RESUMO
Cancer stem cells (CSCs) play a substantial role in cancer onset and recurrence. Anomalous iron and lipid metabolism have been documented in CSCs, suggesting that ferroptosis, a recently discovered form of regulated cell death characterised by lipid peroxidation, could potentially exert a significant influence on CSCs. However, the precise role of ferroptosis in gastric cancer stem cells (GCSCs) remains unknown. To address this gap, we screened ferroptosis-related genes in GCSCs using The Cancer Genome Atlas and corroborated our findings through quantitative polymerase chain reaction and western blotting. These results indicate that stearoyl-CoA desaturase (SCD1) is a key player in the regulation of ferroptosis in GCSCs. This study provides evidence that SCD1 positively regulates the transcription of squalene epoxidase (SQLE) by eliminating transcriptional inhibition of P53. This mechanism increases the cholesterol content and the elevated cholesterol regulated by SCD1 inhibits ferroptosis via the mTOR signalling pathway. Furthermore, our in vivo studies showed that SCD1 knockdown or regulation of cholesterol intake affects the stemness of GCSCs and their sensitivity to ferroptosis inducers. Thus, targeting the SCD1/squalene epoxidase/cholesterol signalling axis in conjunction with ferroptosis inducers may represent a promising therapeutic approach for the treatment of gastric cancer based on GCSCs.
Assuntos
Colesterol , Ferroptose , Células-Tronco Neoplásicas , Transdução de Sinais , Esqualeno Mono-Oxigenase , Estearoil-CoA Dessaturase , Neoplasias Gástricas , Serina-Treonina Quinases TOR , Ferroptose/genética , Neoplasias Gástricas/patologia , Neoplasias Gástricas/genética , Neoplasias Gástricas/metabolismo , Estearoil-CoA Dessaturase/metabolismo , Estearoil-CoA Dessaturase/genética , Serina-Treonina Quinases TOR/metabolismo , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Humanos , Esqualeno Mono-Oxigenase/metabolismo , Esqualeno Mono-Oxigenase/genética , Colesterol/metabolismo , Linhagem Celular Tumoral , Animais , Camundongos , Regulação Neoplásica da Expressão GênicaRESUMO
Azole antifungals inhibit the sterol C14-demethylase (CYP51/Erg11) of the ergosterol biosynthesis pathway. Here we show that the azole-induced synthesis of fungicidal cell wall carbohydrate patches in the pathogenic mold Aspergillus fumigatus strictly correlates with the accumulation of the CYP51 substrate eburicol. A lack of other essential ergosterol biosynthesis enzymes, such as sterol C24-methyltransferase (Erg6A), squalene synthase (Erg9) or squalene epoxidase (Erg1) does not trigger comparable cell wall alterations. Partial repression of Erg6A, which converts lanosterol into eburicol, increases azole resistance. The sterol C5-desaturase (ERG3)-dependent conversion of eburicol into 14-methylergosta-8,24(28)-dien-3ß,6α-diol, the "toxic diol" responsible for the fungistatic activity against yeasts, is not required for the fungicidal effects in A. fumigatus. While ERG3-lacking yeasts are azole resistant, ERG3-lacking A. fumigatus becomes more susceptible. Mutants lacking mitochondrial complex III functionality, which are much less effectively killed, but strongly inhibited in growth by azoles, convert eburicol more efficiently into the supposedly "toxic diol". We propose that the mode of action of azoles against A. fumigatus relies on accumulation of eburicol which exerts fungicidal effects by triggering cell wall carbohydrate patch formation.
