RESUMO
Mutations in the RNA helicase DDX3 have emerged as a frequent cause of intellectual disability in humans. Because many individuals carrying DDX3 mutations have additional defects in craniofacial structures and other tissues containing neural crest (NC)-derived cells, we hypothesized that DDX3 is also important for NC development. Using Xenopus tropicalis as a model, we show that DDX3 is required for normal NC induction and craniofacial morphogenesis by regulating AKT kinase activity. Depletion of DDX3 decreases AKT activity and AKT-dependent inhibitory phosphorylation of GSK3ß, leading to reduced levels of ß-catenin and Snai1: two GSK3ß substrates that are crucial for NC induction. DDX3 function in regulating these downstream signaling events during NC induction is likely mediated by RAC1, a small GTPase whose translation depends on the RNA helicase activity of DDX3. These results suggest an evolutionarily conserved role of DDX3 in NC development by promoting AKT activity, and provide a potential mechanism for the NC-related birth defects displayed by individuals harboring mutations in DDX3 and its downstream effectors in this signaling cascade.
Assuntos
RNA Helicases DEAD-box/metabolismo , Crista Neural/embriologia , Crista Neural/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas de Xenopus/metabolismo , Xenopus/embriologia , Xenopus/metabolismo , Animais , Cartilagem/embriologia , Cartilagem/metabolismo , Embrião não Mamífero/metabolismo , Face/embriologia , Regulação da Expressão Gênica no Desenvolvimento , Glicogênio Sintase Quinase 3 beta/metabolismo , Morfogênese/genética , Fosforilação , Estabilidade Proteica , Crânio/embriologia , Crânio/metabolismo , Fatores de Transcrição da Família Snail/metabolismo , Via de Sinalização Wnt , Xenopus/genética , beta Catenina/metabolismo , Proteínas rac1 de Ligação ao GTP/metabolismoRESUMO
Ephrin-B signaling has been implicated in many normal and pathological processes, including neural crest development and tumor metastasis. We showed previously that proteolysis of ephrin-B ligands by the disintegrin metalloprotease ADAM13 is necessary for canonical Wnt signal activation and neural crest induction in Xenopus, but it was unclear if these mechanisms are conserved in mammals. Here, we report that mammalian ADAM9 cleaves ephrin-B1 and ephrin-B2 and can substitute for Xenopus ADAM13 to induce the neural crest. We found that ADAM9 expression is elevated in human colorectal cancer (CRC) tissues and that knockdown (KD) of ADAM9 inhibits the migration and invasion of SW620 and HCT116 CRC cells by reducing the activity of Akt kinase, which is antagonized by ephrin-Bs. Akt is a signaling node that activates multiple downstream pathways, including the Wnt and mTOR pathways, both of which can promote CRC cell migration/invasion. Surprisingly, we also found that KD of ADAM9 downregulates Wnt signaling but has negligible effects on mTOR signaling in SW620 cells; in contrast, mTOR activity is suppressed while Wnt signaling remains unaffected by ADAM9 KD in HCT116 cells. These results suggest that mammalian ADAM9 cleaves ephrin-Bs to derepress Akt and promote CRC migration and invasion; however, the signaling pathways downstream of Akt are differentially regulated by ADAM9 in different CRC cell lines, reflecting the heterogeneity of CRC cells in responding to manipulations of upstream Akt regulators.
Assuntos
Proteínas ADAM/metabolismo , Neoplasias Colorretais , Efrinas , Animais , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Neoplasias Colorretais/patologia , Humanos , Ligantes , Mamíferos/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Metaloproteases/metabolismo , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Via de Sinalização WntRESUMO
High pliability and promiscuity are observed widely exist in plant specialized metabolism, especially the hydroxycinnamic acid metabolism. Here, we identified an addition BAHD acyltransferase (EpHMT) that catalyzes phaselic acid biosynthesis and found that the substrate promiscuities of identified BAHD and SCPL acyltransferases are responsible for the diversity of hydroxycinnamic acid derivatives in purple coneflower.
