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Advanced Science (Weinheim,... Mar 2024Various forms of programmed cell death (PCD) exhibit distinct characteristics depending on their specific molecular mechanisms, and there are interactions among these...
Various forms of programmed cell death (PCD) exhibit distinct characteristics depending on their specific molecular mechanisms, and there are interactions among these different forms. Ferroptosis, which is related to autophagy and apoptosis, has an unknown potential interaction with pyroptosis. This study revealed a mutually antagonistic relationship between ferroptosis and pyroptosis, with 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGCR) playing a key role in their interaction. It is found that HMGCR predominantly localized to mitochondria during ferroptosis but shifted to the endoplasmic reticulum following treatment with a pyroptosis inducer. Furthermore, this study demonstrated that BRCC36 (BRCA1/BRCA2-containing complex subunit 36) deubiquitinated HMGCR in a manner dependent on deubiquitinating enzyme (DUB) activity, and inhibited ferroptosis and promoted pyroptosis. Moreover, as an oncogene in hepatocellular carcinoma (HCC), BRCC36 promoted cancer cell proliferation, migration, invasion, and tumor growth. Thiolutin, an inhibitor of BRCC36, effectively suppressed the interaction between BRCC36 and HMGCR, leading to the inhibition of HCC growth. Therefore, targeting BRCC36 can offer a novel and promising therapeutic strategy for HCC treatment. In conclusion, these findings provide new theoretical evidence for further characterizing tumor heterogeneity and offer new molecular targets for the diagnosis and treatment of HCC.
Topics: Humans; Pyroptosis; Carcinoma, Hepatocellular; Oxidoreductases; Ferroptosis; Liver Neoplasms; Hydroxymethylglutaryl CoA Reductases
PubMed: 38178583
DOI: 10.1002/advs.202304263 -
Journal of Thoracic Disease Jul 2023Lung adenocarcinoma (LUAD) is a prevalent subtype of lung cancer associated with high mortality rates. We aimed to utilize single-cell multiomics analysis to identify...
BACKGROUND
Lung adenocarcinoma (LUAD) is a prevalent subtype of lung cancer associated with high mortality rates. We aimed to utilize single-cell multiomics analysis to identify the key molecules involved in ubiquitination modification, which plays a role in LUAD development and progression.
METHODS
We use a systematic approach to analyze LUAD-related single-cell and bulk transcriptome datasets from Gene Expression Omnibus (GEO) and The Cancer Genome Atlas (TCGA) databases. Single-cell RNA sequencing (scRNA-seq) data were normalized, clustered, and annotated with the Seurat package in R. InferCNV was used to distinguish malignant from epithelial cells, and AUCell evaluated the area under the curve (AUC) score of ubiquitination-related enzymes. Survival and differential analyses identified significant molecular markers associated with ubiquitination. expression was confirmed using reverse-transcription quantitative polymerase chain reaction (RT-qPCR) and Western blot assays, and its knockdown cell lines were assessed for effects on cellular processes and tumor formation in mice. 's interacting proteins were predicted, and its impact on AGR2 protein half-life and ubiquitination was evaluated. Rescue experiments involving overexpression and silencing assessed their impact on malignant behaviors.
RESULTS
By means of single-cell sequencing analysis, we probed the ubiquitination modification landscape in the LUAD microenvironment. Malignant cells had elevated scores for enzymes and ubiquitin-binding domains compared to normal epithelial cells, with 53 ubiquitination-related molecules showing prognostic disparities. , , and were identified as genes with prognostic significance, with showing higher expression in epithelial and malignant cells. Two missense mutation sites were identified in , which had a high copy number amplification ratio and positive correlation with messenger RNA (mRNA) expression. expression and tumor stage were found to be independent prognostic factors, and interfering with expression reduced the malignant behavior of LUAD cells. was found to bind to AGR2 protein and reduce its ubiquitination, leading to increased stability. Knockdown of inhibited the enhancement of cell viability, invasion, and migration resulting from overexpression.
CONCLUSIONS
This study examined ubiquitination modifications in LUAD using sequencing data, identifying 's critical role in malignancy regulation and its potential as a prognostic and therapeutic biomarker. These insights enhance understanding of LUAD mechanisms and treatment.
PubMed: 37559628
DOI: 10.21037/jtd-23-795 -
Cell Death & Disease Sep 2023Ovarian tumor family deubiquitinase 4 (OTUD4), a member of the OTU deubiquitinating enzyme, is implicated to decrease in cancer to regulate cell apoptosis. However, the...
