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The histone deacetylase HDAC3 is associated with the NCoR/SMRT co-repressor complex, and its canonical function is in transcriptional repression, but it can also activate transcription. Here, we show that the repressor and activator functions of HDAC3 can be genetically separated in Drosophila. A lysine substitution in the N terminus (K26A) disrupts its catalytic activity and activator function, whereas a combination of substitutions (HEBI) abrogating the interaction with SMRTER enhances repressor activity beyond wild type in the early embryo. We conclude that the crucial functions of HDAC3 in embryo development involve catalytic-dependent gene activation and non-enzymatic repression by several mechanisms, including tethering of loci to the nuclear periphery.
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Proteínas de Drosophila , Drosophila , Histona Desacetilases , Proteínas Repressoras , Animais , Drosophila/metabolismo , Regulação da Expressão Gênica , Proteínas Repressoras/metabolismo , Proteínas de Drosophila/metabolismo , Histona Desacetilases/metabolismoRESUMO
Anomalous pulmonary venous return (APVR) is a potentially lethal congenital heart disease. Elucidating the genetic etiology is crucial for understanding its pathogenesis and improving clinical practice, whereas its genetic basis remains largely unknown because of complex genetic etiology. We thus performed whole-exome sequencing for 144 APVR patients and 1636 healthy controls and report a comprehensive atlas of APVR-related rare genetic variants. Novel singleton, loss-of-function and deleterious missense variants (DVars) were enriched in patients, particularly for genes highly expressed in the developing human heart at the critical time point for pulmonary veins draining into the left atrium. Notably, PLXND1, encoding a receptor for semaphorins, represents a strong candidate gene of APVR (adjusted P = 1.1e-03, odds ratio: 10.9-69.3), accounting for 4.17% of APVR. We further validated this finding in an independent cohort consisting of 82 case-control pairs. In these two cohorts, eight DVars were identified in different patients, which convergently disrupt the GTPase-activating protein-related domain of PLXND1. All variant carriers displayed strikingly similar clinical features, in that all anomalous drainage of pulmonary vein(s) occurred on the right side and incorrectly connected to the right atrium, which may represent a novel subtype of APVR for molecular diagnosis. Studies in Plxnd1 knockout mice further revealed the effects of PLXND1 deficiency on severe heart and lung defects and cellular abnormalities related to APVR such as abnormal migration and vascular formation of vascular endothelial cells. These findings indicate the important role of PLXND1 in APVR pathogenesis, providing novel insights into the genetic etiology and molecular subtyping for APVR.
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Cardiopatias Congênitas , Veias Pulmonares , Síndrome de Cimitarra , Animais , Células Endoteliais , Átrios do Coração , Cardiopatias Congênitas/genética , Humanos , Peptídeos e Proteínas de Sinalização Intracelular , Glicoproteínas de Membrana , Camundongos , Veias Pulmonares/anormalidades , Síndrome de Cimitarra/genéticaRESUMO
SEH1 like nucleoporin (SEH1L) is an important component of nuclear pore complex (NPC), which is crucial in the regulation of cell division. However, the interrelation between SEH1L expression and tumor progression is less studied. In this research, we performed a systematic bioinformatic analysis about SEH1L using TCGA, Timer 2.0, Cbioportal, UCLAN and CellMiner™ databases in pan-cancer. Besides, we further validated the bioinformatic results through in vitro and in vivo experiments in HCC, including transcriptome sequencing, real-time quantitative PCR (RT-qPCR), western blotting (WB), immunohistochemistry (IHC), cell proliferation assays, clone formation, EdU, transwell, flow cytometry and subcutaneous tumor model. Our results suggested that SEH1L was significantly up-regulated and related to poor prognosis in most cancers, and may serve as a potential biomarker. SEH1L could promote HCC progression in vitro and in vivo. Besides, the next generation sequencing suggested that 684 genes was significantly up-regulated and 678 genes was down-regulated after the knock down of SEH1L. SEH1L siliencing could activate ATF3/HMOX1/GPX4 axis, decrease mitochondrial membrane potential and GSH, but increase ROS and MDA, and these effects could be reversed by the knock down of ATF3. This study indicated that SEH1L siliencing could induce ferroptosis and suppresses hepatocellular carcinoma (HCC) progression via ATF3/HMOX1/GPX4 axis.
