RESUMO
The recruitment of RNA polymerase II (Pol II) to core promoters is highly regulated during rapid induction of genes. In response to heat shock, heat shock transcription factor 1 (HSF1) is activated and occupies heat shock gene promoters. Promoter-bound HSF1 recruits general transcription factors and Mediator, which interact with Pol II, but stress-specific mechanisms of Pol II recruitment are unclear. Here, we show in comparative analyses of HSF1 paralogs and their mutants that HSF1 interacts with the pericentromeric adaptor protein shugoshin 2 (SGO2) during heat shock in mouse cells, in a manner dependent on inducible phosphorylation of HSF1 at serine 326, and recruits SGO2 to the HSP70 promoter. SGO2-mediated binding and recruitment of Pol II with a hypophosphorylated C-terminal domain promote expression of HSP70, implicating SGO2 as one of the coactivators that facilitate Pol II recruitment by HSF1. Furthermore, the HSF1-SGO2 complex supports cell survival and maintenance of proteostasis in heat shock conditions. These results exemplify a proteotoxic stress-specific mechanism of Pol II recruitment, which is triggered by phosphorylation of HSF1 during the heat shock response.
Assuntos
Proteínas de Ciclo Celular/metabolismo , Fatores de Transcrição de Choque Térmico/metabolismo , Resposta ao Choque Térmico/fisiologia , RNA Polimerase II/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Proteínas de Ciclo Celular/genética , Regulação da Expressão Gênica , Técnicas de Silenciamento de Genes , Proteínas de Choque Térmico HSP70/genética , Proteínas de Choque Térmico HSP70/metabolismo , Resposta ao Choque Térmico/genética , Camundongos , Camundongos Knockout , Fosforilação , Ligação ProteicaRESUMO
Filamentous fungi synthesize natural products as an ecological function. In this study, an interesting indigenous fungus producing orange pigment exogenously was investigated in detail as it possesses additional attributes along with colouring properties. An interesting fungus was isolated from a dicot plant, Maytenus rothiana. After a detailed study, the fungal isolate turned out to be a species of Gonatophragmium belonging to the family Acrospermaceae. Based on the morphological, cultural, and sequence-based phylogenetic analysis, the identity of this fungus was confirmed as Gonatophragmium triuniae. Although this fungus grows moderately, it produces good amounts of pigment on an agar medium. The fermented crude extract isolated from G. triuniae has shown antioxidant activity with an IC50 value of 0.99 mg/mL and antibacterial activity against Gram-positive bacteria (with MIC of 3.91 µg/mL against Bacillus subtilis, and 15.6 µg/mL and 31.25 µg/mL for Staphylococcus aureus and Micrococcus luteus, respectively). Dyeing of cotton fabric mordanted with FeSO4 using crude pigment was found to be satisfactory based on visual observation, suggesting its possible use in the textile industry. The orange pigment was purified from the crude extract by preparative HP-TLC. In addition, UV-Vis, FTIR, HRMS and NMR (1H NMR, 13C NMR), COSY, and DEPT analyses revealed the orange pigment to be "1,2-dimethoxy-3H-phenoxazin-3-one" (C14H11NO4, m/z 257). To our understanding, the present study is the first comprehensive report on Gonatophragmium triuniae as a potential pigment producer, reporting "1,2-dimethoxy-3H-phenoxazin-3-one" as the main pigment from the crude hexane extract. Moreover, this is the first study reporting antioxidant, antibacterial, and dyeing potential of crude extract of G. triuniae, suggesting possible potential applications of pigments and other bioactive secondary metabolites of the G. triuniae in textile and pharmaceutical industry.