Assuntos
Produtos Biológicos , Echinacea , Aciltransferases/genética , Aciltransferases/metabolismo , Ácidos Cumáricos , Echinacea/metabolismo , Plantas/metabolismoRESUMO
BACKGROUND: The eight phenotypically distinguishable indigenous chicken breeds in Guizhou province of China are great resources for high-quality development of the poultry industry in China. However, their full value and potential have yet to be understood in depth. To illustrate the genetic diversity, the relationship and population structure, and the genetic variation patterns shaped by selection in Guizhou indigenous chickens, we performed a genome-wide analysis of 240 chickens from 8 phenotypically and geographically representative Guizhou chicken breeds and 60 chickens from 2 commercial chicken breeds (one broiler and one layer), together with 10 red jungle fowls (RJF) genomes available from previous studies. RESULTS: The results obtained in this present study showed that Guizhou chicken breed populations harbored higher genetic diversity as compared to commercial chicken breeds, however unequal polymorphisms were present within Guizhou indigenous chicken breeds. The results from the population structure analysis markedly reflected the breeding history and the geographical distribution of Guizhou indigenous chickens, whereas, some breeds with complex genetic structure were ungrouped into one cluster. In addition, we confirmed mutual introgression within Guizhou indigenous chicken breeds and from commercial chicken breeds. Furthermore, selective sweep analysis revealed candidate genes which were associated with specific and common phenotypic characteristics evolved rapidly after domestication of Guizhou local chicken breeds and economic traits such as egg production performance, growth performance, and body size. CONCLUSION: Taken together, the results obtained from the comprehensive analysis of the genetic diversity, genetic relationships and population structures in this study showed that Guizhou indigenous chicken breeds harbor great potential for commercial utilization, however effective conservation measures are currently needed. Additionally, the present study drew a genome-wide selection signature draft for eight Guizhou indigenous chicken breeds and two commercial breeds, as well as established a resource that can be exploited in chicken breeding programs to manipulate the genes associated with desired phenotypes. Therefore, this study will provide an essential genetic basis for further research, conservation, and breeding of Guizhou indigenous chickens.
Assuntos
Galinhas , Polimorfismo de Nucleotídeo Único , Animais , Galinhas/genética , Genoma , Fenótipo , China , Variação GenéticaRESUMO
Mutations of the RNA granule component TDRD7 (OMIM: 611258) cause pediatric cataract. We applied an integrated approach to uncover the molecular pathology of cataract in Tdrd7-/- mice. Early postnatal Tdrd7-/- animals precipitously develop cataract suggesting a global-level breakdown/misregulation of key cellular processes. High-throughput RNA sequencing integrated with iSyTE-bioinformatics analysis identified the molecular chaperone and cytoskeletal modulator, HSPB1, among high-priority downregulated candidates in Tdrd7-/- lens. A protein fluorescence two-dimensional difference in-gel electrophoresis (2D-DIGE)-coupled mass spectrometry screen also identified HSPB1 downregulation, offering independent support for its importance to Tdrd7-/- cataractogenesis. Lens fiber cells normally undergo nuclear degradation for transparency, posing a challenge: how is their cell morphology, also critical for transparency, controlled post-nuclear degradation? HSPB1 functions in cytoskeletal maintenance, and its reduction in Tdrd7-/- lens precedes cataract, suggesting cytoskeletal defects may contribute to Tdrd7-/- cataract. In agreement, scanning electron microscopy (SEM) revealed abnormal fiber cell morphology in Tdrd7-/- lenses. Further, abnormal phalloidin and wheat germ agglutinin (WGA) staining of Tdrd7-/- fiber cells, particularly those exhibiting nuclear degradation, reveals distinct regulatory mechanisms control F-actin cytoskeletal and/or membrane maintenance in post-organelle degradation maturation stage fiber cells. Indeed, RNA immunoprecipitation identified Hspb1 mRNA in wild-type lens lysate TDRD7-pulldowns, and single-molecule RNA imaging showed co-localization of TDRD7 protein with cytoplasmic Hspb1 mRNA in differentiating fiber cells, suggesting that TDRD7-ribonucleoprotein complexes may be involved in optimal buildup of key factors. Finally, Hspb1 knockdown in Xenopus causes eye/lens defects. Together, these data uncover TDRD7's novel upstream role in elevation of stress-responsive chaperones for cytoskeletal maintenance in post-nuclear degradation lens fiber cells, perturbation of which causes early-onset cataracts.
