RESUMO
Pancreatic cancer (PC) is a challenging and heterogeneous disease with a high mortality rate. Despite advancements in treatment, the prognosis for PC patients remains poor, with a high chance of disease recurrence. Biomarkers are crucial for diagnosing cancer, predicting patient prognosis and selecting treatments. However, the current lack of effective biomarkers for PC could contribute to the insufficiency of existing treatments. These findings underscore the urgent need to develop novel strategies to fight this disease. This study utilized multiple comprehensive bioinformatic analyses to identify potential therapeutic target genes in PC, focusing on histone lysine demethylases (KDMs). We found that high expression levels of KDM family genes, particularly KDM1A, KDM5A and KDM5B, were associated with improved overall survival in the cohort. Furthermore, the infiltration of various immune cells, including B cells, neutrophils, CD8+ T cells, dendritic cells, and macrophages, was positively correlated with KDM1A, KDM5A, and KDM5B expression. Moreover, MetaCore pathway analysis revealed interesting connections between KDM1A and the cell cycle and proliferation, between KDM5A and DNA damage and double-strand break repair through homologous recombination, and between KDM5B and WNT/ß-catenin signaling. These findings suggest that KDM1A, KDM5A and KDM5B may serve as promising biomarkers and therapeutic targets for PC, a disease of high importance due to its aggressive nature and urgent need for novel biomarkers to improve diagnosis and treatment.
Assuntos
Biomarcadores Tumorais , Regulação Neoplásica da Expressão Gênica , Histona Desmetilases com o Domínio Jumonji , Neoplasias Pancreáticas , Humanos , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patologia , Neoplasias Pancreáticas/metabolismo , Histona Desmetilases com o Domínio Jumonji/metabolismo , Histona Desmetilases com o Domínio Jumonji/genética , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Prognóstico , Biologia Computacional , Proteínas F-Box/metabolismo , Proteínas F-Box/genética , Histona Desmetilases/metabolismo , Histona Desmetilases/genética , Terapia de Alvo Molecular/métodos , Proteína 2 de Ligação ao Retinoblastoma/metabolismo , Proteína 2 de Ligação ao Retinoblastoma/genética , Via de Sinalização Wnt/genética , Proliferação de Células/genética , Proteínas Nucleares , Proteínas RepressorasRESUMO
BACKGROUND: Macrophage pyroptosis is a pivotal inflammatory mechanism in sepsis-induced lung injury, however, the underlying mechanisms remain inadequately elucidated. METHODS: Lipopolysaccharides (LPS)/adenosine triphosphate (ATP)-stimulated macrophages and cecal ligation and puncture (CLP)-induced mouse model for sepsis were established. The levels of key molecules were examined by qRT-PCR, Western blotting, immunohistochemistry (IHC) and ELISA assay. The subcellular localization of circMAPK1 was detected by RNA fluorescence in situ hybridization (FISH). Cell viability, LDH release and caspase-1 activity were monitored by CCK-8, LDH assays, and flow cytometry. The bindings between KDM2B/H3K36me2 and WNK1 promoter was detected by chromatin immunoprecipitation (ChIP) assay and luciferase assay, and associations among circMAPK1, UPF1 and KDM2B mRNA were assessed by RNA pull-down or RNA immunoprecipitation (RIP) assays. The pathological injury of lung tissues was evaluated by lung wet/dry weight ratio and hematoxylin and eosin (H&E) staining. RESULTS: CircMAPK1 was elevated in patients with septic lung injury. Knockdown of circMAPK1 protected against LPS/ATP-impaired cell viability and macrophage pyroptosis via WNK1/NLRP3 axis. Mechanistically, loss of circMAPK1 enhanced the association between KDM2B and WNK1 promoter to promote the demethylation of WNK1 and increase its expression. CircMAPK1 facilitated KDM2B mRNA decay by recruiting UPF1. Functional experiments showed that silencing of KDM2B or WNK1 counteracted circMAPK1 knockdown-suppressed macrophage pyroptosis. In addition, silencing of circMAPK1 alleviated CLP-induced lung injury in mice via KDM2B/WNK1/NLRP3 axis. CONCLUSION: CircMAPK1 exacerbates sepsis-induced lung injury by destabilizing KDM2B mRNA to suppress WNK1 expression, thus facilitating NLRP3-driven macrophage pyroptosis.