Assuntos
AscomicetosRESUMO
Epithelial-mesenchymal transition (EMT) is a programed course of developmental changes resulting in the acquisition of invasiveness and mobility in cells. In cancer, this course is used by epithelial cells to attain movability. Translationally controlled tumor protein (TCTP) has been extensively characterized following the observation on tumor reversion ensuing its depletion. However, the role of TCTP in cancer progression is still elusive. Here, we demonstrate for the first time that TCTP is a target of transforming growth factor-ß1 (TGF-ß1), a key regulator of EMT in A549 cells. We here present changes in expression patterns of intermediate filament markers (vimentin and cytokeratin 18a) of EMT following TCTP knockdown or over expression. The TCTP over-expression in cancer cells is associated with mesenchymal characters, while downregulation promotes the epithelial markers in the cells. Interaction of TCTP with ß-catenin seems to stabilize ß-catenin, preparative to its nuclear localization highlighting a role for ß-catenin signaling in EMT. Moreover, the induction of urokinase plasminogen activator (uPA) following ectopic expression of TCTP leads to destabilization of ECM. The cells knocked down for TCTP show diminished invasiveness and migration under TGF-ß1 treatment. The present results for the first time demonstrate that TGF-ß1 dependent TCTP expression is required for EMT in cells.
Assuntos
Biomarcadores Tumorais/fisiologia , Citoesqueleto/metabolismo , Transição Epitelial-Mesenquimal , Fator de Crescimento Transformador beta1/farmacologia , Células A549 , Biomarcadores Tumorais/genética , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Movimento Celular/genética , Citoesqueleto/efeitos dos fármacos , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Transição Epitelial-Mesenquimal/genética , Células HEK293 , Humanos , Células MCF-7 , Metástase Neoplásica , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Fator de Crescimento Transformador beta1/metabolismo , Proteína Tumoral 1 Controlada por TraduçãoRESUMO
Chikungunya virus (CHIKV) and dengue virus (DENV) are important arboviruses transmitted by Aedes mosquitoes. These viruses are known to coexist within the same vector and coinfect the same host. Although information is available on the mechanism of replication of CHIKV and DENV when present independently in a vector, reports are lacking on the dynamics of virus-vector interactions when these viruses coexist in a mosquito. The current study attempts to understand the perturbations in the proteome of Aedes mosquitoes when infected with CHIKV and DENV either independently or together. Global proteome profiling of chikungunya and dengue mono- and coinfection revealed 28 proteins to be significantly regulated. Validation of the transcripts of these proteins using qRT-PCR indicated differences in the expression patterns between transcript profiling and quantitative proteome analyses. Pathway analysis of the 28 differentially regulated proteins revealed 11 significant pathways, which include oxidative phosphorylation, carbon metabolism, and glycolysis/gluconeogenesis.
Assuntos
Aedes/metabolismo , Coinfecção/metabolismo , Proteínas de Insetos/metabolismo , Mosquitos Vetores/metabolismo , Proteoma/metabolismo , Infecções por Vírus de RNA/metabolismo , Aedes/genética , Aedes/virologia , Animais , Vírus Chikungunya/fisiologia , Coinfecção/genética , Coinfecção/virologia , Vírus da Dengue/fisiologia , Perfilação da Expressão Gênica/métodos , Interações Hospedeiro-Patógeno , Proteínas de Insetos/genética , Mosquitos Vetores/genética , Mosquitos Vetores/virologia , Proteoma/genética , Proteômica/métodos , Infecções por Vírus de RNA/genética , Infecções por Vírus de RNA/virologiaRESUMO
Chikungunya virus (CHIKV) and Dengue virus (DENV) spread via the bite of infected Aedes mosquitoes. Both these viruses exist as co-infections in the host as well as the vector and are known to exploit their cellular machinery for their replication. While there are studies reporting the changes in Aedes transcriptome when infected with DENV and CHIKV individually, the effect both these viruses have on the mosquitoes when present as co-infections is not clearly understood. In the present study, we infected Aedes aegypti mosquitoes with DENV and CHIKV individually and as co-infection through nanoinjections. We performed high throughput RNA sequencing of the infected Aedes aegypti to understand the changes in the Aedes transcriptome during the early stages of infection, i.e., 24 h post infection and compared the transcriptome profiles during DENV and CHIKV mono-infections with that of co-infections. We identified 190 significantly regulated genes identified in CHIKV infected library, 37 genes from DENV library and 100 genes from co-infected library and they were classified into different pathways. Our study reveal that distinct pathways and transcripts are being regulated during the three types of infection states in Aedes aegypti mosquitoes.