Assuntos
Antifúngicos , Aspergillus fumigatus , Azóis , Proteínas Fúngicas , Aspergillus fumigatus/efeitos dos fármacos , Aspergillus fumigatus/metabolismo , Aspergillus fumigatus/genética , Antifúngicos/farmacologia , Proteínas Fúngicas/metabolismo , Proteínas Fúngicas/genética , Azóis/farmacologia , Ergosterol/metabolismo , Ergosterol/biossíntese , Parede Celular/metabolismo , Parede Celular/efeitos dos fármacos , Farmacorresistência Fúngica/genética , Monoterpenos Bicíclicos/farmacologia , Monoterpenos Bicíclicos/metabolismo , Testes de Sensibilidade Microbiana , Esterol 14-Desmetilase/metabolismo , Esterol 14-Desmetilase/genética , Sistema Enzimático do Citocromo P-450/metabolismo , Sistema Enzimático do Citocromo P-450/genética , Oxirredutases/metabolismo , Oxirredutases/genética , Metiltransferases/metabolismo , Metiltransferases/genética , Esqualeno Mono-Oxigenase/metabolismo , Esqualeno Mono-Oxigenase/genética , Lanosterol/análogos & derivadosRESUMO
Wolfiporia cocos, a versatile fungus acclaimed for its nutritional and therapeutic benefits in Traditional Chinese Medicine, holds immense potential for pharmaceutical and industrial applications. In this study, we aimed to optimize liquid fermentation techniques and culture medium composition to maximize mycelial biomass (MB) yield, pachymic acid (PA) concentration, and overall PA production. Additionally, we investigated the molecular basis of our findings by quantifying the expression levels of genes associated with PA and MB biosynthesis using quantitative real-time polymerase chain reaction. Under the optimized fermentation conditions, significant results were achieved, with maximum MB reaching 6.68 g l-1, PA content peaking at 1.25 mg g-1, and a total PA yield of 4.76 g l-1. Notably, among the four examined genes, squalene monooxygenase, exhibited enhanced expression at 0.06 ratio under the optimized conditions. Furthermore, within the realm of carbohydrate-active enzymes, the glycoside hydrolases 16 family displayed elevated expression levels at 21 ratios, particularly during MB production. This study enhances understanding of genetic mechanism governing MB and PA production in W. cocos, highlighting the roles of squalene monooxygenase and glycoside hydrolases 16 carbohydrate-active enzymes.
Assuntos
Biomassa , Meios de Cultura , Fermentação , Micélio , Triterpenos , Wolfiporia , Wolfiporia/genética , Wolfiporia/metabolismo , Micélio/crescimento & desenvolvimento , Micélio/metabolismo , Micélio/genética , Triterpenos/metabolismo , Meios de Cultura/química , Regulação Fúngica da Expressão Gênica , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Esqualeno Mono-Oxigenase/genética , Esqualeno Mono-Oxigenase/metabolismo , Expressão GênicaRESUMO
Dermatophytosis is a cutaneous infection that is able to degrade the keratinized tissues of the animal/human body, like skin, nails, and hair, causing chronic or subacute infection with the contact of some specific fungal strains. Trichophyton mentagrophytes are the most potential fungal pathogen causing dermatophytoses. The present study focuses on computationally based in silico antifungal activity of selected phytocompounds of Leucas aspera (Willd.) Link. against dermatophytic fungus, T. mentagrophytes. Validation and screening of derived phytocompounds is performed using Lipinski rule of five and toxicity test through Protox-II. Five target genes involved in dermatophytosis, induced by T. mentagrophytes are retrieved from the UniProt Database, and the corresponding proteins such as glucan 1,3-beta-glucosidase ARB_02797, Probable class II chitinase ARB_00204, squalene monooxygenase, actin, and ubiquitin are selected for in silico study. Three-dimensional structures of the target protein were computationally determined and validated through modeling tools and techniques due to the lack of validated protein structures in the database. Then, these proteins are used for in silico molecular docking through the AutoDock Vina tool to find out the promising phytocompounds. This study could be utilized in designing more effective drugs against T. mentagrophytes. Based on this work, a plant-based natural alternative can be added to the treatment of dermatophytosis rather than synthetic supplements.
Assuntos
Antifúngicos , Simulação de Acoplamento Molecular , Compostos Fitoquímicos , Compostos Fitoquímicos/farmacologia , Compostos Fitoquímicos/química , Antifúngicos/farmacologia , Antifúngicos/química , Proteínas Fúngicas/antagonistas & inibidores , Proteínas Fúngicas/metabolismo , Proteínas Fúngicas/química , Arthrodermataceae/efeitos dos fármacos , Tinha/microbiologia , Tinha/tratamento farmacológico , Esqualeno Mono-Oxigenase/antagonistas & inibidores , Esqualeno Mono-Oxigenase/metabolismo , Esqualeno Mono-Oxigenase/química , Humanos , Simulação por Computador , Quitinases/metabolismo , Quitinases/antagonistas & inibidores , Extratos Vegetais/farmacologia , Extratos Vegetais/química , Biologia Computacional , Actinas/metabolismoRESUMO
BACKGROUND: While de novo cholesterol biosynthesis plays a crucial role in chemotherapy resistance of colorectal cancer (CRC), the underlying molecular mechanism remains poorly understood. METHODS: We conducted cell proliferation assays on CRC cells with or without depletion of squalene epoxidase (SQLE), with or without 5-fluorouracil (5-FU) treatment. Additionally, a xenograft mouse model was utilized to explore the impact of SQLE on the chemosensitivity of CRC to 5-FU. RNA-sequencing analysis and immunoblotting analysis were performed to clarify the mechanism. We further explore the effect of SQLE depletion on the ubiquitin of NF-κB inhibitor alpha (IκBα) and (S)-2,3-epoxysqualene on the binding of IκBα to beta-transducin repeat containing E3 ubiquitin protein ligase (BTRC) by using immunoprecipitation assay. In addition, a cohort of 272 CRC patients were selected for our clinical analyses. RESULTS: Mechanistically, (S)-2,3-epoxysqualene promotes IκBα degradation and subsequent NF-κB activation by enhancing the interaction between BTRC and IκBα. Activated NF-κB upregulates the expression of baculoviral IAP repeat containing 3 (BIRC3), sustains tumor cell survival after 5-FU treatment and promotes 5-FU resistance of CRC in vivo. Notably, the treatment of terbinafine, an inhibitor of SQLE commonly used as antifungal drug in clinic, enhances the sensitivity of CRC to 5-FU in vivo. Additionally, the expression of SQLE is associated with the prognosis of human CRC patients with 5-FU-based chemotherapy. CONCLUSIONS: Thus, our finding not only demonstrates a new role of SQLE in chemoresistance of CRC, but also reveals a novel mechanism of (S)-2,3-epoxysqualene-dependent NF-κB activation, implicating the combined potential of terbinafine for 5-FU-based CRC treatment.