Assuntos
Epigênese Genética , Histona Desmetilases com o Domínio Jumonji , Piroptose , Sepse , Proteína Quinase 1 Deficiente de Lisina WNK , Animais , Piroptose/genética , Sepse/complicações , Sepse/genética , Sepse/metabolismo , Camundongos , Histona Desmetilases com o Domínio Jumonji/metabolismo , Histona Desmetilases com o Domínio Jumonji/genética , Masculino , Proteína Quinase 1 Deficiente de Lisina WNK/metabolismo , Proteína Quinase 1 Deficiente de Lisina WNK/genética , Humanos , Estabilidade de RNA , Lesão Pulmonar/etiologia , Lesão Pulmonar/metabolismo , Lesão Pulmonar/genética , Modelos Animais de Doenças , Feminino , Macrófagos/metabolismo , Camundongos Endogâmicos C57BL , Proteínas F-BoxRESUMO
Emerging evidence have demonstrated that F-box only protein 2 (FBXO2) is intimately associated with malignant tumor development and occurrence. However, neither the functions nor the molecular mechanisms underlying FBXO2 have been determined in the papillary thyroid carcinoma (PTC). The quantitative real-time PCR (qRT-PCR), western blotting and immunohistochemistry were carried out to detect the FBXO2 expression in PTC tissues. CCK-8 assay, EdU assay and flow cytometry were used to assess cell proliferation, cell cycle and apoptosis. The trans-well assay was conducted to determine the cell invasiveness. The effect of FBXO2 on PTC cell proliferation in vivo was observed through a subcutaneous tumor formation experiment in nude mice. Immunoprecipitation were conducted to detect the interaction between FBXO2 and p53. The ubiquitination assays were conducted to assess the regulation of p53 ubiquitination by FBXO2. FBXO2 was overexpressed in both PTC tissues and cell lines. FBXO2 expression positively correlated with PTC tumor size, lymphatic metastasis, and extramembranous invasion. Furthermore, silencing FBXO2 inhibited PTC cell proliferation and promoted apoptosis. The overexpression of FBXO2 significantly promotes PTC cell proliferation. Mechanistic studies revealed that FBXO2 could directly bind to p53 and promote its ubiquitination degradation. Knockdown of p53 partially reversed the progression arrest induced by FBXO2 Knockdown in PTC cells. FBXO2 knockdown inhibited PTC cell proliferation and promoted apoptosis by targeting p53 for ubiquitination and degradation. This process represents a research foundation for its diagnostic and therapeutic applications.
Assuntos
Apoptose , Proliferação de Células , Proteínas F-Box , Câncer Papilífero da Tireoide , Neoplasias da Glândula Tireoide , Proteína Supressora de Tumor p53 , Ubiquitinação , Adulto , Animais , Feminino , Humanos , Masculino , Camundongos , Pessoa de Meia-Idade , Apoptose/genética , Linhagem Celular Tumoral , Progressão da Doença , Proteínas F-Box/metabolismo , Proteínas F-Box/genética , Regulação Neoplásica da Expressão Gênica , Camundongos Nus , Transdução de Sinais , Câncer Papilífero da Tireoide/metabolismo , Câncer Papilífero da Tireoide/patologia , Câncer Papilífero da Tireoide/genética , Neoplasias da Glândula Tireoide/patologia , Neoplasias da Glândula Tireoide/metabolismo , Neoplasias da Glândula Tireoide/genética , Proteína Supressora de Tumor p53/metabolismo , Proteína Supressora de Tumor p53/genéticaRESUMO
The pathogen Agrobacterium tumefaciens is known for causing crown gall tumours in plants. However, it has also been harnessed as a valuable tool for plant genetic transformation. Apart from the T-DNA, Agrobacterium also delivers at least five virulence proteins into the host plant cells, which are required for an efficient infection. One of these virulence proteins is VirD5. F-box proteins, encoded in the host plant genome or the Ti plasmid, and the ubiquitin/26S proteasome system (UPS) also play an important role in facilitating Agrobacterium infection. Our study identified two Arabidopsis F-box proteins, D5BF1 and D5BF2, that bind VirD5 and facilitate its degradation via the UPS. Additionally, we found that Agrobacterium partially suppresses the expression of D5BF1 and D5BF2. Lastly, stable transformation and tumorigenesis efficiency assays revealed that D5BF1 and D5BF2 negatively regulate the Agrobacterium infection process, showing that the plant F-box proteins and UPS play a role in defending against Agrobacterium infection.
Assuntos
Agrobacterium tumefaciens , Proteínas de Arabidopsis , Arabidopsis , Proteínas F-Box , Transformação Genética , Arabidopsis/microbiologia , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Agrobacterium tumefaciens/genética , Agrobacterium tumefaciens/patogenicidade , Proteínas F-Box/metabolismo , Proteínas F-Box/genética , Carcinogênese/genética , Tumores de Planta/microbiologia , Complexo de Endopeptidases do Proteassoma/metabolismo , Regulação da Expressão Gênica de PlantasRESUMO
Intervertebral disc disease (IDD) is a debilitating spine condition that can be caused by intervertebral disc (IVD) damage which progresses towards IVD degeneration and dysfunction. Recently, human pluripotent stem cells (hPSCs) were recognized as a valuable resource for cell-based regenerative medicine in skeletal diseases. Therefore, adult somatic cells reprogrammed into human induced pluripotent stem cells (hiPSCs) represent an attractive cell source for the derivation of notochordal-like cells (NCs) as a first step towards the development of a regenerative therapy for IDD. Utilizing a differentiation method involving treatment with a four-factor cocktail targeting the BMP, FGF, retinoic acid, and Wnt signaling pathways, we differentiate CRISPR/Cas9-generated mCherry-reporter knock-in hiPSCs into notochordal-like cells. Comprehensive analysis of transcriptomic changes throughout the differentiation process identified regulation of histone methylation as a pivotal driver facilitating the differentiation of hiPSCs into notochordal-like cells. We further provide evidence that specific inhibition of histone demethylases KDM2A and KDM7A/B enhanced the lineage commitment of hiPSCs towards notochordal-like cells. Our results suggest that inhibition of KDMs could be leveraged to alter the epigenetic landscape of hiPSCs to control notochord-specific gene expression. Thus, our study highlights the importance of epigenetic regulators in stem cell-based regenerative approaches for the treatment of disc degeneration.
