RESUMEN
Mutations of the CBP/p300 histone acetyltransferase (HAT) domain can be linked to leukemic transformation in humans, suggestive of a checkpoint of leukocyte compartment sizes. Here, we examined the impact of reversible inhibition of this domain by the small-molecule A485. We found that A485 triggered acute and transient mobilization of leukocytes from the bone marrow into the blood. Leukocyte mobilization by A485 was equally potent as, but mechanistically distinct from, granulocyte colony-stimulating factor (G-CSF), which allowed for additive neutrophil mobilization when both compounds were combined. These effects were maintained in models of leukopenia and conferred augmented host defenses. Mechanistically, activation of the hypothalamus-pituitary-adrenal gland (HPA) axis by A485 relayed shifts in leukocyte distribution through corticotropin-releasing hormone receptor 1 (CRHR1) and adrenocorticotropic hormone (ACTH), but independently of glucocorticoids. Our findings identify a strategy for rapid expansion of the blood leukocyte compartment via a neuroendocrine loop, with implications for the treatment of human pathologies.
Asunto(s)
Médula Ósea , Histona Acetiltransferasas , Humanos , Histona Acetiltransferasas/metabolismo , Médula Ósea/metabolismo , Histonas/metabolismo , Neutrófilos/metabolismo , Sistema Hipotálamo-Hipofisario/metabolismoRESUMEN
CBP/p300 are transcription co-activators whose binding is a signature of enhancers, cis-regulatory elements that control patterns of gene expression in multicellular organisms. Active enhancers produce bi-directional enhancer RNAs (eRNAs) and display CBP/p300-dependent histone acetylation. Here, we demonstrate that CBP binds directly to RNAs in vivo and in vitro. RNAs bound to CBP in vivo include a large number of eRNAs. Using steady-state histone acetyltransferase (HAT) assays, we show that an RNA binding region in the HAT domain of CBP-a regulatory motif unique to CBP/p300-allows RNA to stimulate CBP's HAT activity. At enhancers where CBP interacts with eRNAs, stimulation manifests in RNA-dependent changes in the histone acetylation mediated by CBP, such as H3K27ac, and by corresponding changes in gene expression. By interacting directly with CBP, eRNAs contribute to the unique chromatin structure at active enhancers, which, in turn, is required for regulation of target genes.
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Histona Acetiltransferasas/metabolismo , ARN no Traducido/metabolismo , Factores de Transcripción p300-CBP/metabolismo , Acetilación , Animales , Línea Celular , Elementos de Facilitación Genéticos , Fibroblastos/metabolismo , Histonas/metabolismo , RatonesRESUMEN
Epigenome reprogramming after fertilization enables transcriptionally quiescent maternal and paternal chromatin to acquire a permissive state for subsequent zygotic genome activation (ZGA). H3K27 acetylation (H3K27ac) is a well-established chromatin marker of active enhancers and promoters. However, reprogramming dynamics of H3K27ac during maternal-to-zygotic transition (MZT) in mammalian embryos are not well-studied. By profiling the allelic landscape of H3K27ac during mouse MZT, we show that H3K27ac undergoes three waves of rapid global transitions between oocyte stage and 2-cell stage. Notably, germinal vesicle oocyte and zygote chromatin are globally hyperacetylated, with noncanonical, broad H3K27ac domains that correlate with broad H3K4 trimethylation (H3K4me3) and open chromatin. H3K27ac marks genomic regions primed for activation including ZGA genes, retrotransposons, and active alleles of imprinted genes. We show that CBP/p300 and HDAC activities play important roles in regulating H3K27ac dynamics and are essential for preimplantation development. Specifically, CBP/p300 acetyltransferase broadly deposits H3K27ac in zygotes to induce the opening of condensed chromatin at putative enhancers and ensure proper ZGA. On the contrary, HDACs revert broad H3K27ac domains to canonical domains and safeguard ZGA by preventing premature expression of developmental genes. In conclusion, coordinated activities of CBP/p300 and HDACs during mouse MZT are essential for ZGA and preimplantation development.