Assuntos
Neoplasias Colorretais , Resistencia a Medicamentos Antineoplásicos , Fluoruracila , NF-kappa B , Esqualeno Mono-Oxigenase , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Neoplasias Colorretais/metabolismo , Humanos , Esqualeno Mono-Oxigenase/metabolismo , Esqualeno Mono-Oxigenase/genética , NF-kappa B/metabolismo , Resistencia a Medicamentos Antineoplásicos/genética , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Animais , Fluoruracila/farmacologia , Fluoruracila/uso terapêutico , Camundongos , Linhagem Celular Tumoral , Camundongos Nus , Camundongos Endogâmicos BALB C , Feminino , Masculino , Proliferação de Células/efeitos dos fármacos , Inibidor de NF-kappaB alfa/metabolismo , Inibidor de NF-kappaB alfa/genética , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
The importance of trained immunity in antitumor immunity has been increasingly recognized, but the underlying metabolic regulation mechanisms remain incompletely understood. In this study, we find that squalene epoxidase (SQLE), a key enzyme in cholesterol synthesis, is required for ß-glucan-induced trained immunity in macrophages and ensuing antitumor activity. Unexpectedly, the shunt pathway, but not the classical cholesterol synthesis pathway, catalyzed by SQLE, is required for trained immunity induction. Specifically, 24(S),25-epoxycholesterol (24(S),25-EC), the shunt pathway metabolite, activates liver X receptor and increases chromatin accessibility to evoke innate immune memory. Meanwhile, SQLE-induced reactive oxygen species accumulation stabilizes hypoxia-inducible factor 1α protein for metabolic switching into glycolysis. Hence, our findings identify 24(S),25-EC as a key metabolite for trained immunity and provide important insights into how SQLE regulates trained-immunity-mediated antitumor activity.
Assuntos
Camundongos Endogâmicos C57BL , Esqualeno Mono-Oxigenase , Animais , Esqualeno Mono-Oxigenase/metabolismo , Camundongos , Colesterol/metabolismo , Colesterol/biossíntese , Colesterol/análogos & derivados , Receptores X do Fígado/metabolismo , Macrófagos/metabolismo , Macrófagos/imunologia , Macrófagos/efeitos dos fármacos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Imunidade Inata/efeitos dos fármacos , Humanos , Linhagem Celular TumoralRESUMO
As people age, the risk and progression of colorectal cancer (CRC), along with cholesterol levels, tend to increase. Nevertheless, epidemiological studies on serum lipids and CRC have produced conflicting results. We previously demonstrated that the reduction of squalene epoxidase (SQLE) due to accumulated cholesterol within cells accelerates CRC progression through the activation of the ß-catenin pathway. This study aimed to investigate the mechanism by which age-related cholesterol accumulation within tissue accelerates CRC progression and to assess the clinical significance of SQLE in older individuals with elevated CRC risk. Using machine learning-based digital image analysis with fluorescence-immunohistochemistry, we assessed SQLE, GSK3ßpS9 (GSK3ß activity inhibition through serine 9 phosphorylation at GSK3ß), p53 wild-type (p53WT), and p53 mutant (p53MT) levels in CRC tissues. Our analysis revealed a significant reduction in SQLE, p53WT, and p53MT and increase in GSK3ßpS9 levels, all associated with the substantial accumulation of intra-tissue cholesterol in aged CRCs. Cox analysis underscored the significant influence of SQLE on overall survival and progression-free survival in grade 2-3 CRC patients aged over 50. SQLE and GSK3ßpS9 consistently exhibited outstanding prognostic and diagnostic performance, particularly in older individuals. Furthermore, combining SQLE with p53WT, p53MT, and GSK3ßpS9 demonstrated a robust diagnostic ability in the older population. In conclusion, we have identified that individuals aged over 50 face an increased risk of CRC progression due to aging-linked cholesterol accumulation within tissue and the subsequent reduction in SQLE levels. This study also provides valuable biomarkers, including SQLE and GSK3ßpS9, for older patients at elevated risk of CRC.