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BACKGROUND: Pure red cell aplasia (PRCA) in neuromyelitis optica spectrum disorder (NMOSD) has not been reported before. This study presents a patient with NMOSD who developed PRCA. CASE PRESENTATION: A 54-year-old female was admitted in January 2023 for dysuria and progressive numbness and weakness of lower limbs. She had difficulty standing and walking in a straight line. Both lower limbs were positive for the Babinski and Chaddock signs. MRI showed abnormal signals in the spinal cord. Aquaporin-4-IgG (AQP-4-IgG) was positive (1:320), and NMOSD was confirmed. Intravenous immunoglobulin and methylprednisolone were given, and the symptoms improved. She received maintenance treatment with methylprednisolone tablets, and the dosage was gradually reduced. She was readmitted for fatigue, palpitations, and shortness of breath in May 2023. Bone marrow aspiration and biopsy showed elevated erythroid precursors and erythroid hypoplasia, with normal megakaryocytes and myeloid precursors. Chest CT showed no mediastinal lymph node enlargement or thymoma. PRCA secondary to NMOSD was diagnosed. Recombinant human erythropoietin was given. Her condition improved after 1.5 months, as indicated by blood cell count and imaging. CONCLUSIONS: This case suggests that PRCA can be secondary to NMOSD. A comprehensive immune function and bone marrow evaluation might be necessary if abnormal blood cells are found while managing NMOSD.
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Neuromielite Óptica , Aplasia Pura de Série Vermelha , Humanos , Feminino , Neuromielite Óptica/complicações , Neuromielite Óptica/tratamento farmacológico , Neuromielite Óptica/diagnóstico , Neuromielite Óptica/diagnóstico por imagem , Pessoa de Meia-Idade , Aplasia Pura de Série Vermelha/complicações , Aplasia Pura de Série Vermelha/diagnóstico , Aplasia Pura de Série Vermelha/tratamento farmacológico , Aquaporina 4/imunologiaRESUMO
Cardiomyopathy is a common disease of cardiac muscle that negatively affects cardiac function. HDAC3 commonly functions as corepressor by removing acetyl moieties from histone tails. However, a deacetylase-independent role of HDAC3 has also been described. Cardiac deletion of HDAC3 causes reduced cardiac contractility accompanied by lipid accumulation, but the molecular function of HDAC3 in cardiomyopathy remains unknown. We have used powerful genetic tools in Drosophila to investigate the enzymatic and nonenzymatic roles of HDAC3 in cardiomyopathy. Using the Drosophila heart model, we showed that cardiac-specific HDAC3 knockdown (KD) leads to prolonged systoles and reduced cardiac contractility. Immunohistochemistry revealed structural abnormalities characterized by myofiber disruption in HDAC3 KD hearts. Cardiac-specific HDAC3 KD showed increased levels of whole-body triglycerides and increased fibrosis. The introduction of deacetylase-dead HDAC3 mutant in HDAC3 KD background showed comparable results with wild-type HDAC3 in aspects of contractility and Pericardin deposition. However, deacetylase-dead HDAC3 mutants failed to improve triglyceride accumulation. Our data indicate that HDAC3 plays a deacetylase-independent role in maintaining cardiac contractility and preventing Pericardin deposition as well as a deacetylase-dependent role to maintain triglyceride homeostasis.
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Cardiomiopatias , Modelos Animais de Doenças , Proteínas de Drosophila , Drosophila melanogaster , Histona Desacetilases , Animais , Cardiomiopatias/enzimologia , Cardiomiopatias/genética , Cardiomiopatias/metabolismo , Cardiomiopatias/fisiopatologia , Drosophila melanogaster/enzimologia , Drosophila melanogaster/genética , Drosophila melanogaster/metabolismo , Proteínas de Drosophila/deficiência , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Técnicas de Silenciamento de Genes , Coração/fisiologia , Histona Desacetilases/deficiência , Histona Desacetilases/genética , Histona Desacetilases/metabolismo , Histonas/química , Histonas/metabolismo , Miocárdio/metabolismo , Triglicerídeos/metabolismo , HomeostaseRESUMO
BACKGROUND: The significant roles of circular RNAs (circRNAs) in different cancers and diseases have been reported. We now focused on the possible role of a newly recognized circRNA, circ_0004674 in triple-negative breast cancer (TNBC), and the related downstream mechanism. METHODS: The expression of circ_0004674 in TNBC tissues and cells was determined followed by analysis of the correlation between circ_0004674 and TNBC patients' prognosis. The interaction between circ_0004674, miR-377-3p, E2F6, and PNO1 was then identified using bioinformatics analysis combined with FISH, RIP, RNA pull-down, RT-qPCR, and Western blot analysis. Using gain-of-function and loss-of-function methods, we analyzed the effect of circ_0004674, miR-377-3p, E2F6, and PNO1 on TNBC in vivo and in vitro. RESULTS: Increased circ_0004674 and E2F6 but decreased miR-377-3p were observed in TNBC tissues and MDA-MB-231 TNBC cells, all of which findings were associated with poor prognosis in patients with TNBC. Silencing of circ_0004676 remarkably suppressed the proliferation, cell cycle progression, and migration of TNBC cells in vitro, as well as inhibiting tumorigenesis and metastasis in vivo. Additionally, circ_0004676 served as a sponge of miR-377-3p which bound to the transcription factor E2F6. In the presence of overexpression of circ_0004676, E2F6 expression and its target PNO1 expression were elevated, while miR-377-3p expression was decreased. Interestingly, overexpression of E2F6 could reverse the inhibitory effect on tumor growth caused by downregulation of circ_0004676. CONCLUSION: Our study highlighted the carcinogenic effect of circ_0004676 on TNBC through regulation of the miR-377-3p/E2F6/PNO1 axis. 1. Circ_0004674 is highly expressed in TNBC tissues and cells. 2. Circ_0004674 upregulates the expression of E2F6 by sponging miR-377-3p. 3. E2F6 upregulates PNO1 by binding to the PNO1 promoter. 4. Circ_0004674 favors TNBC progression by regulating the miR-377-3p/E2F6/PNO1 axis. 5. This study provides a new target for the treatment of TNBC.
