RESUMEN
Nowadays, thermally activated delayed fluorescence (TADF) compounds with a fused-ring core skeleton are getting increasing research interest because of their use in high-performance organic light-emitting diodes (OLEDs). In this study, TADF compounds featuring a D-A-type fused-ring core skeleton are developed. The challenging compatibility of a planarized D-A arrangement and the TADF property is achieved through linking the D and A moieties with two oxygen atoms within a six-membered ring. Compared with a single-oxygen analogue possessing a flexible skeleton and a twisted D-A arrangement, these fused-ring compounds with higher skeleton rigidity show higher photoluminescence quantum yields and narrower emission spectra in toluene and in doped thin films. Their electroluminescent devices achieve high external quantum efficiencies (up to 19.4%), suggesting the potential of rarely achieved D-A-type fused-ring TADF systems to serve as high-performance emitters of OLEDs.
RESUMEN
Reported herein is a molecular design strategy of deep-blue emitters for resolving the lack of highly efficient deep-blue organic light-emitting diodes (OLEDs) featuring CIEy (Commission Internationale de l'Eclairage) color coordinates matching the display requirements (<0.1). The strategy is to combine weak spiro-donor and spiro-acceptor groups into a linear donor-π-acceptor type of thermally-activated delayed fluorescence molecule through a sterically bulky π-spacer. The strategy endows an emitter with deep-blue emission, a narrower emission bandwidth (51â nm in toluene), a high photoluminescence quantum yield (0.95 in toluene), weak concentration quenching, and efficient triplet-exciton utilization, which are all attractive characteristics for emitters of deep-blue OLEDs with lower CIEy coordinates. Owing to the rational design, the emitter has realized not only highly efficient doped deep-blue OLEDs with external quantum efficiencies (EQEs) up to 25.4 % and CIEy less than 0.1 but also so far the most efficient nondoped deep-blue OLED (EQE up to 22.5 %) with CIEy less than 0.1.
RESUMEN
BACKGROUND: Epstein-Barr virus (EBV)-driven B cell proliferation is critical to its subsequent persistence in the host and is a key event in the development of EBV-associated B cell diseases. Thus, inquiry into early cellular events that precede EBV-driven proliferation of B cells is essential for understanding the processes that can lead to EBV-associated B cell diseases. METHODS: Infection with high titers of EBV of mixed, primary B cells in different stages of differentiation occurs during primary EBV infection and in the setting of T cell-immunocompromise that predisposes to development of EBV-lymphoproliferative diseases. Using an ex vivo system that recapitulates these conditions of infection, we correlated expression of selected B cell-surface markers and intracellular cytokines with expression of EBV latency genes and cell proliferation. RESULTS: We identified CD23, CD58, and IL6, as molecules expressed at early times after EBV-infection. EBV differentially infected B cells into two distinct sub-populations of latently infected CD23+ cells: one fraction, marked as CD23hiCD58+IL6- by day 3, subsequently proliferated; another fraction, marked as CD23loCD58+, expressed IL6, a B cell growth factor, but failed to proliferate. High levels of LMP1, a critical viral oncoprotein, were expressed in individual CD23hiCD58+ and CD23loCD58+ cells, demonstrating that reduced levels of LMP1 did not explain the lack of proliferation of CD23loCD58+ cells. Differentiation stage of B cells did not appear to govern this dichotomy in outcome either. Memory or naïve B cells did not exclusively give rise to either CD23hi or IL6-expressing cells; rather memory B cells gave rise to both sub-populations of cells. CONCLUSIONS: B cells are differentially susceptible to EBV-mediated proliferation despite expression of viral gene products known to be critical for continuous B cell growth. Cellular events, in addition to viral gene expression, likely play a critical role in determining the outcome of EBV infection. By indentifying cells predicted to undergo EBV-mediated proliferation, our study provides new avenues of investigation into EBV pathogenesis.
