulmón de Células no Pequeñas/genética , Antineoplásicos/farmacología , Neoplasias Pulmonares/genética , Daño del ADN , ADN , Enzimas Desubicuitinizantes/genéticaRESUMEN
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.
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Objective To explore the antitumor small molecules targeting the ubiquitin–proteasome system (UPS) on the basis of active molecules from traditional Chinese medicine. Methods UbG76V-GFP stably expressing cell line was constructed to screen novel small molecule inhibitors targeting UPS. The fluorogenic substrates of Suc-LLVY-AMC, Z-LLE-AMC, and Boc-LRR-AMC were used to assess the effect of dioscin on the 20S proteasome hydrolase activity. The Ub-AMC substrate was used to evaluate the effect of dioscin on the intracellular deubiquitinating enzyme activity. Western blot was used to detect the effect of dioscin on intracellular ubiquitination levels. CCK-8 and colony formation assays were used to detect the inhibitory effect of dioscin on the tumor cell proliferation. Results Dioscin is a UPS inhibitor discovered through the UbG76V-GFP reporter system. It enhances intracellular ubiquitination and inhibits tumor cell proliferation and colony formation by targeting deubiquitinating enzymes. Conclusion Dioscin could significantly inhibit tumor cell proliferation by targeting ubiquitin–proteasome.
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Polycomb group (PcG) proteins are transcriptional repressors that control cell fate and the development of embryo, and they elicit function mainly in the form of polycomb repressive complex (PRC). Chromobox protein homolog 6 (CBX6) is one of the core protein subunits of PRC1, which plays an important role in gene expression regulation, cell renewal and differentiation, tumorigenesis and development, and stem cell maintenance. In this study, CBX6 was found to be degraded through a ubiquitin-proteasome dependent pathway. Then the gene expression library containing 92 deubiquitinating enzymes (DUB) was used to screen DUB targeting CBX6 and results found that ubiquitin-specific protease 29 (USP29) could obviously stabilize CBX6 protein level and extend its half-life (P < 0.05). Immunoprecipitation experiments found that CBX6 interacted with USP29 through its C-terminal domains; Further studies found that USP29 regulated the protein stability of CBX6 by deubiquitination in an enzymatic-activity dependent manner. Cell proliferation assay also found that USP29 inhibited the proliferation of MCF7 cells (P<0.0001). Taken together, through screening, this study found that USP29 could stabilize CBX6 protein level through deubiquitinating CBX6 and inhibit the cell proliferation of MCF7.
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Ubiquitin is a widespread form of post-translational modification of proteins in cells, which plays an important role in regulating immune response, inflammatory response, cell connection, cell cycle, apoptosis, DNA damage repair and many other life processes. The ubiquitin modification of proteins is also reversible. Deubiquitinating enzymes regulate the life span or function of proteins through hydrolyzing polyubiquitin chains to deubiquitinate substrate proteins, thus playing a role in ubiquitin-mediated signal transduction pathways. Ovarian tumor-associated proteases (OTUs) belong to the cysteine family. It has been found that many members of the OTUs family are closely related to the regulation of viral infection. This paper reviewed the role and mechanism of OTUs in host antivirus response in recent years.
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Cylindromatosis gene is a kind of tumor suppressor genes, whose mutation or deletion will lead to the development of a cylindrical tumor. The deubiquitinating enzyme CYLD protein encoded by it is a member of the deubiquitinating enzyme family. CYLD alters the function of the target molecules by removing the ubiquitin chain linked to the substrate protein K63, and participates in the regulation of signaling pathways, such as NF-κB, JNK and Wnt. This article reviews the recent year’s research progress of CYLD, especially its negative regulatory role in the progression of liver-related diseases.
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Ubiquitination and deubiquitination play important roles in the regulation of protein stability and function. Deubiquitinating enzymes (DUBs) are involved in the regulation of survival, migration and proliferation of cancer cells, by participating in a variety of signaling pathways. Most of the DUBs promote the malignant transformation and progression, while the others may function as tumor-suppressors. Given the central roles of DUBs in tumorigenesis and malignant progression, some of these enzymes have been regarded as promising anti-cancer targets. This paper reviews the recent advances in tumor-related DUBs and inhibitors.