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Epigenetics & Chromatin May 2024UHRF1 as a member of RING-finger type E3 ubiquitin ligases family, is an epigenetic regulator with five structural domains. It has been involved in the regulation of a... (Review)
Review
UHRF1 as a member of RING-finger type E3 ubiquitin ligases family, is an epigenetic regulator with five structural domains. It has been involved in the regulation of a series of biological functions, such as DNA replication, DNA methylation, and DNA damage repair. Additionally, aberrant overexpression of UHRF1 has been observed in over ten cancer types, indicating that UHRF1 is a typical oncogene. The overexpression of UHRF1 repressed the transcription of such tumor-suppressor genes as CDKN2A, BRCA1, and CDH1 through DNMT1-mediated DNA methylation. In addition to the upstream transcription factors regulating gene transcription, post-translational modifications (PTMs) also contribute to abnormal overexpression of UHRF1 in cancerous tissues. The types of PTM include phosphorylation, acetylation, methylationand ubiquitination, which regulate protein stability, histone methyltransferase activity, intracellular localization and the interaction with binding partners. Recently, several novel PTM types of UHRF1 have been reported, but the detailed mechanisms remain unclear. This comprehensive review summarized the types of UHRF1 PTMs, as well as their biological functions. A deep understanding of these crucial mechanisms of UHRF1 is pivotal for the development of novel UHRF1-targeted anti-cancer therapeutic strategies in the future.
Topics: Humans; Ubiquitin-Protein Ligases; Protein Processing, Post-Translational; Neoplasms; CCAAT-Enhancer-Binding Proteins; DNA Methylation; Animals; Ubiquitination; Gene Expression Regulation, Neoplastic
PubMed: 38725075
DOI: 10.1186/s13072-024-00540-y -
Cell Death & Disease May 2024RAD18, an important ubiquitin E3 ligase, plays a dual role in translesion DNA synthesis (TLS) and homologous recombination (HR) repair. However, whether and how the...
RAD18, an important ubiquitin E3 ligase, plays a dual role in translesion DNA synthesis (TLS) and homologous recombination (HR) repair. However, whether and how the regulatory mechanism of O-linked N-acetylglucosamine (O-GlcNAc) modification governing RAD18 and its function during these processes remains unknown. Here, we report that human RAD18, can undergo O-GlcNAcylation at Ser130/Ser164/Thr468, which is important for optimal RAD18 accumulation at DNA damage sites. Mechanistically, abrogation of RAD18 O-GlcNAcylation limits CDC7-dependent RAD18 Ser434 phosphorylation, which in turn significantly reduces damage-induced PCNA monoubiquitination, impairs Polη focus formation and enhances UV sensitivity. Moreover, the ubiquitin and RAD51C binding ability of RAD18 at DNA double-strand breaks (DSBs) is O-GlcNAcylation-dependent. O-GlcNAcylated RAD18 promotes the binding of RAD51 to damaged DNA during HR and decreases CPT hypersensitivity. Our findings demonstrate a novel role of RAD18 O-GlcNAcylation in TLS and HR regulation, establishing a new rationale to improve chemotherapeutic treatment.
Topics: Humans; DNA-Binding Proteins; Ubiquitin-Protein Ligases; Recombinational DNA Repair; Acetylglucosamine; Rad51 Recombinase; Proliferating Cell Nuclear Antigen; Phosphorylation; DNA Replication; Ubiquitination; DNA Breaks, Double-Stranded; DNA-Directed DNA Polymerase; Cell Cycle Proteins; DNA Damage; DNA; HEK293 Cells; Ultraviolet Rays; Protein Binding; Glycosylation; Translesion DNA Synthesis
PubMed: 38719812
DOI: 10.1038/s41419-024-06700-y -
The Journal of Biological Chemistry May 2024Base excision repair (BER) requires a tight coordination between the repair enzymes through protein-protein interactions and involves gap filling by DNA polymerase (pol)...