Assuntos
Catarata/genética , Proteínas do Olho/genética , Proteínas de Choque Térmico/genética , Chaperonas Moleculares/genética , Ribonucleoproteínas/genética , Animais , Catarata/patologia , Núcleo Celular/genética , Citoesqueleto/genética , Modelos Animais de Doenças , Oftalmopatias , Humanos , Cristalino/metabolismo , Cristalino/patologia , Camundongos , Microscopia Eletrônica de Varredura , Mutação/genética , RNA Mensageiro/genética , Xenopus laevis/genéticaRESUMO
The disordered tubulin C-terminal tail (CTT), which possesses a higher degree of heterogeneity, is the target for the interaction of many proteins and cellular components. Compared to the seven well-described binding sites of microtubule-targeting agents (MTAs) that localize on the globular tubulin core, tubulin CTT is far less explored. Therefore, tubulin CTT can be regarded as a novel site for the development of MTAs with distinct biochemical and cell biological properties. Here, we designed and synthesized linear and cyclic peptides containing multiple arginines (RRR), which are complementary to multiple acidic residues in tubulin CTT. Some of them showed moderate induction and promotion of tubulin polymerization. The most potent macrocyclic compound 1f was found to bind to tubulin CTT and thus exert its bioactivity. Such RRR containing compounds represent a starting point for the discovery of tubulin CTT-targeting agents with therapeutic potential.
Assuntos
Antineoplásicos , Tubulina (Proteína) , Tubulina (Proteína)/metabolismo , Microtúbulos/metabolismo , Antineoplásicos/farmacologia , Sítios de Ligação , Moduladores de Tubulina/farmacologia , Moduladores de Tubulina/metabolismoRESUMO
Farnesoid X receptor (FXR) ligands have been actively pursued to treat metabolic disorders, liver and bile diseases, among others. Starting from a widely occurring natural product, oleanolic acid (OA), we discovered potent and selective FXR modulator from the 12ß-oxygenated OA alkyl esters, with the assistance of molecular modeling. The representative compound 7b modulated some FXR downstream genes involved in glucose and lipid metabolism in cells, and significantly improved hyperglycemia in KKay fat mice fed with high fat diet, through the reduction of mRNA expression of gluconeogenesis genes PEPCK and G6Pase. This study provides a new series of selective FXR modulator, as well as the in vitro and in vivo evidence for their potential to improve hyperglycemia in diabetic mice through FXR antagonism.
Assuntos
Diabetes Mellitus Experimental , Diabetes Mellitus Tipo 2 , Hiperglicemia , Hipoglicemiantes , Ácido Oleanólico , Receptores Citoplasmáticos e Nucleares , Animais , Camundongos , Ácidos e Sais Biliares/metabolismo , Diabetes Mellitus Experimental/induzido quimicamente , Diabetes Mellitus Experimental/tratamento farmacológico , Diabetes Mellitus Experimental/metabolismo , Ésteres/química , Ésteres/farmacologia , Ésteres/uso terapêutico , Hiperglicemia/tratamento farmacológico , Fígado/metabolismo , Camundongos Endogâmicos C57BL , Ácido Oleanólico/química , Ácido Oleanólico/farmacologia , Ácido Oleanólico/uso terapêutico , Receptores Citoplasmáticos e Nucleares/antagonistas & inibidores , Hipoglicemiantes/química , Hipoglicemiantes/farmacologia , Hipoglicemiantes/uso terapêutico , Diabetes Mellitus Tipo 2/tratamento farmacológico , Gluconeogênese/efeitos dos fármacosRESUMO
BACKGROUND: Currently, the rate of morbidity and mortality in acute respiratory distress syndrome (ARDS) remains high. One of the potential reasons for the poor and ineffective therapies is the lack of early and credible indicator of risk prediction that would help specific treatment of severely affected ARDS patients. Nevertheless, assessment of the clinical outcomes with transcriptomics of ARDS by alveolar macrophage has not been performed. METHODS: The expression data GSE116560 was obtained from the Gene Expression Omnibus databases (GEO) in NCBI. This dataset consists of 68 BAL samples from 35 subjects that were collected within 48 h of ARDS. Differentially expressed genes (DEGs) of different outcomes were analyzed using R software. The top 10 DEGs that were up- or down-regulated were analyzed using receiver operating characteristic (ROC) analysis. Kaplan-Meier survival analysis within two categories according to cut-off and the value of prediction of the clinical outcomes via DEGs was verified. GO enrichment, KEGG pathway analysis, and protein-protein interaction were also used for functional annotation of key genes. RESULTS: 24,526 genes were obtained, including 235 up-regulated and 292 down-regulated DEGs. The gene ADORA3 was chosen as the most obvious value to predict the outcome according to the ROC and survival analysis. For functional annotation, ADORA3 was significantly augmented in sphingolipid signaling pathway, cGMP-PKG signaling pathway, and neuroactive ligand-receptor interaction. Four genes (ADORA3, GNB1, NTS, and RHO), with 4 nodes and 6 edges, had the highest score in these clusters in the protein-protein interaction network. CONCLUSIONS: Our results show that the prognostic prediction of early biomarkers of transcriptomics as identified in alveolar macrophage in ARDS can be extended for mechanically ventilated critically ill patients. In the long term, generalizing the concept of biomarkers of transcriptomics in alveolar macrophage could add to improving precision-based strategies in the ICU patients and may also lead to identifying improved strategy for critically ill patients.