Asunto(s)
Linfocitos B/citología , Proliferación Celular , Infecciones por Virus de Epstein-Barr/fisiopatología , Herpesvirus Humano 4/fisiología , Linfocitos B/inmunología , Linfocitos B/virología , Células Cultivadas , Infecciones por Virus de Epstein-Barr/genética , Infecciones por Virus de Epstein-Barr/inmunología , Infecciones por Virus de Epstein-Barr/virología , Femenino , Expresión Génica , Herpesvirus Humano 4/aislamiento & purificación , Humanos , Interleucina-6/genética , Interleucina-6/inmunología , Masculino , Receptores de IgE/genética , Receptores de IgE/inmunologíaRESUMEN
Diverse stimuli reactivate the Epstein-Barr virus (EBV) lytic cycle in Burkitt lymphoma (BL) cells. In HH514-16 BL cells, two histone deacetylase (HDAC) inhibitors, sodium butyrate (NaB) and trichostatin A (TSA), and the DNA methyltransferase inhibitor azacytidine (AzaCdR) promote lytic reactivation. Valproic acid (VPA), which, like NaB, belongs to the short-chain fatty acid class of HDAC inhibitors, fails to induce the EBV lytic cycle in these cells. Nonetheless, VPA behaves as an HDAC inhibitor; it causes hyperacetylation of histone H3 (J. K. Countryman, L. Gradoville, and G. Miller, J. Virol. 82:4706-4719, 2008). Here we show that VPA blocked the induction of EBV early lytic proteins ZEBRA and EA-D in response to NaB, TSA, or AzaCdR. The block in lytic activation occurred prior to the accumulation of BZLF1 transcripts. Reactivation of EBV in Akata cells, in response to anti-IgG, and in Raji cells, in response to tetradecanoyl phorbol acetate (TPA), was also inhibited by VPA. MS-275 and apicidin, representing two additional classes of HDAC inhibitors, and suberoylanilide hydroxamic acid (SAHA) reactivated EBV in HH514-16 cells; this activity was also inhibited by VPA. Although VPA potently blocked the expression of viral lytic-cycle transcripts, it did not generally block the transcription of cellular genes and was not toxic. The levels and kinetics of specific cellular transcripts, such as Stat3, Frmd6, Mad1, Sepp1, c-fos, c-jun, and egr1, which were activated by NaB and TSA, were similar in HH514-16 cells treated with VPA. When combined with NaB or TSA, VPA did not inhibit the activation of these cellular genes. Changes in cellular gene expression in response to VPA, NaB, or TSA were globally similar as assessed by human genome arrays; however, VPA selectively stimulated the expression of some cellular genes, such as MEF2D, YY1, and ZEB1, that could repress the EBV lytic cycle. We describe a novel example of functional antagonism between HDAC inhibitors.
Asunto(s)
Antivirales/farmacología , Herpesvirus Humano 4/patogenicidad , Inhibidores de Histona Desacetilasas/metabolismo , Ácido Valproico/farmacología , Activación Viral/efectos de los fármacos , Línea Celular , Regulación Viral de la Expresión Génica/efectos de los fármacos , Herpesvirus Humano 4/crecimiento & desarrollo , Humanos , Transcripción Genética/efectos de los fármacosRESUMEN
The Epstein-Barr virus (EBV) lytic activator genes bzlf1 and brlf1 are conventionally referred to as immediate-early (IE) genes. However, previous studies showed that the earliest expression of these genes was blocked by cycloheximide when the EBV lytic cycle was induced by histone deacetylase (HDAC) inhibitors and protein kinase C agonists. Anti-IgG activates a complex signal transduction pathway that leads to EBV lytic activation in the Akata cell line. Here we demonstrate that in Akata cells, where lytic cycle activation occurs very rapidly after anti-IgG treatment, de novo protein synthesis is also required for induction of bzlf1 and brlf1 expression. New protein synthesis is required up to 1.25 h after application of anti-IgG; bzlf1 and brlf1 mRNAs can be detected 1.5 h after anti-IgG. Five cellular IE genes were shown to be expressed by 1 h after addition of anti-IgG, and their expression preceded that of bzlf1 and brlf1. These include early growth response genes (egr1, egr2, and egr3) and nuclear orphan receptors (nr4a1 and nr4a3). These genes were activated by anti-IgG treatment of Akata cells with and without the EBV genome; therefore, their expression was not dependent on expression of any EBV gene product. EGR1, EGR2, and EGR3 proteins were kinetically upstream of ZEBRA and Rta proteins. Expression of EGR1, ZEBRA, and Rta proteins were inhibited by bisindolylmaleimide X, a selective inhibitor of PKC. The findings suggest a revised model in which the signal transduction cascade activated by cross-linking of the B cell receptor induces expression of cellular IE genes, such as early growth response and nuclear orphan receptor genes, whose products, in turn, regulate bzlf1 and brlf1 expression.