Assuntos
Diferenciação Celular , Células-Tronco Pluripotentes Induzidas , Histona Desmetilases com o Domínio Jumonji , Notocorda , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Células-Tronco Pluripotentes Induzidas/citologia , Notocorda/metabolismo , Notocorda/citologia , Histona Desmetilases com o Domínio Jumonji/metabolismo , Histona Desmetilases com o Domínio Jumonji/genética , Histona Desmetilases/metabolismo , Histona Desmetilases/genética , Proteínas F-BoxRESUMO
Recruitment of non-canonical BCOR-PRC1.1 to non-methylated CpG islands via KDM2B plays a fundamental role in transcription control during developmental processes and cancer progression. However, the mechanism is still largely unknown on how this recruitment is regulated. Here, we unveiled the importance of the Poly-D/E regions within the linker of BCOR for its binding to KDM2B. Interestingly, we also demonstrated that these negatively charged Poly-D/E regions on BCOR play autoinhibitory roles in liquid-liquid phase separation (LLPS) of BCORANK-linker-PUFD/PCGF1RAWUL. Through neutralizing negative charges of these Poly-D/E regions, Ca2+ not only weakens the interaction between BCOR/PCGF1 and KDM2B, but also promotes co-condensation of the enzymatic core of BCOR-PRC1.1 with KDM2B into liquid-like droplet. Accordingly, we propose that Ca2+ could modulate the compartmentation and recruitment of the enzymatic core of BCOR-PRC1.1 on KDM2B target loci. Thus, our finding advances the mechanistic understanding on how the tethering of BCOR-PRC1.1 enzymatic core to KDM2B is regulated.
Assuntos
Cálcio , Histona Desmetilases com o Domínio Jumonji , Complexo Repressor Polycomb 1 , Proteínas Proto-Oncogênicas , Proteínas Repressoras , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas/genética , Proteínas Repressoras/metabolismo , Proteínas Repressoras/genética , Cálcio/metabolismo , Humanos , Histona Desmetilases com o Domínio Jumonji/metabolismo , Histona Desmetilases com o Domínio Jumonji/genética , Histona Desmetilases com o Domínio Jumonji/química , Complexo Repressor Polycomb 1/metabolismo , Complexo Repressor Polycomb 1/genética , Ligação Proteica , Separação de Fases , Proteínas F-BoxRESUMO
The ubiquitin-proteasome system is frequently employed to degrade viral proteins, thereby inhibiting viral replication and pathogenicity. Through an analysis of the degradation kinetics of all the SARS-CoV-2 proteins, our study revealed rapid degradation of several proteins, particularly NSP5. Additionally, we identified FBXO22, an E3 ubiquitin ligase, as the primary regulator of NSP5 ubiquitination. Moreover, we validated the interaction between FBXO22 and NSP5, demonstrating that FBXO22-mediated ubiquitination of NSP5 facilitated its recognition by the proteasome, leading to subsequent degradation. Specifically, FBXO22 catalyzed the formation of K48-linked polyubiquitin chains on NSP5 at lysine residues 5 and 90. Knockdown of FBXO22 resulted in decreased NSP5 ubiquitination levels, increased stability, and enhanced ability to evade the host innate immune response. Notably, the protein level of FBXO22 were negatively correlated with SARS-CoV-2 load, highlighting its importance in inhibiting viral replication. This study elucidates the molecular mechanism by which FBXO22 mediates the degradation of NSP5 and underscores its critical role in limiting viral replication. The identification of FBXO22 as a regulator of NSP5 stability provides new insights and potential avenues for targeting NSP5 in antiviral strategies.