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MicroRNAs , RNA Circular , Neoplasias de Mama Triplo Negativas , Humanos , Carcinogênese/genética , Linhagem Celular Tumoral , Proliferação de Células/genética , Transformação Celular Neoplásica , Biologia Computacional , Fator de Transcrição E2F6 , MicroRNAs/genética , Proteínas de Ligação a RNA , Neoplasias de Mama Triplo Negativas/genética , RNA Circular/genéticaRESUMO
T-lymphokine-activated killer cell-originated protein kinase (TOPK) is a serine-threonine kinase that is overexpressed in gastric cancer (GC) and promotes tumor progression. Polyphyllin VII (PPVII), a pennogenin isolated from the rhizomes of Paris polyphylla, shows anticancer effects. Here, we explored the antitumor activity and mechanism of PPVII in GC. Ferroptosis was detected by transmission electron microscope, malondialdehyde, and iron determination assays. Autophagy and its upstream signaling pathway were detected by Western blot, and gene alterations. The binding of PPVII and TOPK was examined through microscale thermophoresis and drug affinity responsive target stability assays. An in vivo mouse model was performed to evaluate the therapeutic of PPVII. PPVII inhibits GC by inducing autophagy-mediated ferroptosis. PPVII promotes the degradation of ferritin heavy chain 1, which is responsible for autophagy-mediated ferroptosis. PPVII activates the Unc-51-like autophagy-activating kinase 1 (ULK1) upstream of autophagy. PPVII inhibits the activity of TOPK, thereby weakening the inhibition of downstream ULK1. PPVII stabilizes the dimer of the inactive form of TOPK by direct binding. PPVII inhibits tumor growth without causing obvious toxicity in vivo. Collectively, this study suggests that PPVII is a potential agent for the treatment of GC by targeting TOPK to activate autophagy-mediated ferroptosis.
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Ferroptose , Neoplasias Gástricas , Humanos , Animais , Camundongos , Quinases de Proteína Quinase Ativadas por Mitógeno/metabolismo , Neoplasias Gástricas/tratamento farmacológico , Células Matadoras Ativadas por Linfocina/metabolismo , Autofagia , Linhagem Celular TumoralRESUMO
Millimeter-wave (mmWave) communication is considered a promising technology for fifth-generation (5G) wireless communications systems since it can greatly improve system throughput. Unfortunately, because of extremely high frequency, mmWave transmission suffers from the signal blocking problem, which leads to the deterioration of transmission performance. In this paper, we solve this problem by the combination of ultra-dense network (UDN) and user-centric virtual cell architecture. The deployment of dense small base stations (SBSs) in UDN can reduce transmission distance of signals. The user-centric virtual cell architecture mitigates and exploits interference to improve throughput by using coordinated multipoint (CoMP) transmission technology. Nonetheless, the backhaul burden is heavy and interbeam interference still severe. Therefore, we propose a novel iterative backhaul capacity-limited joint user association and power allocation (JUAPA) scheme in ultra-dense mmWave networks under user-centric virtual cell architecture. To mitigate interference and satisfy quality of service (QoS) requirements of users, a nonconvex system throughput optimization problem is formulated. To solve this intractable optimization problem, we divide it into two alternating optimization subproblems, i.e., user association and power allocation. During each iteration, a many-to-many matching algorithm is designed to solve user association. Subsequently, we perform power allocation optimization using a successive convex approximation (SCA) algorithm. The results confirm that the performance of the proposed scheme is close to that of the exhaustive searching scheme, which greatly reduces complexity, and clearly superior to that of traditional schemes in improving system throughput and satisfying QoS requirements.