Asunto(s)
Regulación de la Expresión Génica/fisiología , Herpesvirus Humano 4/fisiología , Proteínas Inmediatas-Precoces/metabolismo , Receptores de Antígenos de Linfocitos B/metabolismo , Transducción de Señal/fisiología , Transactivadores/metabolismo , Activación Viral/fisiología , Anticuerpos Antiidiotipos/farmacología , Northern Blotting , Línea Celular Tumoral , Reactivos de Enlaces Cruzados , Cartilla de ADN/genética , Factores de Transcripción de la Respuesta de Crecimiento Precoz/genética , Factores de Transcripción de la Respuesta de Crecimiento Precoz/metabolismo , Regulación de la Expresión Génica/efectos de los fármacos , Herpesvirus Humano 4/metabolismo , Humanos , Proteínas Inmediatas-Precoces/genética , Immunoblotting , Análisis por Micromatrices , Plásmidos/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Transactivadores/genéticaRESUMEN
Epstein-Barr virus (EBV) can be reactivated from latency into the lytic cycle by many stimuli believed to operate by different mechanisms. Cell lines containing EBV differ in their responses to inducing stimuli, yet all stimuli require de novo protein synthesis (44). A crucial step preliminary to identifying these proteins and determining when they are required is to measure the duration of stimulus and response time needed for activation of expression of EBV BRLF1 and BZLF1, the earliest viral indicators of reactivation. Here we show, with four EBV-containing cell lines that respond to different inducing agents, that stimuli that are effective at reactivating EBV can be divided into two main groups. The histone deacetylase inhibitors sodium butyrate and trichostatin A require a relatively long period of exposure, from 2 to 4 h or longer. Phorbol esters, anti-immunoglobulin G (anti-IgG), and, surprisingly, 5-aza-2'-deoxycytidine require short exposures of 15 min or less. The cell/virus background influences the response time. Expression of the EBV BZLF1 and BRLF1 genes can be detected before 2 h in Akata cells treated with anti-IgG, but both long- and short-duration stimuli required 4 or more hr to activate BZLF1 and BRLF1 expression in HH514-16, Raji, or B95-8 cells. Thus, stimulus duration and response time are independent variables. Neither stimulus duration nor response time can be predicted by the number of cells activated into the lytic cycle. These experiments shed new light on the earliest events leading to lytic cycle reactivation of EBV.
Asunto(s)
Anticuerpos Antiidiotipos/farmacología , Inhibidores Enzimáticos/farmacología , Herpesvirus Humano 4/fisiología , Inhibidores de Histona Desacetilasas , Linfocitos/virología , Ésteres del Forbol/farmacología , Activación Viral/efectos de los fármacos , Línea Celular , Herpesvirus Humano 4/metabolismo , Histona Desacetilasas/farmacología , Humanos , Proteínas Inmediatas-Precoces/genética , Proteínas Inmediatas-Precoces/metabolismo , Factores de Tiempo , Transactivadores/genética , Transactivadores/metabolismo , Proteínas Virales/genética , Proteínas Virales/metabolismoRESUMEN
The oncogenic human gammaherpesviruses, Epstein-Barr virus (EBV) and Kaposi's sarcoma-associated herpesvirus (KSHV), are latent in cultured lymphoma cells. We asked whether reactivation from latency of either virus requires de novo protein synthesis. Using Northern blotting and quantitative reverse transcriptase PCR, we measured the kinetics of expression of the lytic cycle activator genes and determined whether abundance of mRNAs encoding these genes from either virus was reduced by treatment with cycloheximide (CHX), an inhibitor of protein synthesis. CHX blocked expression of mRNAs of EBV BZLF1 and BRLF1, the two EBV lytic cycle activator genes, when HH514-16 Burkitt lymphoma cells were treated with histone deacetylase (HDAC) inhibitors, sodium butyrate or trichostatin A, or a DNA methyltransferase inhibitor, 5-Aza-2'-deoxycytidine. CHX also inhibited EBV lytic cycle activation in B95-8 marmoset lymphoblastoid cells by phorbol ester phorbol-12-myristate-13-acetate (TPA). EBV lytic cycle induction became resistant to CHX between 4 and 6 h after application of the inducing stimulus. KSHV lytic cycle activation, as assessed by ORF50 mRNA expression, was rapidly induced by the HDAC