Assuntos
Síndrome do Desconforto Respiratório , Transcriptoma , Biomarcadores , Estado Terminal , Perfilação da Expressão Gênica/métodos , Humanos , Macrófagos Alveolares , Prognóstico , Síndrome do Desconforto Respiratório/genéticaRESUMO
BACKGROUND: 2-Oxoglutarate and Fe(II)-dependent dioxygenases (2ODDs) belong to the 2-oxoglutarate-dependent dioxygenase (2OGD) superfamily and are involved in various vital metabolic pathways of plants at different developmental stages. These proteins have been extensively investigated in multiple model organisms. However, these enzymes have not been systematically analyzed in tomato. In addition, type I flavone synthase (FNSI) belongs to the 2ODD family and contributes to the biosynthesis of flavones, but this protein has not been characterized in tomato. RESULTS: A total of 131 2ODDs from tomato were identified and divided into seven clades by phylogenetic classification. The Sl2ODDs in the same clade showed similar intron/exon distributions and conserved motifs. The Sl2ODDs were unevenly distributed across the 12 chromosomes, with different expression patterns among major tissues and at different developmental stages of the tomato growth cycle. We characterized several Sl2ODDs and their expression patterns involved in various metabolic pathways, such as gibberellin biosynthesis and catabolism, ethylene biosynthesis, steroidal glycoalkaloid biosynthesis, and flavonoid metabolism. We found that the Sl2ODD expression patterns were consistent with their functions during the tomato growth cycle. These results indicated the significance of Sl2ODDs in tomato growth and metabolism. Based on this genome-wide analysis of Sl2ODDs, we screened six potential FNSI genes using a phylogenetic tree and coexpression analysis. However, none of them exhibited FNSI activity. CONCLUSIONS: Our study provided a comprehensive understanding of the tomato 2ODD family and demonstrated the significant roles of these family members in plant metabolism. We also suggest that no FNSI genes in tomato contribute to the biosynthesis of flavones.
Assuntos
Dioxigenases , Solanum lycopersicum , Dioxigenases/genética , Compostos Ferrosos , Regulação da Expressão Gênica de Plantas , Ácidos Cetoglutáricos , Família Multigênica , Filogenia , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Solanum lycopersicum/genética , Solanum lycopersicum/metabolismoRESUMO
During vertebrate gastrulation, canonical Wnt signaling induces the formation of neural plate border (NPB). Wnt is also thought to be required for the subsequent specification of neural crest (NC) lineage at the NPB, but the direct evidence is lacking. We found previously that the disintegrin metalloproteinase ADAM13 is required for Wnt activation and NC induction in Xenopus Here, we report that knockdown of ADAM13 or its close paralog ADAM19 severely downregulates Wnt activity at the NPB, inhibiting NC specification without affecting earlier NPB formation. Surprisingly, ADAM19 functions nonproteolytically in NC specification by interacting with ADAM13 and inhibiting its proteasomal degradation. Ectopic expression of stabilized ADAM13 mutants that function independently of ADAM19 can induce the NC marker/specifier snail2 in the future epidermis via Wnt signaling. These results unveil the essential roles of a novel protease-protease interaction in regulating a distinct wave of Wnt signaling, which directly specifies the NC lineage.