Base excision repair (BER) requires a tight coordination between the repair enzymes through protein-protein interactions and involves gap filling by DNA polymerase (pol) β and subsequent nick sealing by DNA ligase (LIG) 1 or LIGIIIα at the downstream steps. Apurinic/apyrimidinic-endonuclease 1 (APE1), by its exonuclease activity, proofreads 3' mismatches incorporated by polβ during BER. We previously reported that the interruptions in the functional interplay between polβ and the BER ligases result in faulty repair events. Yet, how the protein interactions of LIG1 and LIGIIIα could affect the repair pathway coordination during nick sealing at the final steps remains unknown. Here, we demonstrate that LIGIIIα interacts more tightly with polβ and APE1 than LIG1, and the N-terminal noncatalytic region of LIG1 as well as the catalytic core and BRCT domain of LIGIIIα mediate interactions with both proteins. Our results demonstrated less efficient nick sealing of polβ nucleotide insertion products in the absence of LIGIIIα zinc-finger domain and LIG1 N-terminal region. Furthermore, we showed a coordination between APE1 and LIG1/LIGIIIα during the removal of 3' mismatches from the nick repair intermediate on which both BER ligases can seal noncanonical ends or gap repair intermediate leading to products of single deletion mutagenesis. Overall results demonstrate the importance of functional coordination from gap filling by polβ coupled to nick sealing by LIG1/LIGIIIα in the presence of proofreading by APE1, which is mainly governed by protein-protein interactions and protein-DNA intermediate communications, to maintain repair efficiency at the downstream steps of the BER pathway.
PubMed: 38718860
DOI: 10.1016/j.jbc.2024.107355 -
Revista Da Associacao Medica Brasileira... 2024This prospective study aimed to provide a comprehensive analysis of the methylation status of two pivotal genes, CDKN2A/p16INK4A (cyclin-dependent kinase inhibitor 2A)...
OBJECTIVE
This prospective study aimed to provide a comprehensive analysis of the methylation status of two pivotal genes, CDKN2A/p16INK4A (cyclin-dependent kinase inhibitor 2A) and RB1 (retinoblastoma transcriptional corepressor 1), in breast cancer patients.
METHODS
Samples were obtained from 15 women diagnosed with breast cancer and who underwent a total mastectomy. DNA was extracted from the tumor, non-tumor tissue, and peripheral blood (circulating cell-free DNA). The methylation pattern of cell-free DNA extracted from blood collected on the day of mastectomy was compared with the methylation pattern of cell-free DNA from blood collected 1 year post-surgery. The methylation analysis was carried out by sodium bisulfite conversion and polymerase chain reaction, followed by electrophoresis.
RESULTS
Methylation of CDKN2A/p16INK4A was identified in 13 tumor samples and 12 non-tumor tissue samples. Two patients exhibited CDKN2A/p16INK4A methylation in the cell-free DNA of the first blood collection, while another showed methylation only in the cell-free DNA of the subsequent blood collection. Regarding RB1, 11 tumors and 8 non-tumor tissue samples presented methylation of the gene.
CONCLUSION
This study presents a novel approach for monitoring breast cancer patients through the analysis of cell-free DNA methylation. This analysis can detect changes in methylation patterns before any visible sign of cancer appears in breast tissue and could help predict the recurrence of malignant breast tumors.
Topics: Adult; Aged; Female; Humans; Middle Aged; Biomarkers, Tumor; Breast Neoplasms; Cell-Free Nucleic Acids; Cyclin-Dependent Kinase Inhibitor p16; DNA Methylation; Mastectomy; Polymerase Chain Reaction; Prospective Studies; Retinoblastoma Binding Proteins; Ubiquitin-Protein Ligases
PubMed: 38716944
DOI: 10.1590/1806-9282.20231358 -
International Immunopharmacology Jun 2024Asthma is a prevalent chronic respiratory disease, yet understanding its ecology and pathogenesis remains a challenge. Trim27, a ubiquitination ligase belonging to the...