inhibitors, sodium butyrate and trichostatin A, in HH-B2 primary effusion lymphoma cells. In HH-B2 cells, CHX did not inhibit, but enhanced, expression of the KSHV lytic cycle activator gene, ORF50. In BC-1, a primary effusion lymphoma cell line that is dually infected with EBV and KSHV, CHX blocked EBV BRLF1 lytic gene expression induced by TPA and sodium butyrate; KSHV ORF50 mRNA induced simultaneously in the same cells by the same inducing stimuli was resistant to CHX. The experiments show, for the cell lines and inducing agents studied, that the EBV BZLF1 and BRLF1 genes do not behave with "immediate-early" kinetics upon reactivation from latency. KSHV ORF50 is a true "immediate-early" gene. Our results indicate that the mechanism by which HDAC inhibitors and TPA induce lytic cycle gene expression of the two viruses differs and suggest that EBV but not KSHV requires one or more proteins to be newly synthesized between 4 and 6 h after application of an inducing stimulus.
Asunto(s)
Herpesvirus Humano 4/crecimiento & desarrollo , Herpesvirus Humano 8/crecimiento & desarrollo , Inhibidores de Histona Desacetilasas , Biosíntesis de Proteínas , Proteína Quinasa C/metabolismo , Activación Viral/efectos de los fármacos , Azacitidina/análogos & derivados , Azacitidina/farmacología , Northern Blotting , Línea Celular , Cicloheximida/farmacología , Proteínas de Unión al ADN/genética , Decitabina , Inhibidores Enzimáticos/farmacología , Expresión Génica , Humanos , Ácidos Hidroxámicos/farmacología , Proteínas Inmediatas-Precoces/genética , Inhibidores de la Síntesis de la Proteína/farmacología , ARN Mensajero/biosíntesis , ARN Viral/biosíntesis , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Acetato de Tetradecanoilforbol/análogos & derivados , Acetato de Tetradecanoilforbol/farmacología , Transactivadores/genética , Proteínas Virales/biosíntesis , Proteínas Virales/genéticaRESUMEN
The seminal experiments of George and Eva Klein helped to define the two life cycles of Epstein-Barr Virus (EBV), namely latency and lytic or productive infection. Their laboratories described latent nuclear antigens expressed during latency and discovered several chemicals that activated the viral lytic cycle. The mechanism of the switch between latency and the lytic cycle of EBV and Kaposi's sarcoma-associated herpesvirus (KSHV) can be studied in cultured B cell lines. Lytic cycle activation of EBV is controlled by two viral transcription factors, ZEBRA and Rta. The homologue of Rta encoded in ORF50 is the lytic cycle activator of KSHV. Control of the lytic cycle can be divided into two distinct phases. Upstream events control expression of the virally encoded lytic cycle activator genes. Downstream events represent tasks carried out by the viral proteins in driving expression of lytic cycle genes and lytic viral DNA replication. In this chapter, we report three recent groups of experiments relating to upstream and downstream events. Azacytidine (AzaC) is a DNA methyltransferase inhibitor whose lytic cycle activation capacity was discovered by G. Klein and coworkers. We find that AzaC rapidly activates the EBV lytic cycle but does not detectably alter DNA methylation or histone acetylation on the promoters of the EBV lytic cycle activator genes. AzaC probably acts via a novel, yet to be elucidated, mechanism. The lytic cycle of both EBV and KSHV can be activated by sodium butyrate (NaB), a histone deacetylase inhibitor whose activity in disrupting latency was also discovered by G. Klein and coworkers. Activation of EBV by NaB requires protein synthesis; activation of KSHV is independent of protein synthesis. Thus, NaB works by a different pathway on the two closely related viruses. ZEBRA, the major downstream mediator of EBV lytic cycle activation is both a transcription activator and an essential replication protein. We show that phosphorylation of ZEBRA at its casein kinase 2 (CK2) site separates these two functions. Phosphorylation by CK2 is required for ZEBRA to activate lytic replication but not to induce expression of early lytic cycle genes. We discuss a number of unsolved mysteries about lytic cycle activation which should provide fertile territory for future research.