Assuntos
Aedes/metabolismo , Aedes/virologia , Febre de Chikungunya/metabolismo , Vírus Chikungunya/fisiologia , Vírus da Dengue/fisiologia , Dengue/metabolismo , Transcriptoma , Animais , Febre de Chikungunya/virologia , Coinfecção/metabolismo , Coinfecção/virologia , Dengue/virologia , Perfilação da Expressão Gênica , Insetos Vetores/metabolismo , Insetos Vetores/virologiaRESUMO
Upon heat shock, activated heat shock transcription factor 1 (HSF1) binds to the heat shock response elements (HSEs) in the promoters of mammalian heat shock protein (HSP)-encoding genes and recruits the preinitiation complex and coactivators, including Mediator. These transcriptional regulators may be concentrated in phase-separated condensates around the promoters, but they are too minute to be characterized in detail. We herein established HSF1-/- mouse embryonic fibroblasts harbouring HSP72-derived multiple HSE arrays and visualized the condensates of fluorescent protein-tagged HSF1 with liquid-like properties upon heat shock. Using this experimental system, we demonstrate that endogenous MED12, a subunit of Mediator, is concentrated in artificial HSF1 condensates upon heat shock. Furthermore, the knockdown of MED12 markedly reduces the size of condensates, suggesting an important role for MED12 in HSF1 condensate formation.
Assuntos
Proteínas de Ligação a DNA , Fibroblastos , Animais , Camundongos , Fatores de Transcrição de Choque Térmico/genética , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Fibroblastos/metabolismo , Fatores de Transcrição/metabolismo , Resposta ao Choque Térmico/genética , Mamíferos/metabolismoRESUMO
Soybean rust (SBR), caused by Phakopsora pachyrhizi, is a damaging fungal disease of soybean (Glycine max). Although solar radiation can reduce SBR urediniospore survival, limited information is available on how solar radiation affects SBR progress within soybean canopies. Such information can aid in developing accurate SBR prediction models. To manipulate light penetration into soybean canopies, structures of shade cloth attenuating 30, 40, and 60% sunlight were constructed over soybean plots. In each plot, weekly evaluations of severity in lower, middle, and upper canopies, and daily temperature and relative humidity were recorded. Final plant height and leaf area index were also recorded for each plot. The correlation between amount of epicuticular wax and susceptibility of leaves in the lower, middle, and upper canopies was assessed with a detached leaf assay. Final disease severity was 46 to 150% greater in the lower canopy of all plots and in the middle canopy of 40 and 60% shaded plots. While daytime temperature within the canopy of nonshaded soybean was greater than shaded soybean by 2 to 3°C, temperatures recorded throughout typical evenings and mornings of the growing season in all treatments were within the range (10 to 28.5°C) for SBR development as was relative humidity. This indicates temperature and relative humidity were not limiting factors in this experiment. Epicuticular wax and disease severity in detached leaf assays from the upper canopy had significant negative correlation (P = 0.009, R = -0.84) regardless of shade treatment. In laboratory experiments, increasing simulated total solar radiation (UVA, UVB, and PAR) from 0.15 to 11.66 MJ m(-2) increased mortality of urediniospores from 2 to 91%. Variability in disease development across canopy heights in early planted soybean may be attributed to the effects of solar radiation not only on urediniospore viability, but also on plant height, leaf area index, and epicuticular wax, which influence disease development of SBR. These results provide an understanding of the effect solar radiation has on the progression of SBR within the soybean canopy.