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This study aims to investigate the effect and mechanism of hydnocarpin(HC) in treating triple negative breast cancer(TNBC). Cell counting kit-8(CCK-8), xCELLigence real-time cellular analysis(RTCA), and colony formation assay were employed to determine the effects of HC on the proliferation of two TNBC cell lines: MDA-MB-231 and MDA-MB-436. The effects of HC on the migration and invasion of TNBC cells were detected by high-content analysis, wound-healing assay, and Transwell assay. The changes in the epithelial-mesenchymal transition(EMT) and the expression of invasion-and migration-associated proteins [E-cadherin, vimentin, Snail, matrix metalloproteinase-2(MMP-2), and MMP-9] were detected by Western blot. Western blot and RT-qPCR were employed to determine the protein and mRNA levels of Yes-associated protein(YAP) and downstream targets(CTGF and Cyr61). TNBC cells were transfected with Flag-YAP for the overexpression of YAP, and the role of YAP as a key target for HC to inhibit TNBC malignant progression was examined by CCK-8 assay, Transwell assay, and wound-healing assay. The pathway of HC-induced YAP degradation was detected by the co-treatment of proteasome inhibitor with HC and ubiquitination assay. The binding of HC to YAP and the E3 ubiquitin ligase Ccr4-not transcription complex subunit 4(CNOT4) was detected by microscale thermophoresis(MST) assay and drug affinity responsive target stability(DARTS) assay. The results showed that HC significantly inhibited the proliferation, colony formation, invasion, and EMT of TNBC cells. HC down-regulated the protein and mRNA levels of CTGF and Cyr61. HC down-regulated the total protein level of YAP, while it had no effect on the mRNA level of YAP. The overexpression of YAP antagonized the inhibitory effects of HC on the proliferation, migration, and invasion of TNBC cells. HC promoted the degradation of YAP through the proteasome pathway and up-regulated the ubiquitination level of YAP. The results of MST and DARTS demonstrated direct binding between HC, YAP, and CNOT4. The above results indicated that HC inhibited the malignant progression of TNBC via CNOT4-mediated degradation and ubiquitination of YAP.
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Neoplasias de Mama Triplo Negativas , Humanos , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Neoplasias de Mama Triplo Negativas/genética , Neoplasias de Mama Triplo Negativas/metabolismo , Metaloproteinase 2 da Matriz/metabolismo , Linhagem Celular Tumoral , Proliferação de Células , Movimento Celular , Ubiquitinação , RNA Mensageiro/metabolismo , Transição Epitelial-Mesenquimal , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
RATIONALE: Transposition of the great arteries (TGA) is one of the most severe types of congenital heart diseases. Understanding the clinical characteristics and pathogenesis of TGA is, therefore, urgently needed for patient management of this severe disease. However, the clinical characteristics and genetic cause underlying TGA remain largely unexplored. OBJECTIVE: We sought to systematically examine the clinical characteristics and genetic cause for isolated nonsyndromic TGA. METHODS AND RESULTS: We recruited 249 patients with TGA (66 family trios) and performed whole-exome sequencing. The incidence of patent ductus arteriosus in dextro-TGA (52.7%) and dextrocardia/mesocardia in congenitally corrected TGA (32.8%) were significantly higher than that in other subtypes. A high prevalence of bicuspid pulmonic valve (9.6%) was observed in patients with TGA. Similar results were observed in a replication group of TGA (n=132). Through a series of bioinformatics filtering steps, we obtained 82 candidate genes harboring potentially damaging de novo, loss of function, compound heterozygous, or X-linked recessive variants. Established congenital heart disease-causing genes, such as FOXH1, were found among the list of candidate genes. A total of 19 ciliary genes harboring rare potentially damaging variants were also found; for example, DYNC2LI1 with a de novo putatively damaging variant. The enrichment of ciliary genes supports the roles of cilia in the pathogenesis of TGA. In total, 33% of the TGA probands had >1 candidate gene hit by putatively deleterious variants, suggesting that a portion of the TGA cases were probably affected by oligogenic or polygenic inheritance. CONCLUSIONS: The findings of clinical characteristic analyses have important implications for TGA patient stratification. The results of genetic analyses highlight the pathogenic role of ciliary genes and a complex genetic architecture underlying TGA.
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Cílios/metabolismo , Exoma/genética , Predisposição Genética para Doença/genética , Mutação de Sentido Incorreto , Transposição dos Grandes Vasos/genética , Sequência de Aminoácidos , Sequência de Bases , Estudos de Casos e Controles , Dineínas do Citoplasma/genética , Feminino , Humanos , Masculino , Homologia de Sequência de Aminoácidos , Sequenciamento do Exoma/métodosRESUMO
Dysregulation of the Hippo signaling pathway seen in many types of cancer is usually associated with a poor prognosis. Paris saponin VII (PSVII) is a steroid saponin isolated from traditional Chinese herbs with therapeutic action against various human cancers. In this study we investigated the effects of PSVII on human breast cancer (BC) cells and its anticancer mechanisms. We showed that PSVII concentration-dependently inhibited the proliferation of MDA-MB-231, MDA-MB-436 and MCF-7 BC cell lines with IC50 values of 3.16, 3.45, and 2.86 µM, respectively, and suppressed their colony formation. PSVII (1.2-1.8 µM) induced caspase-dependent apoptosis in the BC cell lines. PSVII treatment also induced autophagy and promoted autophagic flux in the BC cell lines. PSVII treatment decreased the expression and nuclear translocation of Yes-associated protein (YAP), a downstream transcriptional effector in the Hippo signaling pathway; overexpression of YAP markedly attenuated PSVII-induced autophagy. PSVII-induced, YAP-mediated autophagy was associated with increased active form of LATS1, an upstream effector of YAP. The activation of LATS1 was involved the participation of multiple proteins (including MST2, MOB1, and LATS1 itself) in an MST2-dependent sequential activation cascade. We further revealed that PSVII promoted the binding of LATS1 with MST2 and MOB1, and activated LATS1 in the BC cell lines. Molecular docking showed that PSVII directly bound to the MST2-MOB1-LATS1 ternary complex. Microscale thermophoresis analysis and drug affinity responsive targeting stability assay confirmed the high affinity between PSVII and the MST2-MOB1-LATS1 ternary complex. In mice bearing MDA-MB-231 cell xenograft, administration of PSVII (1.5 mg/kg, ip, 4 times/week, for 4 weeks) significantly suppressed the tumor growth with increased pLATS1, LC3-II and Beclin 1 levels and decreased YAP, p62 and Ki67 levels in the tumor tissue. Overall, this study demonstrates that PSVII is a novel and direct Hippo activator that has great potential in the treatment of BC.