Assuntos
Proteínas ADAM/metabolismo , Padronização Corporal/fisiologia , Crista Neural/metabolismo , Proteínas Wnt/metabolismo , Proteínas de Xenopus/metabolismo , Xenopus/metabolismo , Animais , Regulação da Expressão Gênica no Desenvolvimento , Proteínas de Membrana/metabolismo , Crista Neural/embriologia , Placa Neural/metabolismo , Transdução de Sinais , Via de Sinalização Wnt/fisiologia , Xenopus/embriologiaRESUMO
Microtubules are polymers of tubulin dimers, and conformational transitions in the microtubule lattice drive microtubule dynamic instability and affect various aspects of microtubule function. The exact nature of these transitions and their modulation by anticancer drugs such as Taxol and epothilone, which can stabilize microtubules but also perturb their growth, are poorly understood. Here, we directly visualize the action of fluorescent Taxol and epothilone derivatives and show that microtubules can transition to a state that triggers cooperative drug binding to form regions with altered lattice conformation. Such regions emerge at growing microtubule ends that are in a pre-catastrophe state, and inhibit microtubule growth and shortening. Electron microscopy and in vitro dynamics data indicate that taxane accumulation zones represent incomplete tubes that can persist, incorporate tubulin dimers and repeatedly induce microtubule rescues. Thus, taxanes modulate the material properties of microtubules by converting destabilized growing microtubule ends into regions resistant to depolymerization.
Assuntos
Microtúbulos/efeitos dos fármacos , Microtúbulos/metabolismo , Taxoides/farmacologia , Células HeLa , Humanos , Cinética , Tubulina (Proteína)/metabolismoRESUMO
Traumatic brain injury (TBI), characterized by acute neurological dysfunction, is one of the best known environmental risk factors for chronic traumatic encephalopathy and Alzheimer's disease, the defining pathologic features of which include tauopathy made of phosphorylated tau protein (P-tau). However, tauopathy has not been detected in the early stages after TBI, and how TBI leads to tauopathy is unknown. Here we find robust cis P-tau pathology after TBI in humans and mice. After TBI in mice and stress in vitro, neurons acutely produce cis P-tau, which disrupts axonal microtubule networks and mitochondrial transport, spreads to other neurons, and leads to apoptosis. This process, which we term 'cistauosis', appears long before other tauopathy. Treating TBI mice with cis antibody blocks cistauosis, prevents tauopathy development and spread, and restores many TBI-related structural and functional sequelae. Thus, cis P-tau is a major early driver of disease after TBI and leads to tauopathy in chronic traumatic encephalopathy and Alzheimer's disease. The cis antibody may be further developed to detect and treat TBI, and prevent progressive neurodegeneration after injury.
Assuntos
Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/farmacologia , Lesões Encefálicas/patologia , Lesões Encefálicas/prevenção & controle , Tauopatias/prevenção & controle , Proteínas tau/antagonistas & inibidores , Proteínas tau/química , Doença de Alzheimer/complicações , Doença de Alzheimer/prevenção & controle , Animais , Anticorpos Monoclonais/uso terapêutico , Afinidade de Anticorpos , Axônios/metabolismo , Axônios/patologia , Encéfalo/metabolismo , Encéfalo/patologia , Lesões Encefálicas/complicações , Lesões Encefálicas/metabolismo , Modelos Animais de Doenças , Epitopos/química , Epitopos/imunologia , Feminino , Humanos , Masculino , Camundongos , Fosfoproteínas/antagonistas & inibidores , Fosfoproteínas/biossíntese , Fosfoproteínas/imunologia , Fosfoproteínas/toxicidade , Estresse Fisiológico , Tauopatias/complicações , Tauopatias/metabolismo , Tauopatias/patologia , Proteínas tau/biossíntese , Proteínas tau/imunologia , Proteínas tau/toxicidadeRESUMO
A completely non-blocking M×N electrically controlled optofluidic matrix switch that uses a 1×3 optical switch with a V-shaped microchannel as the switching unit is proposed. Its light paths and output ports are selected by a micro-actuator matrix and a control circuit. There are few reports of optofluidic matrix switches. Here the given electrostatic micro-actuator and the basic switch structure provide an effective feasible manner for the matrix switch due to the simple and compact structure as well as the operation style. The impacts of microchannels and intersecting waveguides on the switch performance are discussed, and multiple optimization schemes are proposed to reduce the insertion loss efficiently and significantly. The research results indicate that the M×N matrix switch has the advantages of good matrix controllability, simple structure, wide waveband (400-1700 nm), negligible polarization-dependent loss, small insertion loss, and low cross talk. For 1550 nm wavelength, the insertion loss of a 2×6 matrix switch is about 0.17-0.55 dB, and the maximum cross talk is less than -26.8dB. In addition, the performance parameters of a 4×8 matrix switch are given and compared with other reported matrix switches. The proposed M×N matrix switch solves the problem of large insertion loss of general optical matrix switches and can be expanded to a large-scale matrix switch. Moreover, the design of multiple output ports has more flexible applications in systems with multiple branch optical paths and network nodes.