Assuntos
Complexo de Endopeptidases do Proteassoma , SARS-CoV-2 , Ubiquitinação , Replicação Viral , Humanos , Complexo de Endopeptidases do Proteassoma/metabolismo , SARS-CoV-2/fisiologia , SARS-CoV-2/metabolismo , COVID-19/virologia , COVID-19/metabolismo , Proteínas F-Box/metabolismo , Proteínas F-Box/genética , Células HEK293 , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitina-Proteína Ligases/genética , Proteólise , Proteases Semelhantes à Papaína de Coronavírus/metabolismo , Receptores Citoplasmáticos e NuclearesRESUMO
BACKGROUNDS: F-box protein 45 (FBXO45) has been implicated in the progression of several diseases. Whether FBXO45 is involved in the development of bladder cancer remains unclear. Thus, this study focused on the effect of FBXO45 on the malignant progression of bladder cancer cells. METHODS: FBXO45 small-interference fragment was transfected into RT4 and 5637 cells by liposome-mediated transfection, and the knockdown efficiency of FBXO45 was verified by Western blot assay. The growth rate between FBXO45 knockdown cell lines and control cell lines was compared by counting kit 8 and plate cloning experiments. The motility of bladder cancer cells was observed via the Transwell test and Wound healing test. The effects of FBXO45 silencing on apoptosis and cell division were confirmed by flow cytometry. Western blot assay was performed to determine the function of FBXO45 knockdown on key proteins of cell apoptosis and the ERK/Cyclin D1/CDK4 pathway. RESULTS: After FBXO45 knockdown, the proliferation of bladder cancer cells was blocked (p < 0.01), and the migration and invasion abilities were reduced (p < 0.01). FBXO45 knockdown reduced the number of S-phase cells (RT4, p < 0.01; 5637, p < 0.05) and enhanced the apoptotic rate (p < 0.01). FBXO45 knockdown decreased the levels of p-ERK1/2, CDK4 and Cyclin D1 (p < 0.01). CONCLUSIONS: This study revealed that FBXO45 plays a carcinogenic role in bladder cancer via the ERK/Cyclin D1/CDK4 pathway, which provides a reference for the clinical treatment of patients with bladder cancer.
Assuntos
Ciclina D1 , Quinase 4 Dependente de Ciclina , Progressão da Doença , Proteínas F-Box , Técnicas de Silenciamento de Genes , Neoplasias da Bexiga Urinária , Neoplasias da Bexiga Urinária/genética , Neoplasias da Bexiga Urinária/patologia , Neoplasias da Bexiga Urinária/metabolismo , Humanos , Quinase 4 Dependente de Ciclina/metabolismo , Quinase 4 Dependente de Ciclina/genética , Ciclina D1/metabolismo , Ciclina D1/genética , Proteínas F-Box/genética , Proteínas F-Box/metabolismo , Sistema de Sinalização das MAP Quinases , Células Tumorais Cultivadas , Linhagem Celular Tumoral , Proliferação de CélulasRESUMO
Centromere pairing is crucial for synapsis in meiosis. This study delves into the Skp1-Cullin1-F-box protein (SCF) E3 ubiquitin ligase complex, specifically focusing on F-box protein 47 (FBXO47), in mouse meiosis. Here, we revealed that FBXO47 is localized at the centromere and it regulates centromere pairing cooperatively with SKP1 to ensure proper synapsis in pachynema. The absence of FBXO47 causes defective centromeres, resulting in incomplete centromere pairing, which leads to corruption of SC at centromeric ends and along chromosome axes, triggering premature dissociation of chromosomes and pachytene arrest. FBXO47 deficient pachytene spermatocytes exhibited drastically reduced SKP1 expression at centromeres and chromosomes. Additionally, FBXO47 stabilizes SKP1 by down-regulating its ubiquitination in HEK293T cells. In essence, we propose that FBXO47 collaborates with SKP1 to facilitate centromeric SCF formation in spermatocytes. In summary, we posit that the centromeric SCF E3 ligase complex regulates centromere pairing for pachynema progression in mice.
Assuntos
Centrômero , Pareamento Cromossômico , Proteínas F-Box , Espermatócitos , Animais , Masculino , Centrômero/metabolismo , Centrômero/genética , Camundongos , Espermatócitos/metabolismo , Proteínas F-Box/metabolismo , Proteínas F-Box/genética , Humanos , Células HEK293 , Proteínas Ligases SKP Culina F-Box/metabolismo , Proteínas Ligases SKP Culina F-Box/genética , Meiose , Camundongos Knockout , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitinação , Camundongos Endogâmicos C57BLRESUMO
Background: Prostate cancer progression hinges on ß-catenin's stability and activity, a key factor in epithelial-mesenchymal transition (EMT) and metastasis. This study delves into NDR1-dependent phosphorylation's impact on ß-catenin via FBXO11, an E3 ubiquitin ligase, in prostate cancer cells. Methods: Human prostate cancer cell lines underwent various in vitro assays, including real-time PCR, Western blotting, immunoprecipitation, immunofluorescence, and protein stability assays, to explore ß-catenin's interactions and post-translational modifications. NDR1 modulation's in vivo efficacy was assessed using a nude mice lung metastasis model. Small-molecule screening identified a potential NDR1 activator, aNDR1, tested for its effects on metastasis via in vitro and in vivo assays. Results: NDR1 phosphorylated ß-catenin at Ser33/37, facilitating its interaction with FBXO11. This led to FBXO11-mediated ubiquitination and cytoplasmic degradation of ß-catenin, while the NDR1-FBXO11 complex impeded ß-catenin nuclear translocation by inducing JNK2 ubiquitination. Thus, NDR1 and FBXO11 jointly regulate ß-catenin activity in prostate cancer cells through dual phosphorylation-driven ubiquitination, potentially suppressing EMT. Reduced NDR1 expression inhibited FBXO11 and ß-catenin phosphorylation, diminishing ß-catenin and JNK2 ubiquitination, promoting EMT and enhancing prostate cancer cell metastasis. The inhibitory effects of aNDR1 on prostate cancer metastasis were validated. Conclusion: The NDR1/FBXO11 axis outlines a non-canonical ß-catenin degradation pathway crucial in regulating EMT and prostate cancer cell metastasis. NDR1 activation, particularly with aNDR1, could offer a promising therapeutic avenue against prostate cancer metastasis.