Ovarian tumor family deubiquitinase 4 (OTUD4), a member of the OTU deubiquitinating enzyme, is implicated to decrease in cancer to regulate cell apoptosis. However, the role of OTUD4 in cigarette smoke induced epithelial cell apoptosis and its mechanism have not been elucidated. In this study, we showed that OTUD4 protein reduced in CSE treated mice and airway epithelial cells. OTUD4 silence aggravated cell apoptosis and emphysematous change in the lung tissue of cigarette smoke extract (CSE) treated mice. Additionally, restoration of OTUD4 in the lung of mice alleviated CSE induced apoptosis and emphysematous morphology change. The effect of OTUD4 on cell apoptosis was also confirmed in vitro. Through protein profile screening, we identified that OTUD4 may interact with plasminogen activator inhibitor 1(PAI-1). We further confirmed that OTUD4 interacted with PAI-1 for de-ubiquitination and inhibiting CSE induced PAI-1 degradation. Furthermore, the protective role of OTUD4 in airway epithelial cells apoptosis was blocked by PAI-1 deactivation. Taken together, our data suggest that OTUD4 regulates cigarette smoke (CS)-triggered airway epithelial cell apoptosis via modulating PAI-1 degradation. Targeting OUTD4/PAI-1 signaling might potentially provide a therapeutic target against the lung cell apoptosis in cigarette smoke (CS)-induced emphysema.
Topics: Animals; Mice; Female; Humans; Plasminogen Activator Inhibitor 1; Ovarian Neoplasms; Apoptosis; Epithelial Cells; Lung; Ubiquitin-Specific Proteases
PubMed: 37726265
DOI: 10.1038/s41419-023-06131-1 -
Nucleic Acids Research Nov 2023Post-translational modifications of histones are important regulators of the DNA damage response (DDR). By using affinity purification mass spectrometry (AP-MS) we...
Post-translational modifications of histones are important regulators of the DNA damage response (DDR). By using affinity purification mass spectrometry (AP-MS) we discovered that genetic suppressor element 1 (GSE1) forms a complex with the HDAC1/CoREST deacetylase/demethylase co-repressor complex. In-depth phosphorylome analysis revealed that loss of GSE1 results in impaired DDR, ATR signalling and γH2AX formation upon DNA damage induction. Altered profiles of ATR target serine-glutamine motifs (SQ) on DDR-related hallmark proteins point to a defect in DNA damage sensing. In addition, GSE1 knock-out cells show hampered DNA damage-induced phosphorylation on SQ motifs of regulators of histone post-translational modifications, suggesting altered histone modification. While loss of GSE1 does not affect the histone deacetylation activity of CoREST, GSE1 appears to be essential for binding of the deubiquitinase USP22 to CoREST and for the deubiquitination of H2B K120 in response to DNA damage. The combination of deacetylase, demethylase, and deubiquitinase activity makes the USP22-GSE1-CoREST subcomplex a multi-enzymatic eraser that seems to play an important role during DDR. Since GSE1 has been previously associated with cancer progression and survival our findings are potentially of high medical relevance.
Topics: Cell Nucleus; Co-Repressor Proteins; Deubiquitinating Enzymes; DNA Damage; Histones; Humans; Animals; Mice; Cell Line
PubMed: 37878419
DOI: 10.1093/nar/gkad911 -
American Journal of Physiology. Cell... Oct 2023Breast cancer has become the malignant disease with the highest morbidity and mortality among female cancer patients. The prognosis of metastatic breast cancer is very...
Breast cancer has become the malignant disease with the highest morbidity and mortality among female cancer patients. The prognosis of metastatic breast cancer is very poor, and the therapeutic effects still need to be improved. The molecular mechanism of breast cancer has not been fully clarified. Bioinformatics analysis was used to find the differentially expressed gene that affects the occurrence and development of breast cancer. Furthermore, scratch assays, Transwell assays, immunofluorescence, and Western blotting were used to determine the biological behavior of breast cancer cells affected by DEP domain-containing protein 1B (DEPDC1B). The molecular mechanism was investigated by mass spectrometry analysis, coimmunoprecipitation, and ubiquitin assays. Here, we found that DEPDC1B was highly expressed in breast cancer cells and tissues and was associated with lower overall survival (OS) in patients. We found that DEPDC1B interference significantly inhibited tumor invasion and migration in vitro and tumor metastasis in vivo. Mechanistically, DEPDC1B was first shown to activate the wnt/β-catenin signaling pathway as an oncogene in breast cancer cells. In addition, we also confirmed the interaction between DEPDC1B, ubiquitin-specific protease 5 (USP5), and β-catenin. Then, we found that DEPDC1B mediates the deubiquitination of β-catenin via USP5, which promotes cell invasion and migration. Our findings provide new insights into the carcinogenic mechanism of DEPDC1B, suggesting that DEPDC1B can be considered a potential therapeutic target for breast cancer. By using bioinformatics analysis and the experimental techniques of cell biology and molecular biology, we found that DEP domain-containing protein 1B (DEPDC1B) can promote the invasion and migration of breast cancer cells and that DEPDC1B mediates the deubiquitination of β-catenin by ubiquitin-specific protease 5 (USP5), thus activating the wnt/β-catenin pathway. Our findings provide new insights into the carcinogenic mechanism of DEPDC1B, suggesting that DEPDC1B can be used as a potential therapeutic target for breast cancer.