RESUMO
Spinocerebellar ataxia (SCA) is a hereditary neurodegenerative disease. We have generated SCA17 transgenic mice bearing human TBP with 109 CAG repeats under the Purkinje cell-specific L7/pcp2 promoter. These mice recapitulate the patients' phenotypes and are suitable for the study of the SCA17 pathomechanism. Magnetic resonance imaging (MRI) and immunostainings were performed to identify the neuroimaging spectrum during disease progression. The results indicate that despite an overall normal appearance at birth, postnatal brain damage takes place rapidly in SCA17. Cerebellar atrophy, fourth-ventricle enlargement, and reduced cerebellar N-acetylaspartate levels were detected at the presymptomatic stage, when the mice were juvenile. The aberrations, which included reductions in body weight; cerebral size; striatal size; and the mean, radial, and axial diffusivities of the cerebellum, became more salient as the disease progressed to the old, late-symptomatic stage. Phosphorylated H2A histone family, member X (γH2AX) immunostaining revealed that the cerebellum underwent severe cell senescence in the old stage while the striatum appeared relatively unaffected by aging. Morphometric analysis indicated that the cerebellar atrophy occurred in all subregions with aging. The data establish that the SCA17 mouse brain appears normal at birth but becomes aberrant at the presymptomatic/juvenile stage. More widespread deficits add to the pathological spectrum at the old stage. The study provides information for the expression and expansion of L7/pcp2 promoter and implies the disease progression of SCA17 patients.
Assuntos
Encéfalo/patologia , Ataxias Espinocerebelares/patologia , Animais , Modelos Animais de Doenças , Progressão da Doença , Humanos , Imageamento por Ressonância Magnética , Camundongos , Camundongos Transgênicos , Neuroimagem/métodosRESUMO
Fbw7 is the substrate recognition component of the Skp1-Cullin-F-box (SCF)-type E3 ligase complex and a well-characterized tumor suppressor that targets numerous oncoproteins for destruction. Genomic deletion or mutation of FBW7 has been frequently found in various types of human cancers; however, little is known about the upstream signaling pathway(s) governing Fbw7 stability and cellular functions. Here we report that Fbw7 protein destruction and tumor suppressor function are negatively regulated by the prolyl isomerase Pin1. Pin1 interacts with Fbw7 in a phoshorylation-dependent manner and promotes Fbw7 self-ubiquitination and protein degradation by disrupting Fbw7 dimerization. Consequently, overexpressing Pin1 reduces Fbw7 abundance and suppresses Fbw7's ability to inhibit proliferation and transformation. By contrast, depletion of Pin1 in cancer cells leads to elevated Fbw7 expression, which subsequently reduces Mcl-1 abundance, sensitizing cancer cells to Taxol. Thus, Pin1-mediated inhibition of Fbw7 contributes to oncogenesis, and Pin1 may be a promising drug target for anticancer therapy.