Assuntos
Camundongos Nus , Neoplasias da Próstata , Ubiquitinação , beta Catenina , Masculino , Humanos , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , beta Catenina/metabolismo , Fosforilação , Animais , Linhagem Celular Tumoral , Camundongos , Transição Epitelial-Mesenquimal , Proteínas F-Box/metabolismo , Metástase NeoplásicaRESUMO
The cytosolic peptide:N-glycanase (PNGase) is involved in the quality control of N-glycoproteins via the endoplasmic reticulum-associated degradation (ERAD) pathway. Mutations in the gene encoding cytosolic PNGase (NGLY1 in humans) cause NGLY1 deficiency. Recent findings indicate that the F-box protein FBS2 of the SCFFBS2 ubiquitin ligase complex can be a promising drug target for NGLY1 deficiency. Here, we determined the crystal structure of bovine FBS2 complexed with the adaptor protein SKP1 and a sugar ligand, Man3GlcNAc2, which corresponds to the core pentasaccharide of N-glycan. Our crystallographic data together with NMR data revealed the structural basis of disparate sugar-binding specificities in homologous FBS proteins and identified a potential druggable pocket for in silico docking studies. Our results provide a potential basis for the development of selective inhibitors against FBS2 in NGLY1 deficiency.
Assuntos
Peptídeo-N4-(N-acetil-beta-glucosaminil) Asparagina Amidase , Animais , Bovinos , Humanos , Sequência de Aminoácidos , Sítios de Ligação , Cristalografia por Raios X , Proteínas F-Box/metabolismo , Proteínas F-Box/química , Proteínas F-Box/genética , Modelos Moleculares , Simulação de Acoplamento Molecular , Peptídeo-N4-(N-acetil-beta-glucosaminil) Asparagina Amidase/metabolismo , Peptídeo-N4-(N-acetil-beta-glucosaminil) Asparagina Amidase/genética , Peptídeo-N4-(N-acetil-beta-glucosaminil) Asparagina Amidase/química , Ligação ProteicaRESUMO
Ribonucleoprotein (RNP) granules are membraneless electron-dense structures rich in RNAs and proteins, and involved in various cellular processes. Two RNP granules in male germ cells, intermitochondrial cement and the chromatoid body (CB), are associated with PIWI-interacting RNAs (piRNAs) and are required for transposon silencing and spermatogenesis. Other RNP granules in male germ cells, the reticulated body and CB remnants, are also essential for spermiogenesis. In this study, we disrupted FBXO24, a testis-enriched F-box protein, in mice and found numerous membraneless electron-dense granules accumulated in sperm flagella. Fbxo24 knockout (KO) mice exhibited malformed flagellar structures, impaired sperm motility, and male infertility, likely due to the accumulation of abnormal granules. The amount and localization of known RNP granule-related proteins were not disrupted in Fbxo24 KO mice, suggesting that the accumulated granules were distinct from known RNP granules. Further studies revealed that RNAs and two importins, IPO5 and KPNB1, abnormally accumulated in Fbxo24 KO spermatozoa and that FBXO24 could ubiquitinate IPO5. In addition, IPO5 and KPNB1 were recruited to stress granules, RNP complexes, when cells were treated with oxidative stress or a proteasome inhibitor. These results suggest that FBXO24 is involved in the degradation of IPO5, disruption of which may lead to the accumulation of abnormal RNP granules in sperm flagella.