Asthma is a prevalent chronic respiratory disease, yet understanding its ecology and pathogenesis remains a challenge. Trim27, a ubiquitination ligase belonging to the TRIM (tripartite motif-containing) family, has been implicated in regulating multiple pathophysiological processes such as inflammation, oxidative stress, apoptosis, and cell proliferation. However, the role of Trim27 in asthma has not been investigated. Our study found that Trim27 expression significantly increases in the airway epithelium of asthmatic mice. Knockdown of Trim27 expression effectively relieved ovalbumin (OVA)-induced airway hyperresponsiveness (AHR) and lung tissue histopathological changes. Moreover, Trim27 knockdown exhibited a significant reduction in airway inflammation and oxidative stress in asthmatic mice, and in vitro analysis confirmed the favorable effect of Trim27 deletion on inflammation and oxidative stress in mouse airway epithelial cells. Furthermore, our study revealed that deletion of Trim27 in MLE12 cells significantly decreased NLRP3 inflammasome activation, as evidenced by reduced expression of NLRP3, ASC, and pro-IL-1β mRNA. This downregulation was reversed when Trim27, but not its mutant lacking ubiquitination ligase activity, was replenished in these cells. Consistent with these findings, protein levels of NLRP3, pro-caspase-1, pro-IL-1β, cleaved-caspase-1, and cleaved-IL-1β were higher in Trim27-replenished cells compared to cells expressing Trim27C/A. Functionally, the downregulation of IL-1β and IL-18 levels induced by Trim27 deletion was rescued by replenishing Trim27. Overall, our findings provide evidence that Trim27 contributes to airway inflammation and oxidative stress in asthmatic mice via NLRP3 inflammasome activation, providing crucial insights into potential therapeutic interventions targeting Trim27 as a way to treat asthma.
Topics: Animals; Asthma; NLR Family, Pyrin Domain-Containing 3 Protein; Oxidative Stress; Inflammasomes; Mice; Mice, Inbred BALB C; Ovalbumin; Ubiquitin-Protein Ligases; Lung; Cell Line; Female; Disease Models, Animal; Inflammation; Humans; Mice, Inbred C57BL; Tripartite Motif Proteins; DNA-Binding Proteins
PubMed: 38713938
DOI: 10.1016/j.intimp.2024.112199 -
Biochimica Et Biophysica Acta.... Jun 2024Human DNA polymerase ι (Polι) belongs to the Y-family of specialized DNA polymerases engaged in the DNA damage tolerance pathway of translesion DNA synthesis that is...
Human DNA polymerase ι (Polι) belongs to the Y-family of specialized DNA polymerases engaged in the DNA damage tolerance pathway of translesion DNA synthesis that is crucial to the maintenance of genome integrity. The extreme infidelity of Polι and the fact that both its up- and down-regulation correlate with various cancers indicate that Polι expression and access to the replication fork should be strictly controlled. Here, we identify RNF2, an E3 ubiquitin ligase, as a new interacting partner of Polι that is responsible for Polι stabilization in vivo. Interestingly, while we report that RNF2 does not directly ubiquitinate Polι, inhibition of the E3 ubiquitin ligase activity of RNF2 affects the cellular level of Polι thereby protecting it from destabilization. Additionally, we indicate that this mechanism is more general, as DNA polymerase η, another Y-family polymerase and the closest paralogue of Polι, share similar features.
Topics: Humans; Ubiquitin-Protein Ligases; DNA-Directed DNA Polymerase; DNA Polymerase iota; Ubiquitination; DNA Replication; DNA Damage; HEK293 Cells; Enzyme Stability; Polycomb Repressive Complex 1
PubMed: 38705361
DOI: 10.1016/j.bbamcr.2024.119743 -
Journal of Experimental & Clinical... Apr 2024TP53, the most frequently mutated gene in human cancers, orchestrates a complex transcriptional program crucial for cancer prevention. While certain TP53-dependent genes...
BACKGROUND
TP53, the most frequently mutated gene in human cancers, orchestrates a complex transcriptional program crucial for cancer prevention. While certain TP53-dependent genes have been extensively studied, others, like the recently identified RNF144B, remained poorly understood. This E3 ubiquitin ligase has shown potent tumor suppressor activity in murine Eμ Myc-driven lymphoma, emphasizing its significance in the TP53 network. However, little is known about its targets and its role in cancer development, requiring further exploration. In this work, we investigate RNF144B's impact on tumor suppression beyond the hematopoietic compartment in human cancers.