Assuntos
Proteínas de Ciclo Celular/genética , Proteínas F-Box/genética , Regulação da Expressão Gênica , Peptidilprolil Isomerase/metabolismo , Ubiquitina-Proteína Ligases/genética , Sequência de Aminoácidos , Proteínas de Ciclo Celular/antagonistas & inibidores , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Tumoral , Proteínas F-Box/antagonistas & inibidores , Proteínas F-Box/metabolismo , Proteína 7 com Repetições F-Box-WD , Genes Supressores de Tumor , Humanos , Dados de Sequência Molecular , Peptidilprolil Isomerase de Interação com NIMA , Peptidilprolil Isomerase/genética , Ubiquitina-Proteína Ligases/antagonistas & inibidores , Ubiquitina-Proteína Ligases/metabolismo , UbiquitinaçãoRESUMO
PURPOSE: Rap2B, a member of the GTP-binding proteins, is generally up-regulated in numerous types of tumors. Nevertheless, the influence and regulatory mechanisms of Rap2B in gliomas are still not corroborated. Therefore, we analyzed the expression of Rap2B in glioma tissues and cells, and researched its significance in adhesion, proliferation, migration and invasion of the glioma cell line. METHODS: We analyzed the expression of Rap2B in different pathologic grades of glioma tissues by tissue microarray and immunohistochemistry. We assessed the expression of Rap2B in glioma tissue and non-tumor tissue by Western blot. And the expression of Rap2b protein in glioma cells and normal human astrocytes (NHA) was detected by Western blot. In addition, we disclosed the effect of Rap2B knockdown on cell adhesion, proliferation, migration and invasion by using cell attachment assay, CCK-8 assay, cell migration assay and Wound Healing assay, cell invasion assay, respectively. Western blot was used to detect the changes of expression level of NF-kB, MMP-2 and MMP-9 protein when downregulated the expression of Rap2B. RESULTS: The tissue microarray immunohistochemical results of glioma showed that the expression of Rap2B had no significant correlations between Rap2B expression and the clinicopathologic variables, including patient age (P = 0.352), gender (P = 0.858), WHO Grade (P = 0.693) and histology type (P = 0.877). Western blot analysis showed that the glioma tissue had a dramatically increase of Rap2B expression compared with the non-tumor tissues (P < 0.01). And the expression of Rap2B was markedly up-regulated in all 5 glioma cell lines compared with that in normal human astrocytes (NHA) (P < 0.01). We found that the ability of adhesion, proliferation, migration and invasion of glioma cells were significantly decreased after downregulated Rap2B expression compared with the control group (P < 0.05). In addition, Western blot results showed that the expression levels of NF-kB, MMP-2 and MMP-9 in the interference group were significantly lower than those in the negative control group (P < 0.05). CONCLUSIONS: Rap2B expression is up-regulated in glioma tissues and glioma cell lines. Knockdown of Rap2B inhibits glioma cells' adhesion and proliferation in vitro. Knockdown of Rap2B inhibits glioma cells' migration in vitro. Knockdown of Rap2B inhibits glioma cells' invasion and MMPs activity through NF-kB pathway.
Assuntos
Biomarcadores Tumorais/metabolismo , Adesão Celular , Movimento Celular , Proliferação de Células , Glioma/patologia , Proteínas rap de Ligação ao GTP/metabolismo , Apoptose , Biomarcadores Tumorais/genética , Feminino , Seguimentos , Regulação Neoplásica da Expressão Gênica , Glioma/genética , Glioma/metabolismo , Humanos , Masculino , Pessoa de Meia-Idade , NF-kappa B/genética , NF-kappa B/metabolismo , Invasividade Neoplásica , Prognóstico , RNA Interferente Pequeno/genética , Transdução de Sinais , Células Tumorais Cultivadas , Proteínas rap de Ligação ao GTP/antagonistas & inibidores , Proteínas rap de Ligação ao GTP/genéticaRESUMO
Two ß-secreatase (BACE1) inhibitors from natural products (cinnamic acid and flavone) were linked to furnish potent, cell permeable BACE1 inhibitors with noncompetitive mode of inhibition, with the assistance of saturated transfer difference (STD)-NMR technique. Some of these conjugates also exhibited selective BACE1 inhibition over other aspartyl proteases such as BACE-2 and renin, as well as poor cytotoxicity. Taken together, conjugates 4 represent a new series of BACE inhibitors warrants further investigation for their potential in Alzheimier's disease therapy.