Assuntos
Proteínas F-Box , Infertilidade Masculina , Camundongos Knockout , Cauda do Espermatozoide , Masculino , Animais , Infertilidade Masculina/genética , Infertilidade Masculina/metabolismo , Camundongos , Cauda do Espermatozoide/metabolismo , Proteínas F-Box/metabolismo , Proteínas F-Box/genética , Grânulos Citoplasmáticos/metabolismo , Espermatozoides/metabolismoRESUMO
The circadian clock is an endogenous oscillator, and its importance lies in its ability to impart rhythmicity on downstream biological processes, or outputs. Our knowledge of output regulation, however, is often limited to an understanding of transcriptional connections between the clock and outputs. For instance, the clock is linked to plant growth through the gating of photoreceptors via rhythmic transcription of the nodal growth regulators, PHYTOCHROME-INTERACTING FACTORs (PIFs), but the clock's role in PIF protein stability is less clear. Here, we identified a clock-regulated, F-box type E3 ubiquitin ligase, CLOCK-REGULATED F-BOX WITH A LONG HYPOCOTYL 1 (CFH1), that specifically interacts with and degrades PIF3 during the daytime. Additionally, genetic evidence indicates that CFH1 functions primarily in monochromatic red light, yet CFH1 confers PIF3 degradation independent of the prominent red-light photoreceptor phytochrome B (phyB). This work reveals a clock-mediated growth regulation mechanism in which circadian expression of CFH1 promotes sustained, daytime PIF3 degradation in parallel with phyB signaling.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Fatores de Transcrição Hélice-Alça-Hélice Básicos , Relógios Circadianos , Fitocromo B , Proteínas de Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Arabidopsis/metabolismo , Arabidopsis/genética , Relógios Circadianos/fisiologia , Relógios Circadianos/genética , Fitocromo B/metabolismo , Fitocromo B/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Regulação da Expressão Gênica de Plantas , Proteólise , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitina-Proteína Ligases/genética , Ritmo Circadiano/fisiologia , Proteínas F-Box/metabolismo , Proteínas F-Box/genética , LuzRESUMO
BACKGROUND: Gliomas are the most common primary malignant tumours of the central nervous system, and neddylation may be a potential target for the treatment of gliomas. Our study analysed neddylation's potential role in gliomas of different pathological types and its correlation with immunotherapy. METHODS: Genes required for model construction were sourced from existing literature, and their expression data were extracted from the TCGA and CGGA databases. LASSO regression was employed to identify genes associated with the prognosis of glioma patients in TCGA and to establish a clinical prognostic model. Biological changes in glioma cell lines following intervention with hub genes were evaluated using the CCK-8 assay and transwell assay. The genes implicated in the model construction were validated across various cell lines using Western blot. We conducted analyses to examine correlations between model scores and clinical data, tumor microenvironments, and immune checkpoints. Furthermore, we investigated potential differences in molecular functions and mechanisms among different groups. RESULTS: We identified 249 genes from the Reactome database and analysed their expression profiles in the TCGA and CGGA databases. After using LASSO-Cox, four genes (BRCA1, BIRC5, FBXL16 and KLHL25, p < 0.05) with significant correlations were identified. We selected FBXL16 for validation in in vitro experiments. Following FBXL16 overexpression, the proliferation, migration, and invasion abilities of glioma cell lines all showed a decrease. Then, we constructed the NEDD Index for gliomas. The nomogram indicated that this model could serve as an independent prognostic marker. Analysis of the tumour microenvironment and immune checkpoints revealed that the NEDD index was also correlated with immune cell infiltration and the expression levels of various immune checkpoints. CONCLUSION: The NEDD index can serve as a practical tool for predicting the prognosis of glioma patients, and it is correlated with immune cell infiltration and the expression levels of immune checkpoints.
Assuntos
Biomarcadores Tumorais , Neoplasias Encefálicas , Regulação Neoplásica da Expressão Gênica , Glioma , Humanos , Glioma/genética , Glioma/imunologia , Glioma/patologia , Prognóstico , Linhagem Celular Tumoral , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/imunologia , Neoplasias Encefálicas/patologia , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Proteína NEDD8/genética , Proteína NEDD8/metabolismo , Microambiente Tumoral/genética , Microambiente Tumoral/imunologia , Proteínas de Checkpoint Imunológico/genética , Proteínas de Checkpoint Imunológico/metabolismo , Proliferação de Células/genética , Proteínas F-Box/genética , Proteínas F-Box/metabolismo , Feminino , Perfilação da Expressão Gênica/métodos , Bases de Dados Genéticas , Movimento Celular/genética , MasculinoRESUMO
Proteasome is essential for cell survival, and proteasome inhibition induces proteasomal gene transcription via the activated endoplasmic-reticulum-associated transcription factor nuclear factor erythroid 2-like 1 (Nrf1/NFE2L1). Nrf1 activation requires proteolytic cleavage by DDI2 and N-glycan removal by NGLY1. We previously showed that Nrf1 ubiquitination by SKP1-CUL1-F-box (SCF)FBS2/FBXO6, an N-glycan-recognizing E3 ubiquitin ligase, impairs its activation, although the molecular mechanism remained elusive. Here, we show that SCFFBS2 cooperates with the RING-between-RING (RBR)-type E3 ligase ARIH1 to ubiquitinate Nrf1 through oxyester bonds in human cells. Endo-ß-N-acetylglucosaminidase (ENGASE) generates asparagine-linked N-acetyl glucosamine (N-GlcNAc) residues from N-glycans, and N-GlcNAc residues on Nrf1 served as acceptor sites for SCFFBS2-ARIH1-mediated ubiquitination. We reconstituted the polyubiquitination of N-GlcNAc and serine/threonine residues on glycopeptides and found that the RBR-specific E2 enzyme UBE2L3 is required for the assembly of atypical ubiquitin chains on Nrf1. The atypical ubiquitin chains inhibited DDI2-mediated activation. The present results identify an unconventional ubiquitination pathway that inhibits Nrf1 activation.