METHODS
Employing TP53 wild-type cells, we generated models lacking RNF144B in both non-transformed and cancerous cells of human and mouse origin. By using proteomics, transcriptomics, and functional analysis, we assessed RNF144B's impact in cellular proliferation and transformation. Through in vitro and in vivo experiments, we explored proliferation, DNA repair, cell cycle control, mitotic progression, and treatment resistance. Findings were contrasted with clinical datasets and bioinformatics analysis.
RESULTS
Our research underscores RNF144B's pivotal role as a tumor suppressor, particularly in lung adenocarcinoma. In both human and mouse oncogene-expressing cells, RNF144B deficiency heightened cellular proliferation and transformation. Proteomic and transcriptomic analysis revealed RNF144B's novel function in mediating protein degradation associated with cell cycle progression, DNA damage response and genomic stability. RNF144B deficiency induced chromosomal instability, mitotic defects, and correlated with elevated aneuploidy and worse prognosis in human tumors. Furthermore, RNF144B-deficient lung adenocarcinoma cells exhibited resistance to cell cycle inhibitors that induce chromosomal instability.
CONCLUSIONS
Supported by clinical data, our study suggests that RNF144B plays a pivotal role in maintaining genomic stability during tumor suppression.
Topics: Animals; Humans; Mice; Cell Line, Tumor; Cell Proliferation; Genomic Instability; Lung Neoplasms; Tumor Suppressor Protein p53; Ubiquitin-Protein Ligases
PubMed: 38685100
DOI: 10.1186/s13046-024-03045-4 -
Mikrobiyoloji Bulteni Apr 2024The World Health Organization has included the problem of antibiotic resistance among the top 10 important health problems in the world. Treatment of infectious diseases...
The World Health Organization has included the problem of antibiotic resistance among the top 10 important health problems in the world. Treatment of infectious diseases has become more difficult due to the spread of antibiotic resistance between bacteria via transposable elements. Vancomycin-resistant enterococci (VRE) are of critical medical and public health importance due to their association with serious nosocomial infections and high risk of death. One of the most important features of VREs is that they have multiple antibiotic resistance and treatment options are reduced. Therefore, new treatment methods are needed. The vanA gene constitutes the building block of the vancomycin resistance mechanism and causes high resistance to vancomycin. In this study, it was aimed to investigate the neutralization of the vancomycin resistance mechanism by creating vanA antisense RNA (asRNA). The vanA positive VRE50 strain in our culture collection which was isolated from the clinical sample, was used to amplify the vanA gene by polymerase chain reaction (PCR). The amplified vanA amplicon was inserted inversely into the pUC19 plasmid by means of the enzyme cutting sites in the primers used. The resulting plasmid was combined with the pAT392 plasmid which can replicate in gram-positive bacteria and a fusion plasmid was created. The fusion plasmid whose orientation was confirmed, was transferred to the wild strain VRE50 by electroporation method. Minimum inhibitory concentration (MIC) values of transformed VRE (tVRE50) and wild type VRE50 strains used as control were determined by the E-Test method. The vancomycin MIC value of the wild type VRE50 strain was determined as 1024 µg/mL and that of the tVRE50 strain was 32 µg/mL and it was determined that the vancomycin resistance of the tVRE50 strain decreased with asRNA (antisense RNA). Antisense RNA technology is an important method for neutralizing the expression of genes. This study showed that neutralization of the vancomycin resistance gene may provide a lower MIC value in a vancomycin-resistant enterococcus strain and lead to increased susceptibility. This new approach provides a new method for VRE treatment by neutralizing the vancomycin resistance mechanism. The result obtained in this study needs to be supported by in vivo tests.
Topics: Vancomycin-Resistant Enterococci; Carbon-Oxygen Ligases; RNA, Antisense; Bacterial Proteins; Humans; Vancomycin; Plasmids; Vancomycin Resistance; Microbial Sensitivity Tests; Anti-Bacterial Agents; Gene Silencing
PubMed: 38676581
DOI: 10.5578/mb.202498191 -
International Journal of Molecular... Apr 2024Base excision repair (BER), which involves the sequential activity of DNA glycosylases, apurinic/apyrimidinic endonucleases, DNA polymerases, and DNA ligases, is one of...