Assuntos
Doença de Alzheimer/tratamento farmacológico , Secretases da Proteína Precursora do Amiloide/antagonistas & inibidores , Ácido Aspártico Endopeptidases/antagonistas & inibidores , Permeabilidade da Membrana Celular/efeitos dos fármacos , Descoberta de Drogas/métodos , Inibidores Enzimáticos/síntese química , Doença de Alzheimer/enzimologia , Sítios de Ligação , Ligação Competitiva , Descoberta de Drogas/instrumentação , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacocinética , Inibidores Enzimáticos/farmacologia , Humanos , Espectroscopia de Ressonância Magnética , Estrutura MolecularRESUMO
A series of novel 7,9-O-linked macrocyclic taxoids together with modification at the C2 position were synthesized, and their cytotoxicities against drug-sensitive and P-glycoprotein and ßIII-tubulin overexpressed drug-resistant cancer cell lines were evaluated. It is demonstrated that C-seco taxoids conformationally constrained via carbonate containing-linked macrocyclization display increased cytotoxicity on drug-resistant tumors overexpressing both ßIII and P-gp, among which compound 22b, bearing a 2-m-methoxybenzoyl group together with a five-atom linker, was identified as the most potent. Molecular modeling suggested the improved cytotoxicity of 22b results from enhanced favorable interactions with the T7 loop region of ßIII.
Assuntos
Compostos Macrocíclicos/síntese química , Compostos Macrocíclicos/farmacologia , Taxoides/síntese química , Taxoides/farmacologia , Hidrocarbonetos Aromáticos com Pontes/síntese química , Hidrocarbonetos Aromáticos com Pontes/química , Hidrocarbonetos Aromáticos com Pontes/farmacologia , Morte Celular/efeitos dos fármacos , Docetaxel/síntese química , Docetaxel/química , Docetaxel/farmacologia , Células HeLa , Humanos , Compostos Macrocíclicos/química , Simulação de Acoplamento Molecular , Paclitaxel/síntese química , Paclitaxel/química , Paclitaxel/farmacologia , Homologia Estrutural de Proteína , Taxoides/química , Tubulina (Proteína)/químicaRESUMO
BACE1 inhibitory conjugates derived from two natural products, luteolin (1) and p-hydroxy-cinnamic acid (2), were subjected to systematic structure modifications, including various positions in luteolin segment for conjugation, different linkers (length, bond variation), as well as various substitutions in cinnamic acid segment (various substituents on benzene, and replacement of benzene by heteroaromatics and cycloalkane). Optimal conjugates such as 7c and 7k were chosen on the basis of a series of bioassay data for further investigation.
Assuntos
Secretases da Proteína Precursora do Amiloide/antagonistas & inibidores , Ácido Aspártico Endopeptidases/antagonistas & inibidores , Cinamatos/química , Inibidores Enzimáticos/síntese química , Luteolina/química , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Humanos , Estrutura Molecular , Ligação Proteica , Relação Estrutura-AtividadeRESUMO
CdSe nanocrystals, including quantum dots and quantum rods, are a research hotspot in nanomaterials. Recently, it has been found that two-dimensional CdSe semiconductor nanobelts show a unique quantum confinement effect owing to their homogeneous nanoscale thickness. In this study, a series of (CdSe) n·monoamine ( n = 1, 2, and 4, monoamine = butylamine, hexylamine, and octylamine) layered inorganic/organic hybrid nanobelts was synthesized by the solvothermal method. These hybrids have a lamellar structure with a few inorganic [CdSe] layers and monoamine layers alternatively stacked along the c axis of their crystallites, which exhibit the typical preferred orientation for layered crystallites and isomorphous [CdSe] layers according to their X-ray diffraction patterns. The [CdSe] layer is a contracted (110) superlattice cell of wurtzite (WZ)-type CdSe with unit cell parameter relations of a ≈ â3 aWZ and b ≈ cWZ, and a lamellar nanobelt morphology with the [010] growth direction. These ordered hybrids exhibit prominent split exciton absorption peaks and sharp band edge luminescence owing to the homogeneous thickness of the [CdSe] layers within the hybrids, all of which are comparable with reported WZ-type CdSe colloidal nanosheets and exhibit a prominent two-dimensional exciton quantum confinement effect. The solvothermal reaction was systematically investigated. The multiple stage amine de-intercalation topotactic process was revealed for the WZ-type CdSe nanobelts with the (CdSe) n·monoamine ( n = 1, 2, and 4) hybrids as the intermediate phases. The solvothermal synthesis method (70-150 °C) is facile and milder than the traditional hot injection method (>200 °C). Therefore, these hybrids can be considered to be intermediate phases or precursors of WZ-type CdSe nanosheets with a topotactic intralayer [CdSe] structure and also a type of multiple quantum well with extremely thin [CdSe] layers orderly stacked among the monoamine insulating blocking layers.