Assuntos
Fator 1 Nuclear Respiratório , Ubiquitinação , Humanos , Células HEK293 , Fator 1 Nuclear Respiratório/metabolismo , Fator 1 Nuclear Respiratório/genética , Fator 1 Relacionado a NF-E2/metabolismo , Fator 1 Relacionado a NF-E2/genética , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitina-Proteína Ligases/genética , Acetilglucosamina/metabolismo , Células HeLa , Complexo de Endopeptidases do Proteassoma/metabolismo , Proteínas F-Box/metabolismo , Proteínas F-Box/genéticaRESUMO
Influenza A virus (IAV) infection while primarily affecting the lungs, is often associated with cardiovascular complications. However, the mechanisms underlying this association are not fully understood. Here, we investigated the potential role of FBXL19, a member of the Skp1-Cullin-1-F-box family of E3 ubiquitin ligase, in IAV-induced cardiac inflammation. We demonstrated that FBXL19 overexpression in endothelial cells (ECs) reduced viral titers and IAV matrix protein 1 (M1) levels while increasing antiviral gene expression, including interferon (IFN)-α, -ß, and -γ and RANTES (regulated on activation normal T cell expressed and secreted) in the cardiac tissue of IAV-infected mice. Moreover, EC-specific overexpression of FBXL19 attenuated the IAV infection-reduced interferon regulatory factor 3 (IRF3) level without altering its mRNA level and suppressed cardiac inflammation. Furthermore, IAV infection triggered cellular senescence programs in the heart as indicated by the upregulation of p16 and p21 mRNA levels and the downregulation of lamin-B1 levels, which were partially reversed by FBXL19 overexpression in ECs. Our findings indicate that EC-specific overexpression of FBXL19 protects against IAV-induced cardiac damage by enhancing interferon-mediated antiviral signaling, reducing cardiac inflammation, and suppressing cellular senescence programs.NEW & NOTEWORTHY Our study reveals a novel facet of IAV infection, demonstrating that it can trigger cellular senescence within the heart. Intriguingly, upregulation of endothelial FBXL19 promotes host innate immunity, reduces cardiac senescence, and diminishes inflammation. These findings highlight the therapeutic potential of targeting FBXL19 to mitigate IAV-induced cardiovascular complications.
Assuntos
Senescência Celular , Células Endoteliais , Fator Regulador 3 de Interferon , Infecções por Orthomyxoviridae , Animais , Infecções por Orthomyxoviridae/imunologia , Infecções por Orthomyxoviridae/metabolismo , Células Endoteliais/metabolismo , Células Endoteliais/imunologia , Células Endoteliais/virologia , Fator Regulador 3 de Interferon/metabolismo , Fator Regulador 3 de Interferon/genética , Camundongos Endogâmicos C57BL , Camundongos , Proteínas F-Box/metabolismo , Proteínas F-Box/genética , Humanos , Vírus da Influenza A/patogenicidade , Miocárdio/metabolismo , Miocárdio/imunologia , Miocárdio/patologia , Modelos Animais de Doenças , Transdução de Sinais , Interferons/metabolismo , Interferons/genética , Masculino , Quimiocina CCL5RESUMO
Pancreatic ductal adenocarcinoma (PDAC) poses a significant threat due to its tendency to evade early detection, frequent metastasis, and the subsequent challenges in devising effective treatments. Processes that govern epithelial-mesenchymal transition (EMT) in PDAC hold promise for advancing novel therapeutic strategies. SAMD1 (SAM domain-containing protein 1) is a CpG island-binding protein that plays a pivotal role in the repression of its target genes. Here, we revealed that SAMD1 acts as a repressor of genes associated with EMT. Upon deletion of SAMD1 in PDAC cells, we observed significantly increased migration rates. SAMD1 exerts its effects by binding to specific genomic targets, including CDH2, encoding N-cadherin, which emerged as a driver of enhanced migration upon SAMD1 knockout. Furthermore, we discovered the FBXO11-containing E3 ubiquitin ligase complex as an interactor and negative regulator of SAMD1, which inhibits SAMD1 chromatin-binding genome-wide. High FBXO11 expression in PDAC is associated with poor prognosis and increased expression of EMT-related genes, underlining an antagonistic relationship between SAMD1 and FBXO11. In summary, our findings provide insights into the regulation of EMT-related genes in PDAC, shedding light on the intricate role of SAMD1 and its interplay with FBXO11 in this cancer type.
Assuntos
Carcinoma Ductal Pancreático , Transição Epitelial-Mesenquimal , Proteínas F-Box , Regulação Neoplásica da Expressão Gênica , Neoplasias Pancreáticas , Receptores de LDL , Animais , Humanos , Caderinas/metabolismo , Caderinas/genética , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/metabolismo , Carcinoma Ductal Pancreático/patologia , Linhagem Celular Tumoral , Movimento Celular/genética , Transição Epitelial-Mesenquimal/genética , Proteínas F-Box/metabolismo , Proteínas F-Box/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/genética , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patologia , Prognóstico , Receptores de LDL/genética , Receptores de LDL/metabolismoRESUMO
Accumulating evidence has demonstrated that F-box protein 22 (FBXO22) participates in tumour development and progression in various types of human malignancies. However, the functions and detailed molecular mechanisms of FBXO22 in osteosarcoma tumorigenesis and progression remain elusive. In this study, we aimed to determine the effects of FBXO22 on the cell proliferation, migration and invasion of osteosarcoma cells using cell counting kit-8 and Matrigel Transwell approaches. Moreover, we explored the molecular mechanisms by which FBXO22 mediated oncogenesis and progression in osteosarcoma via Western blotting, immunoprecipitation and ubiquitination. We found that FBXO22 depletion inhibited the proliferation, migration and invasion of osteosarcoma cells, whereas FBXO22 overexpression increased the proliferation and motility of osteosarcoma cells. Mechanistically, FBXO22 promoted the ubiquitination and degradation of FoxO1 in osteosarcoma cells. FBXO22 depletion reduced cell proliferation and motility via regulation of FoxO1. Taken together, our findings provide new insight into FBXO22-induced osteosarcoma tumorigenesis. The inhibition of FBXO22 could be a promising strategy for the treatment of osteosarcoma.