Assuntos
Glycine max/microbiologia , Doenças das Plantas/microbiologia , Luz Solar , Glycine max/efeitos da radiação , TemperaturaRESUMO
INTRODUCTION: Bacopa monnieri (BM) contains several dammarane-type triterpenoid saponins including bacopaside I and bacoside A. These bioactive compounds may be used as chemical markers for the quality control of different BM products used for promoting mental health and intellect. OBJECTIVE: Quantification of bacopaside I and bacoside A in crude plant material of BM stored under the stability study conditions by HPLC. METHODOLOGY: Crude BM samples were stored at long-term (LS; 30°C and 65% RH), accelerated (AS; 40°C and 75% RH) and real-time (RT) study conditions. HPLC of BM extracts was carried out using a LiChroCART Purospher® STAR RP-18 endcapped column along with a guard column, Purospher STAR RP 18e 4.0 4.0 mm 5 µm using a gradient of acetonitrile (A) and water containing 0.05% (v/v) orthophosphoric acid (B) at a flow rate 1.5 mL/min with UV detection at 205 nm. RESULTS: The linear range of bacopaside I and bacoside A was 0.2 to 1 mg/mL. With the help of a regression equation the coefficient of determination (r²) values for bacopaside I and bacoside A were found to be > 0.999 and > 0.994 respectively. Relative standard deviation (RSD) values were < 4.0 for all the concentrations injected (n = 3). The HPLC study indicated that BM samples kept under LS condition are rich in saponin contents as compared with the samples stored under AS and RT study conditions. CONCLUSION: The study indicated that BM plant material should be used fresh to obtain maximum concentration of active saponins or it should be stored under LS conditions up to 3 months.
Assuntos
Bacopa/química , Cromatografia Líquida de Alta Pressão/métodos , Extratos Vegetais/química , Saponinas/isolamento & purificação , Triterpenos/isolamento & purificação , Biomarcadores/química , Armazenamento de Medicamentos , Modelos Lineares , Saponinas/química , Temperatura , Triterpenos/química , Raios UltravioletaRESUMO
The first synthetic route of naturally occurring (±)-5,7-dihydroxy-8-methyl-3-(2',4'-dihydroxybenzyl)chroman-4-one (1) from Gan Luo Xin pill was successfully accomplished. The synthetic route has been developed retro-synthetically in 9 simple steps with a high yield of â¼80%. The synthetic protocol was developed using readily available starting material phloroglucinol. The key intermediate 2,4,6-trihydroxy-3-methyl acetophenone (4) was synthesized via Vilsmeier-Haack reaction, followed by reduction using sodium cyanoborohydride and acylation reaction. LC-MS, IR, 1H NMR, 13C NMR of 1 have been analyzed to confirm the structure of (±)-5,7-dihydroxy-8-methyl-3-(2',4'-dihydroxybenzyl)chroman-4-one (1) and found in agreement with the natural molecule. The target compound showed 97% and 87% antioxidant activity in DPPH and ABTS assay at 1 mg/ml concentration, respectively. The compound (1) also showed ferric ion reducing activity with the absorbance of 0.18 at 700 nm. The present study could be useful in developing synthetic routes of other potential naturally occurring homoisoflavonoid.
Assuntos
Produtos Biológicos , Medicina Tradicional do Leste Asiático , Antioxidantes/farmacologiaRESUMO
The first synthetic route developed for Podocarflavone A reported from Podocarpus macrophyllus and its analogs in 7 steps. Computational analysis for binding with the pantothenate kinase (3AVO) of Mycobacterium tuberculosis showed their docking score (ds) in the range of -8.9 to -9.3 Kcal/mol. MD simulations delineated the stability of the protein-ligand complexes in the TIP3P model. MMGBSA and MMPBSA values of 8d were -42.46 Kcal/mol and -14.58 Kcal/mol, respectively. Further in-vitro antitubercular screening of compounds 8a, 8d, and 8e against M. tuberculosis H37Ra using XRMA protocol exhibited promising antimycobacterial activity with IC50 values 21.82 µg/mL, 15.55 µg/mL, and 16.56 µg/mL, respectively. Compounds 8a, 8d, and 8e showed antibacterial activity with IC50 values 41.56 µg/mL, 24.72 µg/mL, and 72.45 µg/mL respectively against the Staphylococcus aureus. 8a and 8d showed inhibition with IC50 values 39.6 µg/mL and 27.64 µg/mL, respectively, against Bacillus subtilis. The present study could help in the further development of lead molecules against tuberculosis.