Assuntos
Colesterol , Neoplasias Colorretais , Progressão da Doença , Esqualeno Mono-Oxigenase , Idoso , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Envelhecimento/metabolismo , Colesterol/metabolismo , Neoplasias Colorretais/patologia , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/genética , Fatores de Risco , Esqualeno Mono-Oxigenase/metabolismo , Esqualeno Mono-Oxigenase/genéticaRESUMO
Cholesterol is essential for both normal cell viability and cancer cell proliferation. Aberrant activity of squalene monooxygenase (SM, also known as squalene epoxidase), the rate-limiting enzyme of the committed cholesterol synthesis pathway, is accordingly implicated in a growing list of cancers. We previously reported that hypoxia triggers the truncation of SM to a constitutively active form, thus preserving sterol synthesis during oxygen shortfalls. Here, we show SM truncation is upregulated and correlates with the magnitude of hypoxia in endometrial cancer tissues, supporting the in vivo relevance of our earlier work. To further investigate the pathophysiological consequences of SM truncation, we examined its lipid droplet-localized pool using complementary immunofluorescence and cell fractionation approaches and found that it exclusively comprises the truncated enzyme. This partitioning is facilitated by the loss of an endoplasmic reticulum-embedded region at the SM N terminus, whereas the catalytic domain containing membrane-associated C-terminal helices is spared. Moreover, we determined multiple amphipathic helices contribute to the lipid droplet localization of truncated SM. Taken together, our results expand on the striking differences between the two forms of SM and suggest upregulated truncation may contribute to SM-related oncogenesis.
Assuntos
Colesterol , Neoplasias do Endométrio , Gotículas Lipídicas , Esqualeno Mono-Oxigenase , Feminino , Humanos , Linhagem Celular Tumoral , Colesterol/metabolismo , Colesterol/biossíntese , Neoplasias do Endométrio/metabolismo , Neoplasias do Endométrio/patologia , Neoplasias do Endométrio/genética , Retículo Endoplasmático/metabolismo , Regulação Neoplásica da Expressão Gênica , Gotículas Lipídicas/metabolismo , Esqualeno Mono-Oxigenase/metabolismo , Esqualeno Mono-Oxigenase/genética , Regulação para CimaRESUMO
BACKGROUND: Osteosarcoma (OSA) presents a clinical challenge and has a low 5-year survival rate. Currently, the lack of advanced stratification models makes personalized therapy difficult. This study aims to identify novel biomarkers to stratify high-risk OSA patients and guide treatment. METHODS: We combined 10 machine-learning algorithms into 101 combinations, from which the optimal model was established for predicting overall survival based on transcriptomic profiles for 254 samples. Alterations in transcriptomic, genomic and epigenomic landscapes were assessed to elucidate mechanisms driving poor prognosis. Single-cell RNA sequencing (scRNA-seq) unveiled genes overexpressed in OSA cells as potential therapeutic targets, one of which was validated via tissue staining, knockdown and pharmacological inhibition. We characterized changes in multiple phenotypes, including proliferation, colony formation, migration, invasion, apoptosis, chemosensitivity and in vivo tumourigenicity. RNA-seq and Western blotting elucidated the impact of squalene epoxidase (SQLE) suppression on signalling pathways. RESULTS: The artificial intelligence-derived prognostic index (AIDPI), generated by our model, was an independent prognostic biomarker, outperforming clinicopathological factors and previously published signatures. Incorporating the AIDPI with clinical factors into a nomogram improved predictive accuracy. For user convenience, both the model and nomogram are accessible online. Patients in the high-AIDPI group exhibited chemoresistance, coupled with overexpression of MYC and SQLE, increased mTORC1 signalling, disrupted PI3K-Akt signalling, and diminished immune infiltration. ScRNA-seq revealed high expression of MYC and SQLE in OSA cells. Elevated SQLE expression correlated with chemoresistance and worse outcomes in OSA patients. Therapeutically, silencing SQLE suppressed OSA malignancy and enhanced chemosensitivity, mediated by cholesterol depletion and suppression of the FAK/PI3K/Akt/mTOR pathway. Furthermore, the SQLE-specific inhibitor FR194738 demonstrated anti-OSA effects in vivo and exhibited synergistic effects with chemotherapeutic agents. CONCLUSIONS: AIDPI is a robust biomarker for identifying the high-risk subset of OSA patients. The SQLE protein emerges as a metabolic vulnerability in these patients, providing a target with translational potential.