Assuntos
Movimento Celular , Proliferação de Células , Proteínas F-Box , Proteína Forkhead Box O1 , Regulação Neoplásica da Expressão Gênica , Osteossarcoma , Ubiquitinação , Osteossarcoma/metabolismo , Osteossarcoma/patologia , Osteossarcoma/genética , Humanos , Proteína Forkhead Box O1/metabolismo , Proteína Forkhead Box O1/genética , Proteínas F-Box/metabolismo , Proteínas F-Box/genética , Movimento Celular/genética , Linhagem Celular Tumoral , Proteólise , Progressão da Doença , Neoplasias Ósseas/metabolismo , Neoplasias Ósseas/patologia , Neoplasias Ósseas/genética , Invasividade Neoplásica , Carcinogênese/genética , Carcinogênese/metabolismo , Carcinogênese/patologia , Receptores Citoplasmáticos e NuclearesRESUMO
BACKGROUND: Non-small cell lung cancer (NSCLC) is a malignant form of lung cancer, and its prognosis could be improved by identifying key therapeutic targets. Thus, this study investigates the potential role of F-box Only Protein 33 (FBXO33) in NSCLC. METHODS: The expression levels of FBXO33 in NSCLC were determined using University of Alabama at Birmingham Cancer Data Analysis Portal (UALCAN) prediction, and its correlation with overall survival (OS) was analyzed via Kaplan-Meier survival analysis. These results were validated through quantitative polymerase chain reaction (qPCR), western blot (WB), and immunofluorescence (IF). We modulated FBXO33 expression by overexpression or knockdown and analyzed its effects on cell growth, proliferation, migration, invasion, and stemness characteristics in NSCLC cell lines. Additionally, the interaction between FBXO33 and Myelocytomatosis (Myc) and its impact on Myc ubiquitination were examined. An in vivo NSCLC xenograft model was used to corroborate the in vivo experimental results. RESULTS: The study found an inverse correlation between FBXO33 expression in NSCLC and OS. Lower FBXO33 expression enhanced the growth, proliferation, migration, invasion, and stemness characteristics of NSCLC cell lines. FBXO33 interacted with Myc to promote its ubiquitination and subsequent degradation, which suppressed NSCLC development. CONCLUSION: FBXO33 is expressed at low levels in NSCLC and correlates with lower OS. Overexpression of FBXO33 promotes Myc ubiquitination and degradation and inhibits tumor cell proliferation, migration and stemness characteristics, thereby impeding NSCLC progression.
Assuntos
Carcinoma Pulmonar de Células não Pequenas , Proliferação de Células , Proteínas F-Box , Neoplasias Pulmonares , Células-Tronco Neoplásicas , Proteínas Proto-Oncogênicas c-myc , Ubiquitinação , Humanos , Carcinoma Pulmonar de Células não Pequenas/patologia , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Carcinoma Pulmonar de Células não Pequenas/genética , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/metabolismo , Animais , Proteínas F-Box/metabolismo , Proteínas F-Box/genética , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Linhagem Celular Tumoral , Proteínas Proto-Oncogênicas c-myc/metabolismo , Proteínas Proto-Oncogênicas c-myc/genética , Proliferação de Células/genética , Movimento Celular/genética , Camundongos Nus , Camundongos , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitina-Proteína Ligases/genética , Regulação Neoplásica da Expressão Gênica , Feminino , Proteólise , MasculinoRESUMO
Neuroblastoma, the deadliest solid tumor in children, exhibits alarming mortality rates, particularly among high-risk cases. To enhance survival rates, a more precise risk stratification for patients is imperative. Utilizing proteomic data from 34 cases with or without N-Myc amplification, we identified 28 differentially expressed ubiquitination-related proteins (URGs). From these, a prognostic signature comprising 6 URGs was constructed. A nomogram incorporating clinical-pathological parameters yielded impressive AUC values of 0.88, 0.93, and 0.95 at 1, 3, and 5 years, respectively. Functional experiments targeting the E3 ubiquitin ligase FBXO42, a component of the prognostic signature, revealed its TP53-dependent promotion of neuroblastoma cell proliferation. In conclusion, our ubiquitination-related prognostic model robustly predicts patient outcomes, guiding clinical decisions. Additionally, the newfound pro-proliferative role of FBXO42 offers a novel foundation for understanding the molecular mechanisms of neuroblastoma.