Topics: Humans; Female; Breast Neoplasms; beta Catenin; Wnt Signaling Pathway; Ubiquitin-Specific Proteases; GTPase-Activating Proteins; Melanoma, Cutaneous Malignant
PubMed: 37642235
DOI: 10.1152/ajpcell.00249.2023 -
Cancer Science Aug 2023Ubiquitin-specific peptidase 24 (USP24), a member of the deubiquitinase family, plays an important role in tumor regulation. However, the role of USP24 in gastric cancer...
Ubiquitin-specific peptidase 24 (USP24), a member of the deubiquitinase family, plays an important role in tumor regulation. However, the role of USP24 in gastric cancer (GC) is unknown. The aim of our study was to explore the role of USP24 in GC to seek new therapeutic targets for GC. TCGA analysis showed that USP24 was upregulated in GC patients, and its high expression levels were associated with poor prognosis. It was found that overexpressed USP24 promoted GC cell proliferation and abnormal glycolysis. Further analysis and study showed that USP24 could promote the stability and increase the expression of oncogene PLK1. Knocking down USP24 can reduce the stability of PLK1 to reduce Notch 1 activity, thus inhibiting GC glycolysis, proliferation, and other processes and alleviating tumor progression. Knocking down USP24 can inhibit GC progression. In conclusion, USP24 was upregulated in GC and promoted the growth and glycolysis of GC cells, the mechanism of which was related to the deubiquitination of PLK1 and the increase of its stability. Silencing USP24 inhibited the GC process. This study suggests that the USP24/PLK1/Notch 1 axis may be a novel therapeutic target for gastric cancer.
Topics: Humans; Stomach Neoplasms; Oncogenes; Cell Proliferation; Ubiquitin-Specific Proteases; Carcinoma; Cell Line, Tumor; Glycolysis; Gene Expression Regulation, Neoplastic; Receptor, Notch1; Ubiquitin Thiolesterase
PubMed: 37265030
DOI: 10.1111/cas.15847 -
USP4 promotes the proliferation and glucose metabolism of gastric cancer cells by upregulating PKM2.PloS One 2023The pyruvate kinase enzyme PKM2 catalyzes the final step in glycolysis and converts phosphoenolpyruvate (PEP) to pyruvate. PKM2 is often overexpressed in cancer and...
BACKGROUND
The pyruvate kinase enzyme PKM2 catalyzes the final step in glycolysis and converts phosphoenolpyruvate (PEP) to pyruvate. PKM2 is often overexpressed in cancer and plays a role in the Warburg effect. The expression of PKM2 can be regulated at different levels. While it has been proven that PKM2 can be regulated by ubiquitination, little is known about its de-ubiquitination regulation.
METHODS
Immunoprecipitation was applied to identify the PKM2 interaction protein and to determine the interaction region between PKM2 and USP4. Immunofluorescence was performed to determine the cellular localization of USP4 and PKM2. The regulation of PKM2 by USP4 was examined by western blot and ubiquitination assay. MTT assays, glucose uptake, and lactate production were performed to analyze the biological effects of USP4 in gastric cancer cells.
RESULTS
USP4 interacts with PKM2 and catalyzes the de-ubiquitination of PKM2. Overexpression of USP4 promotes cell proliferation, glucose uptake, and lactate production in gastric cancer cells. Knockdown of USP4 reduces PKM2 levels and results in a reduction in cell proliferation and the glycolysis rate.
CONCLUSIONS
USP4 plays a tumor-promoting role in gastric cancer cells by regulating PKM2.
Topics: Humans; Stomach Neoplasms; Cell Proliferation; Lactic Acid; Glucose; Ubiquitin-Specific Proteases
PubMed: 37624791
DOI: 10.1371/journal.pone.0290688 -
MedComm Aug 2023Epigenetic regulators and posttranslational modifications of proteins play important roles in various kinds of cancer cell death, including ferroptosis, a non-apoptotic...