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Neoplasias da Mama , Saponinas , Animais , Autofagia , Neoplasias da Mama/tratamento farmacológico , Proliferação de Células , Feminino , Via de Sinalização Hippo , Humanos , Camundongos , Simulação de Acoplamento Molecular , Proteínas Serina-Treonina Quinases , Saponinas/farmacologia , Saponinas/uso terapêuticoRESUMO
This publication has been retracted by the Editor due to concerns regarding the originality of the figure images. Reference: Yanting Chai, Ke Xiang, Yezi Wu, Te Zhang, Ying Liu, Xuewen Liu, Weiguo Zhen, Yuan Si. Cucurbitacin B Inhibits the Hippo-YAP Signaling Pathway and Exerts Anticancer Activity in Colorectal Cancer Cells. Med Sci Monit, 2018; 24: LBR9251-9258. DOI: 10.12659/MSM.911594.
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With the development of metal-based drugs, Ru(II) compounds present potential applications of PDT (photodynamic therapy) and anticancer reagents. We herein synthesized two naphthyl-appended ruthenium complexes by the combination of the ligand with naphthyl and bipyridyl. The DNA affinities, photocleavage abilities, and photocytotoxicity were studied by various spectral methods, viscosity measurement, theoretical computation method, gel electrophoresis, and MTT method. Two complexes exhibited strong interaction with calf thymus DNA by intercalation. Production of singlet oxygen (1O2) led to obvious DNA photocleavage activities of two complexes under 365 nm light. Furthermore, two complexes displayed obvious photocytotoxicity and low dark cytotoxicity towards Hela, A549, and A375 cells.
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Complexos de Coordenação , Rutênio , Complexos de Coordenação/farmacologia , DNA , Clivagem do DNA , Células HeLa , Humanos , Simulação de Acoplamento Molecular , Rutênio/farmacologiaRESUMO
This study aims to investigate the effect of ethoxysanguinarine(Eth) on cisplatin(DDP)-resistant human gastric cancer cells and decipher the underlying mechanism. The human gastric cancer cell line SGC7901 and the DDP-resistant cell line SGC7901/DDP were used as the cell models. Western blot was employed to determine the expression levels of multidrug resistance-related proteins, and methyl thiazolyl tetrazolium(MTT) assay to detect the proliferation of SGC7901 and SGC7901/DDP cells exposed to DDP. After treatment with different concentrations of Eth, the proliferation of SGC7901 and SGC7901/DDP cells was detected by MTT assay, trypan blue exclusion assay, colony formation assay, and high-content imaging and analysis system. The apoptosis of SGC7901/DDP cells was detected by flow cytometry with Annexin V-FITC/PI staining. GFP-LC3 transfection was carried out to detect the effect of Eth on the autophagy of SGC7901/DDP cells. The expression levels of the multidrug resistance-related protein P-glycoprotein(P-gp), the apoptosis-related proteins [caspase-9, caspase-3, and poly(ADP-ribose) polymerase(PARP)], the autophagy-related protein light chain 3-â ¡(LC3-â ¡), the key effectors [mammalian target of rapamycin(mTOR), 70 kDa ribosomal protein S6 kinase(P70 S6 K), and 4 E binding protein 1(4 E-BP1)] of the mammalian target of rapamycin complex 1(mTORC1) signaling pathway, cancerous inhibitor of protein phosphatase 2A(CIP2A), and protein kinase B(Akt) were measured by Western blot. The mRNA level of CIP2A in the SGC7901/DDP cells exposed to Eth for 24 h was analyzed by RT-qPCR. After SGC7901/DDP cells were transfected with CIP2A expression vector pcDNA3.1-HA-CIP2A and treated with different concentrations of Eth, MTT assay was used to determine the prolife-ration of SGC7901/DDP cells and Western blot to detect the expression levels of related proteins. The interaction sites of Eth and CIP2A were predicted by molecular docking. The affinity between Eth and CIP2A was determined by drug affinity responsive target stability(DARTS) assay. The pharmacokinetic properties and drug-like activity of Eth were predicted by SwissADME. The results indicated that SGC7901/DDP cells were more sensitive to Eth than SGC7901 