Assuntos
Mycobacterium tuberculosis , Tuberculose , Antituberculosos/química , Flavonoides , Humanos , Testes de Sensibilidade Microbiana , Simulação de Acoplamento Molecular , Relação Estrutura-Atividade , Tuberculose/microbiologiaRESUMO
Activated and promoter-bound heat-shock transcription factor 1 (HSF1) induces RNA polymerase II recruitment upon heat shock, and this is facilitated by the core Mediator in Drosophila and yeast. Another Mediator module, CDK8 kinase module (CKM), consisting of four subunits including MED12 and CDK8, plays a negative or positive role in the regulation of transcription; however, its involvement in HSF1-mediated transcription remains unclear. We herein demonstrated that HSF1 interacted with MED12 and recruited MED12 and CDK8 to the HSP70 promoter during heat shock in mammalian cells. The kinase activity of CDK8 (and its paralog CDK19) promoted HSP70 expression partly by phosphorylating HSF1-S326 and maintained proteostasis capacity. These results indicate an important role for CKM in the protection of cells against proteotoxic stress.
Assuntos
Quinase 8 Dependente de Ciclina/genética , Fatores de Transcrição de Choque Térmico/genética , Resposta ao Choque Térmico/genética , Complexo Mediador/genética , Complexos Multiproteicos/genética , Proteostase/genética , Animais , Quinase 8 Dependente de Ciclina/metabolismo , Quinases Ciclina-Dependentes/genética , Quinases Ciclina-Dependentes/metabolismo , Fibroblastos , Regulação da Expressão Gênica , Células HEK293 , Proteínas de Choque Térmico HSP70/genética , Proteínas de Choque Térmico HSP70/metabolismo , Células HeLa , Fatores de Transcrição de Choque Térmico/metabolismo , Humanos , Complexo Mediador/metabolismo , Camundongos , Complexos Multiproteicos/metabolismo , Neurônios , Osteoblastos , Fosforilação , Ligação Proteica , Transdução de Sinais , Transcrição GênicaRESUMO
Boll rots of cotton (Gossypium hirsutum L.) are common in the humid areas of the Southeastern US. One type of boll damage that may be differentiated from others is hardlock, with symptoms that include compression of the fibers within individual locules of mature, open cotton bolls without further obvious disintegration of the lint or damage to the carpel wall. The principal economic effect is that the boll's lint is unharvestable by mechanical cotton pickers. This disease is endemic to the Southeast and can cause severe yield losses up to 70% in some fields. Scanning electron microscopy images of fibers from hardlocked bolls showed flattened and twisted tissue compared to fibers from healthy bolls. Fusarium verticillioides (Saccardo) Nirenberg was the fungus most commonly isolated from seeds of developing cotton bolls. Flowers inoculated with F. verticillioides on the day of bloom by spraying a spore suspension onto the flowers developed significantly (P < 0.05) more hardlock symptoms compared to untreated controls. The infection process was analyzed using a F. verticillioides isolate tagged with green fluorescent protein (GFP). When it was applied to cotton flowers on the day of bloom, the GFP-tagged F. verticillioides strain was detected in the stigma and style by 2 days after bloom (DAB) and in developing seeds at 4, 6, 8, 10, 16, 20, 40, and 60 (open bolls) DAB. By 8 DAB, the GFP F. verticillioides was isolated from over 80% of developing seeds.
Assuntos
Fusarium/isolamento & purificação , Gossypium/microbiologia , Doenças das Plantas/microbiologia , Fusarium/classificação , Genes Reporter , Gossypium/ultraestrutura , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Microscopia Eletrônica de Varredura , Micologia/métodos , Sementes/microbiologia , Sudeste dos Estados Unidos , Coloração e Rotulagem/métodosRESUMO
The mitochondrial unfolded protein response (UPRmt ) is characterized by the transcriptional induction of mitochondrial chaperone and protease genes in response to impaired mitochondrial proteostasis and is regulated by ATF5 and CHOP in mammalian cells. However, the detailed mechanisms underlying the UPRmt are currently unclear. Here, we show that HSF1 is required for activation of mitochondrial chaperone genes, including HSP60, HSP10, and mtHSP70, in mouse embryonic fibroblasts during inhibition of matrix chaperone TRAP1, protease Lon, or electron transfer complex 1 activity. HSF1 bound constitutively to mitochondrial chaperone gene promoters, and we observed that its occupancy was remarkably enhanced at different levels during the UPRmt . Furthermore, HSF1 supported the maintenance of mitochondrial function under the same conditions. These results demonstrate that HSF1 is required for induction of mitochondrial chaperones during the UPRmt , and thus, it may be one of the guardians of mitochondrial function under conditions of impaired mitochondrial proteostasis.