Epigenetic regulators and posttranslational modifications of proteins play important roles in various kinds of cancer cell death, including ferroptosis, a non-apoptotic form of cell death. However, the interplay of chromatin modifiers and deubiquitinase (DUB) in ferroptosis remains unclear. Here, we found that ubiquitin-specific protease 5 (USP5) is regarded as a bona fide DUB of lymphoid-specific helicase (LSH), a DNA methylation repressor, in hepatocellular carcinoma (HCC). Functional studies reveal that USP5 interacts with LSH and stabilizes LSH by a deubiquitylation activity-dependent process. Furthermore, the USP5-mediated deubiquitination of LSH facilitates the tumorigenesis of HCC by upregulating solute carrier family 7 member 11 (SLC7A11) to suppress ferroptosis of liver cancer cells. Moreover, the USP5 inhibitor degrasyn inhibits DUB activities of USP5 to LSH to suppress the progression of HCC. Additionally, USP5 and LSH are positively correlated and both are overexpressed and linked to poor prognosis in HCC patients. Together, our findings show that USP5 interacts with LSH directly and enhances LSH protein stability through deubiquitination, which, in turn, promotes the development of HCC by suppressing ferroptosis of liver cancer cells, suggesting that USP5 may be a potential therapeutic target for HCC.
PubMed: 37492786
DOI: 10.1002/mco2.337 -
Frontiers in Endocrinology 2023The malignant lung cancer has a high morbidity rate and very poor 5-year survival rate. About 80% - 90% of protein degradation in human cells is occurred through the... (Review)
Review
The malignant lung cancer has a high morbidity rate and very poor 5-year survival rate. About 80% - 90% of protein degradation in human cells is occurred through the ubiquitination enzyme pathway. Ubiquitin ligase (E3) with high specificity plays a crucial role in the ubiquitination process of the target protein, which usually occurs at a lysine residue in a substrate protein. Different ubiquitination forms have different effects on the target proteins. Multiple short chains of ubiquitination residues modify substrate proteins, which are favorable signals for protein degradation. The dynamic balance adapted to physiological needs between ubiquitination and deubiquitination of intracellular proteins is beneficial to the health of the organism. Ubiquitination of proteins has an impact on many biological pathways, and imbalances in these pathways lead to diseases including lung cancer. Ubiquitination of tumor suppressor protein factors or deubiquitination of tumor carcinogen protein factors often lead to the progression of lung cancer. Ubiquitin proteasome system (UPS) is a treasure house for research and development of new cancer drugs for lung cancer, especially targeting proteasome and E3s. The ubiquitination and degradation of oncogene proteins with precise targeting may provide a bright prospect for drug development in lung cancer; Especially proteolytic targeted chimerism (PROTAC)-induced protein degradation technology will offer a new strategy in the discovery and development of new drugs for lung cancer.
Topics: Humans; Proteasome Endopeptidase Complex; Lung Neoplasms; Ubiquitination; Ubiquitin; Proteins; Cell Transformation, Neoplastic; Drug Discovery
PubMed: 37795365
DOI: 10.3389/fendo.2023.1220108 -
Journal of Molecular Biology Dec 2023Deubiquitinases (DUBs) are proteolytic enzymes that catalyze the removal of ubiquitin from protein substrates. The critical role of DUBs in regulating protein...
Deubiquitinases (DUBs) are proteolytic enzymes that catalyze the removal of ubiquitin from protein substrates. The critical role of DUBs in regulating protein ubiquitination makes them attractive drug targets in oncology, neurodegenerative disease, and antiviral development. Biochemical assays for quantifying DUB activity have enabled characterization of substrate preferences and discovery of small molecule inhibitors. However, assessing the efficacy of these inhibitors in cellular contexts to support clinical drug development has been limited by a lack of tractable cell-based assays. To address this gap, we developed a two-color flow cytometry-based assay that allows for sensitive quantification of DUB activity and inhibition in living cells. The utility of this system was demonstrated by quantifying the potency of GRL0617 against the viral DUB SARS-CoV-2 PL, identifying potential GRL0617 resistance mutations, and performing structure-function analysis of the vOTU domain from the recently emerged Yezo virus. In addition, the system was optimized for cellular DUBs by modifying a GFP-targeting nanobody to recruit USP7 and USP28 to benchmark a panel of reported inhibitors and assess inhibition kinetics. Together, these results demonstrate the utility of these assays for studying DUB biology in a cellular context with potential to aid in inhibitor discovery and development.
Topics: Humans; Aniline Compounds; Benzamides; Deubiquitinating Enzymes; Neurodegenerative Diseases; Ubiquitin; Ubiquitin Thiolesterase; Ubiquitin-Specific Peptidase 7; Ubiquitination; Flow Cytometry; Protease Inhibitors; Coronavirus Papain-Like Proteases; Single-Domain Antibodies
PubMed: 37858708
DOI: 10.1016/j.jmb.2023.168316