Assuntos
Neoplasias Ósseas , Osteossarcoma , Esqualeno Mono-Oxigenase , Humanos , Inteligência Artificial , Biomarcadores , Osteossarcoma/tratamento farmacológico , Osteossarcoma/genética , Fosfatidilinositol 3-Quinases , Prognóstico , Proteínas Proto-Oncogênicas c-akt , Esqualeno Mono-Oxigenase/genética , Esqualeno Mono-Oxigenase/metabolismoRESUMO
Squalene epoxidase (SQLE) is an essential enzyme involved in cholesterol biosynthesis. However, its role in sarcoma and its correlation with immune infiltration remains unclear. All original data were downloaded from The Cancer Genome Atlas (TCGA). SQLE expression was explored using the TCGA database, and correlations between SQLE and cancer immune characteristics were analyzed via the TISIDB databases. Generally, SQLE is predominantly overexpressed and has diagnostic and prognostic value in sarcoma. Upregulated SQLE was associated with poorer overall survival, poorer disease-specific survival, and tumor multifocality in sarcoma. Mechanistically, we identified a hub gene that included a total of 82 SQLE-related genes, which were tightly associated with histone modification pathways in sarcoma patients. SQLE expression was negatively correlated with infiltrating levels of dendritic cells and plasmacytoid dendritic cells and positively correlated with Th2 cells. SQLE expression was negatively correlated with the expression of chemokines (CCL19 and CX3CL1) and chemokine receptors (CCR2 and CCR7) in sarcoma. In conclusion, SQLE may be used as a prognostic biomarker for determining prognosis and immune infiltration in sarcoma.
Assuntos
Sarcoma , Esqualeno Mono-Oxigenase , Humanos , Prognóstico , Esqualeno Mono-Oxigenase/genética , Esqualeno Mono-Oxigenase/metabolismo , Biomarcadores Tumorais/genética , Sarcoma/genéticaRESUMO
BACKGROUND: Squalene epoxidase is one of the rate-limiting enzymes in the biosynthetic pathway of membrane sterols and triterpenoids. The enzyme catalyzes the formation of oxidized squalene, which is a common precursor of sterols and triterpenoids. RESULT: In this study, the squalene epoxidase gene (PcSE) was evaluated in Poria cocos. Molecular docking between PcSE and squalene was performed and the active amino acids were identified. The sgRNA were designed based on the active site residues. The effect on triterpene synthesis in P. cocos was consistent with the results from ultra-high-performance liquid chromatography-quadruplex time-of-flight-double mass spectrometry (UHPLC-QTOF-MS/MS) analysis. The results showed that deletion of PcSE inhibited triterpene synthesis. In vivo verification of PcSE function was performed using a PEG-mediated protoplast transformation approach. CONCLUSION: The findings from this study provide a foundation for further studies on heterologous biosynthesis of P. cocos secondary metabolites.
Assuntos
Fitosteróis , Triterpenos , Wolfiporia , Espectrometria de Massas em Tandem/métodos , Esqualeno Mono-Oxigenase/genética , Esqualeno Mono-Oxigenase/metabolismo , Wolfiporia/genética , Wolfiporia/metabolismo , Simulação de Acoplamento Molecular , Esqualeno , Sistemas CRISPR-Cas , Edição de Genes , RNA Guia de Sistemas CRISPR-Cas , Triterpenos/metabolismoRESUMO
Cholesterol biosynthesis and metabolism are critical aspects that shape the process of tumor development and associated microenvironmental conditions owing to the ability of cholesterol to drive tumor growth and invasion. Squalene Epoxidase (SQLE) is the second rate-limiting enzyme involved in the synthesis of cholesterol. The functional role of SQLE within the tumor microenvironment, however, has yet to be established. Here we show that SQLE is distinctively expressed across most types of cancer, and the expression level is highly correlated with tumor mutation burden and microsatellite instability. Accordingly, SQLE was identified as a prognostic risk factor in cancer patients. In addition, we observed a negative correlation between SQLE expression and immune cell infiltration across multiple cancers, and murine xenograft model further confirmed that SQLE knockdown was associated with enhanced intratumoral CD8+ T cell infiltration. Using next-generation sequencing, we identified 410 genes distinctively expressed in tumors exhibiting SQLE inhibition. KEGG and GO analysis further verified that SQLE altered the immune response in the tumor microenvironment. Furthermore, we found that the metabolism and translation of proteins is the main binding factor with SQLE. Our findings ascertain that SQLE is a potential target in multiple cancers and suppressing SQLE establishes an essential mechanism for shaping tumor microenvironment.