Assuntos
Fatores de Transcrição de Choque Térmico/metabolismo , Mitocôndrias/metabolismo , Chaperonas Moleculares/genética , Resposta a Proteínas não Dobradas/genética , Animais , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Fibroblastos , Técnicas de Silenciamento de Genes , Células HEK293 , Células HeLa , Fatores de Transcrição de Choque Térmico/genética , Humanos , Potencial da Membrana Mitocondrial/genética , Camundongos , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo , Fosforilação , Regiões Promotoras Genéticas/genética , Interferência de RNA , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismoRESUMO
Most estrogen receptor α (ERα) ligands target the ligand binding domain (LBD). Agonist 17ß-estradiol (E2) and tamoxifen (TM, known SERM), bind to the same site within the LBD. However, structures of ligand-bound complexes show that E2 and TM induce different conformations of helix 12 (H12). During the molecular modelling studies of some naturally occurring flavonoids such as quercetin, luteolin, myricetin, kaempferol, naringin, hesperidin, galangin, baicalein and epicatechin with human ERα (3ERT and 1GWR), we observed that most of the ligands bound to the active site pocket of both 3ERT and 1GWR. The docking scores, interaction analyses, and conformation of H12 provided the data to support for the estrogenic or antiestrogenic potential of these flavonoids to a limited degree. Explicit molecular dynamics for 50 ns was performed to identify the stability and compatibility pattern of protein-ligand complex and RMSD were obtained. Baicalein, epicatechin, and kaempferol with 1GWR complex showed similar RMSD trend with minor deviations in the protein backbone RMSD against 1GWR-E2 complex that provided clear indications that ligands were stable throughout the explicit molecular simulations in the protein and outcome of naringin-3ERT complex had an upward trend but stable throughout the simulations and all molecular dynamics showed stability with less than overall 1 Å deviation throughout the simulations. To examine their estrogenic or antiestrogenic potential, we studied the effect of the flavonoids on viability, progesterone receptor expression and 3xERE/3XERRE-driven reporter gene expression in ERα positive and estrogen responsive MCF-7 breast cancer cells. Epicatechin, myricetin, and kaempferol showed estrogenic potential at 5 µM concentration.
Assuntos
Receptor alfa de Estrogênio/metabolismo , Flavonoides/farmacologia , Receptores de Estrogênio/ultraestrutura , Sítios de Ligação , Avaliação Pré-Clínica de Medicamentos/métodos , Estradiol/análogos & derivados , Estradiol/farmacologia , Antagonistas de Estrogênios/metabolismo , Antagonistas de Estrogênios/farmacologia , Receptor alfa de Estrogênio/agonistas , Receptor alfa de Estrogênio/antagonistas & inibidores , Estrogênios/metabolismo , Flavonoides/química , Humanos , Ligantes , Conformação Molecular , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , Ligação Proteica , Receptores de Estrogênio/metabolismo , Tamoxifeno/química , Tamoxifeno/farmacologiaRESUMO
Antiviral therapy is crucial for the circumvention of viral epidemics. The unavailability of a specific antiviral drug against the chikungunya virus (CHIKV) disease has created an alarming situation to identify or develop potent chemical molecules for remedial management of CHIKV. In the present investigation, in silico studies of dihydrorugosaflavonoid derivatives (5a-f) with non-structural protein-3 (nsP3) were carried out. nsP3 replication protein has recently been considered as a possible antiviral target in which crucial inhibitors fit into the adenosine-binding pocket of the macrodomain. The 4'-halogenated dihydrorugosaflavonoids displayed intrinsic binding with the nsp3 macrodomain (PDB ID: 3GPO) of CHIKV. Compounds 5c and 5d showed docking scores of -7.54 and -6.86 kcal mol-1, respectively. Various in vitro assays were performed to confirm their (5a-f) antiviral potential against CHIKV. The non-cytotoxic dose was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and was found to be <100 µM. The compounds 5c and 5d showed their inhibitory potential for CHIKV, which was determined through cytopathic effect assay and plaque reduction assay, which show inhibition up to 95 and 92% for 70 µM concentration of the compounds, respectively. The quantitative real-time polymerase chain reaction assay result confirmed the ability of 5c and 5d to reduce the viral RNA level at 70 µM concentration of compounds to nearly 95 and 93% concentration, respectively, in cells with CHIKV infection. Further, the CHIKV-inhibitory capacity of these compounds was corroborated by execution of immunofluorescence assay. The executed work will be meaningful for the future research of studied dihydrorugosaflavonoids against prime antiviral entrants, leading to remedial management to preclude CHIKV infection.