Base excision repair (BER), which involves the sequential activity of DNA glycosylases, apurinic/apyrimidinic endonucleases, DNA polymerases, and DNA ligases, is one of the enzymatic systems that preserve the integrity of the genome. Normal BER is effective, but due to single-nucleotide polymorphisms (SNPs), the enzymes themselves-whose main function is to identify and eliminate damaged bases-can undergo amino acid changes. One of the enzymes in BER is DNA polymerase β (Polβ), whose function is to fill gaps in DNA. SNPs can significantly affect the catalytic activity of an enzyme by causing an amino acid substitution. In this work, pre-steady-state kinetic analyses and molecular dynamics simulations were used to examine the activity of naturally occurring variants of Polβ that have the substitutions L19P and G66R in the dRP-lyase domain. Despite the substantial distance between the dRP-lyase domain and the nucleotidyltransferase active site, it was found that the capacity to form a complex with DNA and with an incoming dNTP is significantly altered by these substitutions. Therefore, the lower activity of the tested polymorphic variants may be associated with a greater number of unrepaired DNA lesions.
Topics: DNA Polymerase beta; Humans; Amino Acid Substitution; Polymorphism, Single Nucleotide; Molecular Dynamics Simulation; DNA Repair; Kinetics; Catalytic Domain; DNA; Protein Domains
PubMed: 38673769
DOI: 10.3390/ijms25084182 -
Cells Apr 2024Cervical cancer is a major cause of death in women despite the advancement of current treatment modalities. The conventional therapeutic agent, cisplatin (CCDP), is the...
Cervical cancer is a major cause of death in women despite the advancement of current treatment modalities. The conventional therapeutic agent, cisplatin (CCDP), is the standard treatment for CC; however, resistance often develops due to the cancer's heterogeneity. Therefore, a detailed elucidation of the specific molecular mechanisms driving CC is crucial for the development of targeted therapeutic strategies. Retinoblastoma binding protein 6 () is a potential biomarker associated with cell proliferation and is upregulated in cervical cancer sites, exhibiting apoptosis and dysregulated expression. Furthermore, has been demonstrated to sensitize cancer cells to radiation and certain chemotherapeutic agents by regulating the gene, thus suggesting a crosstalk among / oncogenic signatures. The present study, therefore, investigated the relationship between cisplatin and expression in CC cells. Herein, we first explored bioinformatics simulations and identified that the signaling pathway is overexpressed and correlated with CC. For further analysis, we explored the Genomics of Drug Sensitivity in Cancer (GDSC) and found that most of the CC cell lines are sensitive to CCDP. To validate these findings, was silenced in HeLa and Vero cells using RNAi technology, followed by measurement of wild-type and at the mRNA level using qPCR. Cells co-treated with cisplatin and siRBBP6 were subsequently analyzed for apoptosis induction and real-time growth monitoring using flow cytometry and the xCELLigence system, respectively. Cancer cells in the co-treatment group showed a reduction in apoptosis compared to the cisplatin-treated group. Moreover, the real-time growth monitoring revealed a reduced growth rate in knockdown cells treated with cisplatin. Although wild-type remained unchanged in the co-treatment group of cancer cells, was completely repressed, suggesting that is necessary for sensitizing cervical cancer cells to cisplatin treatment by downregulating . The Vero cell population, which served as a non-cancerous control cell line in this study, remained viable following treatment with both siRBBP6 and cisplatin. Findings from this study suggest that expression promotes cisplatin sensitivity in HeLa cells through downregulation. Knockdown of limits apoptosis induction and delays cell growth inhibition in response to cisplatin. The knowledge obtained here has the potential to help improve cisplatin efficacy through personalized administration based on the expression profile of among individual patients.
Topics: Humans; Cisplatin; Female; Uterine Cervical Neoplasms; Cell Line, Tumor; Drug Resistance, Neoplasm; Tumor Suppressor Protein p53; Gene Expression Regulation, Neoplastic; Proto-Oncogene Proteins c-bcl-2; Apoptosis; Gene Knockdown Techniques; Cell Proliferation; Antineoplastic Agents; Signal Transduction; HeLa Cells; DNA-Binding Proteins; Ubiquitin-Protein Ligases
PubMed: 38667315
DOI: 10.3390/cells13080700