cells. Eth significantly inhibited proliferation and colony formation and changed the morphology, roundness, and area of SGC7901/DDP cells. Eth treatment caused the nucleus shrinking and significantly increased the apoptosis rate of the cells. Furthermore, Eth down-regulated the expression of caspase-9 and caspase-3 precursors and promoted the cleavage of PARP, which suggested that Eth induced the apoptosis of SGC7901/DDP cells. The GFP-LC3 in Eth-treated cells showed speckled aggregation. The up-regulated expression of LC3-â ¡ by Eth indicated that Eth activated the autophagy of SGC7901/DDP cells. Eth down-regulated the expression of P-gp, the phosphorylation of mTOR, P70 S6K, and 4E-BP1, the expression of CIP2A, and the phosphorylation of Akt. Additionally, it increased the activity of PP2A, and had no significant effect on the expression of CIP2A in SGC7901/DDP cells. CIP2A overexpression antagonized the inhibition of cell proliferation and the activation of autophagy by Eth. Molecular docking suggested that Eth bound to CIP2A. The results of DARTS assay further proved the above binding effect. Eth has potential drug-like activity. The above results demonstrated that Eth inhibited the proliferation, induced the apoptosis, and activated the autophagy of SGC7901/DDP cells by targeting CIP2A and then down-regulating PP2A/mTORC1 signaling pathway. This study provided a new target for the treatment of cisplatin-resistant gastric cancer.
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Antineoplásicos , Neoplasias Gástricas , Humanos , Cisplatino/farmacologia , Cisplatino/uso terapêutico , Caspase 9/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Caspase 3/metabolismo , Neoplasias Gástricas/tratamento farmacológico , Neoplasias Gástricas/genética , Neoplasias Gástricas/metabolismo , Resistencia a Medicamentos Antineoplásicos , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Simulação de Acoplamento Molecular , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Inibidores de Poli(ADP-Ribose) Polimerases/uso terapêutico , Autofagia , Apoptose , Proliferação de Células , Proteínas Reguladoras de Apoptose/metabolismo , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Linhagem Celular TumoralRESUMO
The present study investigated the mechanism of polyphyllin A(PPA) in inhibiting gastric cancer(GC) cells. GC cells(SGC7901 and MGC803 cell lines) were treated with PPA at different concentrations. The effect of PPA on the proliferation of GC cells was detected by MTT assay, real-time cell analysis(RTCA) assay, and clone-forming assay, respectively. Reactive oxygen species(ROS) of GC cells was detected by flow cytometry. The change of mitochondrial membrane potential was detected by JC-1 assay. The expression and phosphorylation levels of apoptosis-related proteins(caspase-9, caspase-3, and PARP) and proteins related to the signaling pathway(ETS-1, CIP2 A, and Akt) were detected by Western blot. The binding sites of PPA to ETS-1 were analyzed by molecular docking. The affinity of PPA and ETS-1 was detected by drug affinity responsive target stability(DARTS) assay. PPA had a significant inhibitory effect on the proliferation and colony formation of GC cells at a low concentration. The PPA groups showed increased ROS and decreased mitochondrial membrane potential. PPA down-regulated the precursor expression of caspase-9 and caspase-3 and promoted the cleavage of PARP, suggesting that PPA induced the apoptosis of GC cells through the mitochondrial pathway. PPA significantly reduced expression levels of CIP2 A and the phosphorylation of downstream Akt. Molecular docking showed that PPA bound to the ETS domain of ETS-1, the transcription factor of CIP2 A, and formed hydrogen bonds with Pro319 and Asp317. DARTS assay further confirmed that PPA significantly prevented the hydrolysis of ETS-1 by pronase, which was inductive of the direct binding effect of PPA and ETS-1. PPA inhibits the proliferation and induces the apoptosis of GC cells by directly targeting ETS-1 to down-regulate the ETS-1/CIP2 A/Akt signaling pathway.