RESUMO
Four phenylenediammonium dinitrate salts were prepared and characterized by elemental, Infrared spectroscopy (IR), Ultraviolet spectroscopy (UV) and gravimetric methods. These dinitrates find application in propellant, explosives and pyrotechnics. Their thermal decomposition has been studied using thermogravimetry (TG) and simultaneous thermogravimetry-differential scanning calorimetry (TG-DSC). Kinetics parameters were evaluated by model fitting and isoconversional methods. Their thermolytic pathways have also been suggested, which involves decomposition followed by ignition.
Assuntos
Substâncias Explosivas/química , Nitratos/química , Compostos de Amônio Quaternário/química , Temperatura Alta , Cinética , Modelos QuímicosRESUMO
Novel drug regimens against tuberculosis (TB) are urgently needed and may be developed by targeting essential enzymes of Mtb that sustain the pathogenicity of tuberculosis. In the present investigation, series of compounds (5a-f and 6a-f) based on a naturally occurring rugosaflavonoid moiety were evaluated by in silico molecular modeling studies against ß-ketoacyl-ACP reductase (MabA) (PDB ID: IUZN) and pantothenate kinase (PanK) (PDB ID: 3AF3). Compounds 5a, 5c, 5d, and 6c, which had docking scores of -8.29, -8.36, -8.17 and -7.39 kcal mol-1, respectively, displayed interactions with MabA that were better than those of isoniazid (-6.81 kcal mol-1). Similarly, compounds 5a, 5c, 5d, and 6c, which had docking scores of -7.55, -7.64, -7.40 and -6.7 kcal mol-1, respectively, displayed interactions with PanK that were comparable to those of isoniazid (-7.64 kcal mol-1). Because of their docking scores, these compounds were screened in vitro against Mycobacterium tuberculosis H37Ra (Mtb) using an XRMA protocol. Among the screened compounds, the dihydrorugosaflavonoid derivatives 5a, 5c, and 5d had IC50 values of 12.93, 8.43 and 11.3 µg mL-1, respectively, and exhibited better inhibitory activity than the parent rugosaflavonoid derivatives. The rugosaflavonoid derivative 6c had an IC50 value of 17.57 µg mL-1. The synthesized compounds also displayed inhibitory activity against the Gram-positive bacteria Bacillus subtilis and Staphylococcus aureus. The present study will be helpful for the further development of these molecules into antitubercular lead candidates.
RESUMO
The heat shock response (HSR) is characterized by the rapid and robust induction of heat shock proteins (HSPs), including HSP70, in response to heat shock and is regulated by heat shock transcription factor 1 (HSF1) in mammalian cells. Poly(ADP-ribose) polymerase 1 (PARP1), which can form a complex with HSF1 through the scaffold protein PARP13, has been suggested to be involved in the HSR. However, its effects on and the regulatory mechanisms of the HSR are not well understood. Here we show that prior to heat shock, the HSF1-PARP13-PARP1 complex binds to the HSP70 promoter. In response to heat shock, activated and auto-PARylated PARP1 dissociates from HSF1-PARP13 and is redistributed throughout the HSP70 locus. Remarkably, chromatin in the HSP70 promoter is initially PARylated at high levels and decondensed, whereas chromatin in the gene body is moderately PARylated afterwards. Activated HSF1 then binds to the promoter efficiently and promotes the HSR. Chromatin PARylation and HSF1 binding to the promoter are also facilitated by the phosphorylation-dependent dissociation of PARP13. Furthermore, the HSR and proteostasis capacity are reduced by pretreatment with genotoxic stresses, which disrupt the ternary complex. These results illuminate one of the priming mechanisms of the HSR that facilitates the binding of HSF1 to DNA during heat shock.