Assuntos
Linfócitos T CD8-Positivos , Esqualeno Mono-Oxigenase , Microambiente Tumoral , Animais , Humanos , Camundongos , Linfócitos T CD8-Positivos/metabolismo , Colesterol , Neoplasias/genética , Neoplasias/metabolismo , Esqualeno Mono-Oxigenase/genética , Esqualeno Mono-Oxigenase/metabolismoRESUMO
The transcription factors p53 and MYC are often considered non-druggable targets, but their dysregulation can generate new dependencies and treatment opportunities in cancer cells. The p53 and MYC-regulated squalene epoxidase (SQLE) has been identified as a potential Achilles heel in colorectal cancer. This is of great interest because the FDA-approved anti-fungal SQLE inhibitor Terbinafine could be repurposed to treat colorectal cancer patients.
Assuntos
Neoplasias Colorretais , Esqualeno Mono-Oxigenase , Humanos , Esqualeno Mono-Oxigenase/genética , Esqualeno Mono-Oxigenase/metabolismo , Proteína Supressora de Tumor p53/genética , Terbinafina , Mutação , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/genéticaRESUMO
Pancreatic cancer (PC), a highly lethal malignancy, commonly exhibits metabolic reprogramming that results in therapeutic vulnerabilities. Nevertheless, the mechanisms underlying the impacts of aberrant cholesterol metabolism on PC development and progression remain elusive. In this study, we found that squalene epoxidase (SQLE) is a crucial mediator of cholesterol metabolism in PC growth. We observed a profound upregulation of SQLE in PC tissues, and its high expression was correlated with poor patient outcomes. Our functional experiments demonstrated that SQLE facilitated cell proliferation, induced cell cycle progression, and inhibited apoptosis in vitro, while promoting tumor growth in vivo. Mechanistically, SQLE was found to have a dual role. First, its inhibition led to squalene accumulation-induced endoplasmic reticulum (ER) stress and subsequent apoptosis. Second, it enhanced de novo cholesterol biosynthesis and maintained lipid raft stability, thereby activating the Src/PI3K/Akt signaling pathway. Significantly, employing SQLE inhibitors effectively suppressed PC cell proliferation and xenograft tumor growth. In summary, this study reveals SQLE as a novel oncogene that promotes PC growth by mitigating ER stress and activating lipid raft-regulated Src/PI3K/Akt signaling pathway, highlighting the potential of SQLE as a therapeutic target for PC.
Assuntos
Neoplasias Pancreáticas , Proteínas Proto-Oncogênicas c-akt , Humanos , Linhagem Celular Tumoral , Proliferação de Células , Colesterol , Neoplasias Pancreáticas/genética , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais , Esqualeno Mono-Oxigenase/metabolismo , Quinases da Família srcRESUMO
The acetylation-dependent (Ac/)N-degron pathway degrades proteins through recognition of their acetylated N-termini (Nt) by E3 ligases called Ac/N-recognins. To date, specific Ac/N-recognins have not been defined in plants. Here we used molecular, genetic, and multiomics approaches to characterize potential roles for Arabidopsis (Arabidopsis thaliana) DEGRADATION OF ALPHA2 10 (DOA10)-like E3 ligases in the Nt-acetylation-(NTA)-dependent turnover of proteins at global- and protein-specific scales. Arabidopsis has two endoplasmic reticulum (ER)-localized DOA10-like proteins. AtDOA10A, but not the Brassicaceae-specific AtDOA10B, can compensate for loss of yeast (Saccharomyces cerevisiae) ScDOA10 function. Transcriptome and Nt-acetylome profiling of an Atdoa10a/b RNAi mutant revealed no obvious differences in the global NTA profile compared to wild type, suggesting that AtDOA10s do not regulate the bulk turnover of NTA substrates. Using protein steady-state and cycloheximide-chase degradation assays in yeast and Arabidopsis, we showed that turnover of ER-localized SQUALENE EPOXIDASE 1 (AtSQE1), a critical sterol biosynthesis enzyme, is mediated by AtDOA10s. Degradation of AtSQE1 in planta did not depend on NTA, but Nt-acetyltransferases indirectly impacted its turnover in yeast, indicating kingdom-specific differences in NTA and cellular proteostasis. Our work suggests that, in contrast to yeast and mammals, targeting of Nt-acetylated proteins is not a major function of DOA10-like E3 ligases in Arabidopsis and provides further insight into plant ERAD and the conservation of regulatory mechanisms controlling sterol biosynthesis in eukaryotes.