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Neoplasias Gástricas , Apoptose , Linhagem Celular Tumoral , Proliferação de Células , Humanos , Simulação de Acoplamento Molecular , Neoplasias Gástricas/tratamento farmacológico , Neoplasias Gástricas/genética , Neoplasias Gástricas/metabolismoRESUMO
The role of O-linked N-acetylglucosamine (O-GlcNAc) modification in the cell cycle has been enigmatic. Previously, both O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA) disruptions have been shown to derail the mitotic centrosome numbers, suggesting that mitotic O-GlcNAc oscillation needs to be in concert with mitotic progression to account for centrosome integrity. Here, using both chemical approaches and biological assays with HeLa cells, we attempted to address the underlying molecular mechanism and observed that incubation of the cells with the OGA inhibitor Thiamet-G strikingly elevates centrosomal distances, suggestive of premature centrosome disjunction. These aberrations could be overcome by inhibiting Polo-like kinase 1 (PLK1), a mitotic master kinase. PLK1 inactivation is modulated by the myosin phosphatase targeting subunit 1 (MYPT1)-protein phosphatase 1cß (PP1cß) complex. Interestingly, MYPT1 has been shown to be abundantly O-GlcNAcylated, and the modified residues have been detected in a recent O-GlcNAc-profiling screen utilizing chemoenzymatic labeling and bioorthogonal conjugation. We demonstrate here that MYPT1 is O-GlcNAcylated at Thr-577, Ser-585, Ser-589, and Ser-601, which antagonizes CDK1-dependent phosphorylation at Ser-473 and attenuates the association between MYPT1 and PLK1, thereby promoting PLK1 activity. We conclude that under high O-GlcNAc levels, PLK1 is untimely activated, conducive to inopportune centrosome separation and disruption of the cell cycle. We propose that too much O-GlcNAc is equally deleterious as too little O-GlcNAc, and a fine balance between the OGT/OGA duo is indispensable for successful mitotic divisions.
Assuntos
Centrossomo/metabolismo , Mitose , Fosfatase de Miosina-de-Cadeia-Leve/metabolismo , Proteína Quinase CDC2/genética , Proteína Quinase CDC2/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular , Glicosilação , Humanos , Fosfatase de Miosina-de-Cadeia-Leve/genética , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Quinase 1 Polo-LikeRESUMO
G protein pathway suppressor 2 (GPS2) is expressed in most human tissues, including the stomach. However, the biological functions of GPS2 in cancer, as well as the underlying molecular mechanisms, remain poorly understood. Here, we report that GPS2 expression was aberrantly downregulated in gastric cancer (GC) tissues compared with control tissues. Clinicopathologic analysis showed that low GPS2 expression was significantly correlated with pathological grade, lymph node stage, and invasive depth. Kaplan-Meier analysis indicated that patients with low GPS2 expression showed poorer overall survival rates than those with high GPS2 expression. Moreover, GPS2 overexpression decreased GC cell proliferation, colony formation, tumorigenesis, and invasion. Overexpression of GPS2 reduced the protein expression of epidermal growth factor receptor (EGFR) and inhibited its downstream signaling in GC cells. Interestingly, GPS2 decreased EGFR protein expression, which was reversed by a lysosome inhibitor. Furthermore, GPS2 reduced EGFR protein stability by enhancing the binding of EGFR and an E3 ligase, c-Cbl, which promoted the ubiquitination of EGFR, ultimately leading to its degradation through the lysosomal pathway. Further analysis indicated that GPS2 activated autophagy and promoted the autophagic flux by destabilizing EGFR. Taken together, these results suggest that low GPS2 expression is associated with GC progression and provide insights into the applicability of the GPS2-EGFR axis as a potential therapeutic target in GC.
Assuntos
Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Neoplasias Gástricas/patologia , Estudos de Casos e Controles , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Progressão da Doença , Regulação para Baixo , Receptores ErbB/química , Receptores ErbB/metabolismo , Feminino , Humanos , Metástase Linfática , Masculino , Gradação de Tumores , Transplante de Neoplasias , Prognóstico , Estabilidade Proteica , Neoplasias Gástricas/genética , Neoplasias Gástricas/metabolismo , Análise de Sobrevida , UbiquitinaçãoRESUMO
Sulfurimonas species (class Campylobacteria, phylum Campylobacterota) were globally distributed and especially predominant in deep-sea hydrothermal environments. They were previously identified as chemolithoautotrophic sulfur-oxidizing bacteria (SOB), whereas little is known about their potential in sulfur reduction. In this report, we found that the elemental sulfur reduction is quite common in different species of genus Sulfurimonas. To gain insights into the sulfur reduction mechanism, growth tests, morphology observation, as well as genomic and transcriptomic analyses were performed on a deep-sea hydrothermal vent bacterium Sulfurimonas sp. NW10. Scanning electron micrographs and dialysis tubing tests confirmed that elemental sulfur reduction occurred without direct contact of cells with sulfur particles while direct access strongly promoted bacterial growth. Furthermore, we demonstrated that most species of Sulfurimonas probably employ both periplasmic and cytoplasmic polysulfide reductases, encoded by genes psrA1 B1 CDE and psrA2 B2 , respectively, to accomplish cyclooctasulfur reduction. This is the first report showing two different sulfur reduction pathways coupled to different energy conservations could coexist in one sulfur-reducing microorganism, and demonstrates that most bacteria of Sulfurimonas could employ both periplasmic and cytoplasmic polysulfide reductases to perform cyclooctasulfur reduction. The capability of sulfur reduction coupling with hydrogen oxidation may partially explain the prevalenceof Sulfurimonas in deep-sea hydrothermal vent environments.