Assuntos
DNA/metabolismo , Fatores de Transcrição de Choque Térmico/metabolismo , Resposta ao Choque Térmico/fisiologia , Poli(ADP-Ribose) Polimerase-1/metabolismo , Animais , Linhagem Celular , Cromatina/genética , Cromatina/metabolismo , DNA/genética , Dano ao DNA , Técnicas de Silenciamento de Genes , Proteínas de Choque Térmico HSP70/genética , Proteínas de Choque Térmico HSP70/metabolismo , Resposta ao Choque Térmico/genética , Humanos , Camundongos , Modelos Biológicos , Poli(ADP-Ribose) Polimerase-1/antagonistas & inibidores , Poli(ADP-Ribose) Polimerase-1/genética , Poli(ADP-Ribose) Polimerases/deficiência , Poli(ADP-Ribose) Polimerases/genética , Poli(ADP-Ribose) Polimerases/metabolismo , Regiões Promotoras Genéticas , Ligação Proteica , Proteostase , RNA Mensageiro/genética , RNA Mensageiro/metabolismoRESUMO
The leaf extract of Azadirachta indica (Neem) plant was utilized as reducing agent for the green synthesis of Mn3O4 nanoparticles (NPs). The crystalline analysis demonstrated the typical tetragonal hausmannite crystal structure of Mn3O4, which confirmed the formation of Mn3O4 NPs without the existence of other oxides. Green synthesized Mn3O4 NPs were applied for the catalytic thermal decomposition of ammonium perchlorate (AP) and as working electrode for fabricating the chemical sensor. The excellent catalytic effect for the thermal decomposition of AP was observed by decreasing the decomposition temperature by 175 °C with single decomposing step. The fabricated chemical sensor based on green synthesized Mn3O4 NPs displayed high, reliable and reproducible sensitivity of â¼569.2 µA mM(-1) cm(-2) with reasonable limit of detection (LOD) of â¼22.1 µM and the response time of â¼10 s toward the detection of 2-butanone chemical. A relatively good linearity in the ranging from â¼20 to 160 µM was detected for Mn3O4 NPs electrode based 2-butanone chemical sensor.
Assuntos
Azadirachta/química , Química Verde , Compostos de Manganês/química , Nanopartículas/química , Óxidos/química , Extratos Vegetais/química , Substâncias Redutoras/química , Butanonas/análise , Catálise , Técnicas Eletroquímicas , Temperatura Alta , Nanopartículas/ultraestrutura , Percloratos/química , Folhas de Planta/química , Compostos de Amônio Quaternário/químicaRESUMO
The HIV-1 nucleocapsid protein (NCp7) is a small basic protein with two CysCysHisCys zinc-binding domains that specifically recognizes the Psi-site of the viral RNA. NCp7 plays a number of crucial roles in the viral lifecycle, including reverse transcription and RNA encapsidation. Several classes of potential anti-HIV compounds have been designed to inactivate NCp7 through zinc ejection, including a special class of thioester compounds. We have investigated the mechanism of action of two N-substituted-S-acyl-2-mercaptobenzamide compounds (compounds 1 and 2) that target NCp7. UV/Visible spectroscopy studies demonstrated that both thioesters were able to eject metal from NCp7. NMR and mass spectroscopy studies showed that the thioester compounds specifically ejected zinc from the carboxyl-terminal zinc-binding domain of NCp7 by covalent modification of Cys(39). Exposure of NCp7 to compounds 1 and 2 destroyed its ability to specifically bind RNA, whereas NCp7 already bound to RNA was protected from zinc ejection by the thioesters. The thiol component of the thioesters (compound 3, 2-mercaptobenzoyl-beta-alaninamide) did not eject zinc from NCp7, but when compound 3 was incubated with acetyl CoA prior to incubation with NCp7, we observed extensive metal ejection. Thus, the thiol released by the reaction of compounds 1 and 2 could be re-acylated in vivo by acyl CoA to form a new thioester compound that is able to react with NCp7. These studies provide a better understanding of the mechanism of action of thioester compounds, which is important for future design of anti-HIV-1 compounds that target NCp7.