Assuntos
Arabidopsis , Proteínas de Saccharomyces cerevisiae , Animais , Acetilação , Arabidopsis/genética , Arabidopsis/metabolismo , Mamíferos/metabolismo , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Esqualeno Mono-Oxigenase/metabolismo , Esteróis , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismoRESUMO
Endometrial cancer (EC) is a common malignant tumor that lacks any therapeutic target and, in many cases, recurrence is the leading ca use of morbidity and mortality in women. Widely known EC has a strongly positive correlation with abnormal lipid metabolism. Squalene epoxidase (SQLE), a crucial enzyme in the cholesterol synthesis pathway regulating lipid metabolic processes has been found to be associated with various cancers in recent years. Here, we focused on studying the role of SQLE in EC. Our study revealed that SQLE expression level was upregulated significantly in EC tissues. In vitro experiments showed that SQLE overexpression significantly promoted the proliferation, and inhibited cell apoptosis of EC cells, whereas SQLE knockdown or use of terbinafine showed the opposite results. Furthermore, we found out that the promotional effect of SQLE on the proliferation of EC cells might be achieved by activating the PI3K/AKT pathway. In vivo, studies confirmed that the knockdown of SQLE or terbinafine can observably inhibit tumor growth in nude mice. These results indicate that SQLE may promote the progression of EC by activating the PI3K/AKT pathway. Moreover, SQLE is a potential target for EC treatment and its inhibitor, terbinafine, has the potential to become a targeted drug for EC treatment.
Assuntos
Neoplasias do Endométrio , Proteínas Proto-Oncogênicas c-akt , Humanos , Animais , Camundongos , Feminino , Proteínas Proto-Oncogênicas c-akt/metabolismo , Terbinafina/farmacologia , Esqualeno Mono-Oxigenase/genética , Esqualeno Mono-Oxigenase/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Camundongos Nus , Transdução de Sinais , Neoplasias do Endométrio/tratamento farmacológico , Neoplasias do Endométrio/genética , Proliferação de Células , Linhagem Celular TumoralRESUMO
Cisplatin resistance poses a substantial hurdle in effectively treating head and neck squamous cell carcinoma (HNSCC). Utilizing multiple tumor models and examining an internal HNSCC cohort, squalene epoxidase (SQLE) is pinpointed as a key driver of chemoresistance and tumorigenesis, operating through a cholesterol-dependent pathway. Comprehensive transcriptomic analysis reveals that SQLE is essential for maintaining c-Myc transcriptional activity by stabilizing the c-Myc protein and averting its ubiquitin-mediated degradation. Mechanistic investigation demonstrates that SQLE inhibition diminishes Akt's binding affinity to lipid rafts via a cholesterol-dependent process, subsequently deactivating lipid raft-localized Akt, reducing GSK-3ß phosphorylation at S9, and increasing c-Myc phosphorylation at T58, ultimately leading to c-Myc destabilization. Importantly, employing an Sqle conditional knockout mouse model, SQLE's critical role in HNSCC initiation and progression is established. The preclinical findings demonstrate the potent synergistic effects of combining terbinafine and cisplatin in arresting tumor growth. These discoveries not only provide novel insights into the underlying mechanisms of SQLE-mediated cisplatin resistance and tumorigenesis in HNSCC but also propose a promising therapeutic avenue for HNSCC patients unresponsive to conventional cisplatin-based chemotherapy.
Assuntos
Neoplasias de Cabeça e Pescoço , Esqualeno Mono-Oxigenase , Animais , Camundongos , Humanos , Carcinoma de Células Escamosas de Cabeça e Pescoço/tratamento farmacológico , Esqualeno Mono-Oxigenase/genética , Esqualeno Mono-Oxigenase/metabolismo , Cisplatino/farmacologia , Cisplatino/uso terapêutico , Proteínas Proto-Oncogênicas c-akt , Resistencia a Medicamentos Antineoplásicos , Glicogênio Sintase Quinase 3 beta , Transformação Celular Neoplásica , Carcinogênese , Colesterol , Neoplasias de Cabeça e Pescoço/tratamento farmacológicoRESUMO
KEYMESSAGE: CbSE overexpression increased stigmasterol levels and altered plant morphology. The genes upstream and downstream of CbSE were found to be upregulated, which confirms its regulatory role in the saponin biosynthetic pathway. Chlorophytum borivilianum is a high-value medicinal plant with many promising preclinical applications that include saponins as a major active ingredient. Squalene epoxidase (SE) is one of the major rate-limiting enzymes of the saponin biosynthetic pathway. Here, we functionally characterized C. borivilianum SE (CbSE) by over-expressing heterologously in Nicotiana tabacum. The heterologous expression of CbSE resulted in stunted pant growth with altered leaf and flower morphology. Next, RT-qPCR analysis of transgenic plants overexpressing CbSE revealed increased expression levels of Cycloartenol synthase (CAS), Beta amyrin synthase (ßAS), and cytochrome P450 monooxygenase 51 (CYP51) (Cytochrome P450), which encode key enzymes for triterpenoid and phytosterol biosynthesis in C. borivilianum. Further, Methyl Jasmonate (MeJa) treatment upregulated Squalene synthase (SQS), SE, and Oxidosqualene cyclases (OSCs) to a significant level. GC-MS analysis of the leaf and hairy roots of the transformants showed an increased stigmasterol content (0.5-1.0 fold) compared to wild type (WT) plants. These results indicate that CbSE is a rate-limiting gene, which encodes an efficient enzyme responsible for phytosterol and triterpenoid production in C. borivilianum.