Asunto(s)
Efecto Citopatogénico Viral/fisiología , Regulación Viral de la Expresión Génica/genética , Herpesvirus Humano 4/fisiología , Herpesvirus Humano 8/fisiología , Virus Oncogénicos/fisiología , Azacitidina/farmacología , Cicloheximida/farmacología , Efecto Citopatogénico Viral/efectos de los fármacos , Efecto Citopatogénico Viral/genética , Replicación del ADN , ADN Viral/biosíntesis , ADN Viral/genética , Infecciones por Virus de Epstein-Barr/virología , Regulación Viral de la Expresión Génica/efectos de los fármacos , Infecciones por Herpesviridae/virología , Herpesvirus Humano 4/efectos de los fármacos , Herpesvirus Humano 4/genética , Herpesvirus Humano 8/efectos de los fármacos , Herpesvirus Humano 8/genética , Humanos , Proteínas Inmediatas-Precoces/fisiología , Mutación , Fosforilación , Procesamiento Proteico-Postraduccional , Estructura Terciaria de Proteína , Transactivadores/química , Transactivadores/genética , Transactivadores/fisiología , Infecciones Tumorales por Virus/virología , Latencia del Virus/efectos de los fármacos , Latencia del Virus/genéticaRESUMEN
Kaposi's sarcoma-associated herpesvirus (KSHV) can be driven into the lytic cycle in vitro by phorbol esters and sodium butyrate. This report begins to analyze the process by which butyrate activates the promoter of KSHV open reading frame 50 (ORF50), the key viral regulator of the KSHV latency to lytic cycle switch. A short fragment of the promoter, 134 nucleotides upstream of the translational start of ORF50, retained basal uninduced activity and conferred maximal responsiveness to sodium butyrate. The butyrate response element was mapped to a consensus Sp1-binding site. By means of electrophoretic mobility shift assays, both Sp1 and Sp3 were shown to form complexes in vitro with the ORF50 promoter at the Sp1 site. Butyrate induced the formation of a group of novel complexes, including several Sp3-containing complexes, one Sp1-containing complex, and several other complexes that were not identified with antibodies to Sp1 or Sp3. Formation of all butyrate-induced DNA-protein complexes was mediated by the consensus Sp1 site. In insect and mammalian cell lines, Sp1 significantly activated the ORF50 promoter linked to luciferase. Chromatin immunoprecipitation experiments in a PEL cell line showed that butyrate induced Sp1, CBP, and p300 binding to the ORF50 promoter in vivo in an on-off manner. The results suggest that induction of the KSHV lytic cycle by butyrate is mediated through interactions at the Sp1/Sp3 site located 103 to 112 nucleotides upstream of the translational initiation of ORF50 presumably by enhancing the binding of Sp1 to this site.
Asunto(s)
Butiratos/farmacología , Regulación Viral de la Expresión Génica , Herpesvirus Humano 8/efectos de los fármacos , Proteínas Inmediatas-Precoces/metabolismo , Regiones Promotoras Genéticas/genética , Factor de Transcripción Sp1/genética , Transactivadores/metabolismo , Proteínas Virales/metabolismo , Secuencia de Bases , Línea Celular Tumoral , Herpesvirus Humano 8/fisiología , Humanos , Proteínas Inmediatas-Precoces/genética , Datos de Secuencia Molecular , Elementos de Respuesta/genética , Transactivadores/genética , Proteínas Virales/genética , Replicación ViralRESUMEN
A DICOM Server Mediate Layer is introduced in this paper. It communicates with modalities according to DICOM3.0 standard on the one hand, provides a simple way to interface with other application on the other hand, this mades the implementation of DICOM service much easier for other applications.