Assuntos
Helicobacteraceae/metabolismo , Fontes Hidrotermais/microbiologia , Enxofre/metabolismo , Crescimento Quimioautotrófico , DNA Bacteriano/genética , Helicobacteraceae/classificação , Helicobacteraceae/genética , Helicobacteraceae/isolamento & purificação , Oxirredução , Filogenia , RNA Ribossômico 16S/genética , Água do Mar/microbiologiaRESUMO
Two novel Gram-strain-negative and rod-shaped bacteria, designated strain G1T and G2T, were isolated from sediment samples collected from the coast of Xiamen, PR China. The cells were motile by a single polar flagellum. Growth of strain G1T occurred at 10-40 °C (optimum, 30 °C), at pH 6.0-9.0 (optimum, pH 7.5) and with 5-1530 mM NaCl (optimum, 510 mM), while the temperature, pH and NaCl concentration ranges for G2T were 4-45 °C (optimum, 28 °C), pH 5.5-8.0 (optimum, pH 6.5) and 85-1530 mM NaCl (optimum, 340 mM). The two isolates were obligate chemolithoautotrophs capable of using thiosulfate, sulfide, elemental sulphur or tetrathionate as an energy source. Strain G1T used molecular oxygen or nitrite as an electron acceptor, while strain G2T used molecular oxygen as the sole electron acceptor. The dominant fatty acids of G1T and G2T were summed feature 3 (C16:1 ω7c and/or C16:1 ω6c), C16 : 0 and summed feature 8 (C18:1 ω7c and/or C18:1 ω6c). The DNA G+C content of G1T and G2T were 45.1 and 48.3âmol%, respectively. Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain G1T and G2T were members of the genus Thiomicrorhabdus, and most closely related to Thiomicrorhabdus hydrogeniphila MAS2T (96.0â%) and Thiomicrorhabdus indica 13-15AT (95.4â%), respectively. The 16S rRNA gene sequence similarity between strains G1T and G2T was 95.8â%. Based on the phylogenetic, genomic and phenotypic data presented here, the isolate strains represent novel species of the genus Thiomicrorhabdus, for which the names Thiomicrorhabdus sediminis sp. nov. (type strain G1T=MCCC 1A14511T=KCTC 15841T) and Thiomicrorhabdus xiamenensis sp. nov. (type strain G2T=MCCC 1A14512T=KCTC 15842T) are proposed.
Assuntos
Sedimentos Geológicos/microbiologia , Filogenia , Piscirickettsiaceae/classificação , Água do Mar/microbiologia , Bactérias Redutoras de Enxofre/classificação , Técnicas de Tipagem Bacteriana , Composição de Bases , China , DNA Bacteriano/genética , Ácidos Graxos/química , Oxirredução , Fosfolipídeos/química , Piscirickettsiaceae/isolamento & purificação , RNA Ribossômico 16S/genética , Análise de Sequência de DNA , Enxofre , Bactérias Redutoras de Enxofre/isolamento & purificaçãoRESUMO
A novel marine hydrogen- and sulfur-oxidizing bacterium, designated strain S2-6 T, was isolated from the deep-sea sediment samples at the Longqi hydrothermal system, southwestern Indian Ocean. Cells were Gram-stain-negative, motile, short rods with a single polar flagellum. Growth was observed at 10-45 °C (optimum 33 °C), pH 5.0-8.0 (optimum pH 7.0) and 1.5 to 6.0% (w/v) NaCl with an optimum at 3.0% (w/v). The isolate was an obligate chemolithoautotroph capable of growth using thiosulfate, tetrathionate, elemental sulfur or sodium sulfide as the energy source, and oxygen or nitrate as the sole electron acceptor. When hydrogen was used as the energy source, strain S2-6 T could respire oxygen, nitrate or element sulfur. The major cellular fatty acids of strain S2-6 T were summed feature 3 (C16:1ω7c and/or C16:1ω6c), C16:0 and summed feature 8 (C18:1ω7c and/or C18:1ω6c). The total size of its genome was 2,320,257 bp and the genomic DNA G + C content was 37.3 mol%. Phylogenetic analysis based on 16S rRNA gene sequences and core genes showed that the novel isolate belonged to the genus Sulfurimonas and was most closely related to Sulfurimonas paralvinellae GO25T (96.8% sequence identity) and Sulfurimonas autotrophica OK10T (95.8% sequence identity). The average nucleotide identity and DNA-DNA hybridization values between strain S2-6 T and S. paralvinellae GO25T and S. autotrophica OK10T were 74.6%-81.2% and 19.1%-24.6%, respectively. Based on the polyphase taxonomical data, strain S2-6 T represents a novel species of the genus Sulfurimonas, for which the name Sulfurimonas sediminis sp. nov. is proposed, with the type strain S2-6 T (= MCCC 1A14513T = KCTC 15854 T).