Assuntos
Fitosteróis , Saponinas , Triterpenos , Nicotiana/genética , Nicotiana/metabolismo , Estigmasterol , Esqualeno Mono-Oxigenase/genética , Esqualeno Mono-Oxigenase/metabolismo , Triterpenos/metabolismo , Sistema Enzimático do Citocromo P-450/genética , Sistema Enzimático do Citocromo P-450/metabolismo , Regulação da Expressão Gênica de PlantasRESUMO
Understanding the genetic mechanisms underlying milk production traits contribute to improving the production potential of dairy animals. Squalene epoxidase (SQLE) is one of the rate-limiting enzymes for cholesterol biosynthesis and was highly expressed in the buffalo mammary. The objectives of the present study were to detect the polymorphisms within SQLE in buffalo, the genetic effects of these mutations on milk production traits, and to understand the gene regulatory effects on buffalo mammary epithelial cells (BuMECs). A total of five SNPs were identified by sequencing, g.18858G > A loci were significantly associated with fat yield, and g.22834C > T loci were significantly associated with peak milk yield, milk yield, fat yield, and protein yield. Notably, linkage disequilibrium analysis indicated that 2 SNPs (g.18858G > A and g.22834C > T) formed one haplotype block, which was found to be significantly associated with milk fat yield, fat percentage, and protein yield. Furthermore, expression of SQLE was measured in different tissues of buffalo and was found to be higher in the mammary. Knockdown of SQLE gene expression significantly affected the growth of BuMECs, including proliferation, cell cycle, and apoptosis, and significantly downregulated the expression of related genes MYC, PCNA, and P21. In addition, knockdown of the SQLE gene significantly reduces triglyceride concentrations and the signal intensity of oil red O staining. In addition, silencing of SQLE was also found to regulate the synthesis and secretion of ß-casein and κ-casein negatively. Furthermore, SQLE knockdown is accompanied by the downregulation of critical genes (RPS6KB1, JAK2, eIF4E, and SREBP1) related to milk fat and protein synthesis. The current study showed the potential of the SQLE gene as a candidate for buffalo milk production traits. It provides a new understanding of the physiological mechanisms underlying buffalo milk production regulation.
Assuntos
Leite , Esqualeno Mono-Oxigenase , Animais , Leite/metabolismo , Esqualeno Mono-Oxigenase/genética , Esqualeno Mono-Oxigenase/metabolismo , Fenótipo , Haplótipos , Polimorfismo de Nucleotídeo Único , Búfalos/genéticaRESUMO
BACKGROUND: The mortality of cervical cancer (CC) is quite high and advanced CC is hard to cure. Accordingly, to find the mechanism of CC progression at molecular level is imminent. METHODS: The mRNA expression data were acquired from The Cancer Genome Atlas database, and squalene epoxidase (SQLE) level in the tumor and adjuvant tissues of CC was analyzed. The pathway enrichment analysis of target mRNAs was performed based on the GSEA database. The cancerous tissues and para-cancerous tissues of CC patients were collected for immunohistochemistry. SQLE and p53 mRNA expression was ensured by qRT-polymerase chain reaction. SQLE and p53 protein levels were determined by western blot. Cell functional assays focused on evaluating the malignant behaviors of cancer cells in each treatment group. Nude mouse xenograft models were constructed for tumorigenicity analysis. RESULTS: Bioinformatics analysis revealed that SQLE expression was high in CC tissues, which was linked to the poor prognosis. SQLE could affect the p53 signaling pathway. Cell functional assays demonstrated that SQLE expression was promoted in CC cell lines, and overexpressing SQLE facilitated the malignant phenotypes of CC cells, whereas silencing SQLE suppressed CC progression in vitro and in vivo. Besides, the repressed p53 signaling pathway could reverse the effect caused by silenced SQLE. CONCLUSION: SQLE could promote CC progression by modulating the p53 signaling pathway.