Asunto(s)
Regulación del Desarrollo de la Expresión Génica , Cigoto , Ratones , Animales , Cigoto/metabolismo , Acetilación , Blastocisto/metabolismo , Cromatina/genética , Cromatina/metabolismo , Mamíferos/genéticaRESUMEN
Host cells have evolved an intricate regulatory network to fine tune the type-I interferon responses. However, the full picture of this regulatory network remains to be depicted. In this study, we found that knock out of zinc-finger CCHC-type containing protein 8 (ZCCHC8) impairs the replication of influenza A virus (IAV), Sendai virus (Sev), Japanese encephalitis virus (JEV), and vesicular stomatitis virus (VSV). Further investigation unveiled that ZCCHC8 suppresses the type-I interferon responses by targeting the interferon regulatory factor 3 (IRF3) signaling pathway. Mechanistically, ZCCHC8 associates with phosphorylated IRF3 and disrupts the interaction of IRF3 with the co-activator CREB-binding protein (CBP). Additionally, the direct binding of ZCCHC8 with the IFN-stimulated response element (ISRE) impairs the ISRE-binding of IRF3. Our study contributes to the comprehensive understanding for the negative regulatory network of the type-I interferon responses and provides valuable insights for the control of multiple viruses from a host-centric perspective.IMPORTANCEThe innate immune responses serve as the initial line of defense against invading pathogens and harmful substances. Negative regulation of the innate immune responses plays an essential role in avoiding auto-immune diseases and over-activated immune responses. Hence, the comprehensive understanding of the negative regulation network for innate immune responses could provide novel therapeutic insights for the control of viral infections and immune dysfunction. In this study, we report that ZCCHC8 negatively regulates the type-I interferon responses. We illustrate that ZCCHC8 impedes the IRF3-CBP association by interacting with phosphorylated IRF3 and competes with IRF3 for binding to ISRE. Our study demonstrates the role of ZCCHC8 in the replication of multiple RNA viruses and contributes to a deeper understanding of the negative regulation system for the type-I interferon responses.
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Proteína de Unión a CREB , Inmunidad Innata , Factor 3 Regulador del Interferón , Interferón Tipo I , Virus Sendai , Transducción de Señal , Replicación Viral , Factor 3 Regulador del Interferón/metabolismo , Interferón Tipo I/metabolismo , Humanos , Células HEK293 , Virus Sendai/fisiología , Virus Sendai/genética , Proteína de Unión a CREB/metabolismo , Proteína de Unión a CREB/genética , Virus ARN/fisiología , Virus ARN/inmunología , Virus ARN/genética , Animales , Células A549 , Virus de la Influenza A/fisiología , Virus de la Influenza A/inmunología , Fosforilación , Interacciones Huésped-Patógeno , Vesiculovirus/fisiología , Virus de la Encefalitis Japonesa (Especie)/fisiología , Virus de la Encefalitis Japonesa (Especie)/inmunologíaRESUMEN
There are diverse phenotypes of castration-resistant prostate cancer, including neuroendocrine disease, that vary in their sensitivity to drug treatment. The efficacy of BET and CBP/p300 inhibitors in prostate cancer is attributed, at least in part, to their ability to decrease androgen receptor (AR) signalling. However, the activity of BET and CBP/p300 inhibitors in prostate cancers that lack the AR is unclear. In this study, we showed that BRD4, CBP, and p300 were co-expressed in AR-positive and AR-null prostate cancer. A combined inhibitor of these three proteins, NEO2734, reduced the growth of both AR-positive and AR-null organoids, as measured by changes in viability, size, and composition. NEO2734 treatment caused consistent transcriptional downregulation of cell cycle pathways. In neuroendocrine models, NEO2734 treatment reduced ASCL1 levels and other neuroendocrine markers, and reduced tumour growth in vivo. Collectively, these results show that epigenome-targeted inhibitors cause decreased growth and phenotype-dependent disruption of lineage regulators in neuroendocrine prostate cancer, warranting further development of compounds with this activity in the clinic. © 2024 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.
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Proteína p300 Asociada a E1A , Receptores Androgénicos , Transducción de Señal , Masculino , Humanos , Receptores Androgénicos/metabolismo , Receptores Androgénicos/genética , Animales , Proteína p300 Asociada a E1A/metabolismo , Proteína p300 Asociada a E1A/genética , Factores de Transcripción/metabolismo , Factores de Transcripción/genética , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Proteínas de Ciclo Celular/metabolismo , Proteínas de Ciclo Celular/genética , Neoplasias de la Próstata Resistentes a la Castración/patología , Neoplasias de la Próstata Resistentes a la Castración/metabolismo , Neoplasias de la Próstata Resistentes a la Castración/tratamiento farmacológico , Neoplasias de la Próstata Resistentes a la Castración/genética , Neoplasias de la Próstata/patología , Neoplasias de la Próstata/metabolismo , Neoplasias de la Próstata/tratamiento farmacológico , Neoplasias de la Próstata/genética , Regulación Neoplásica de la Expresión Génica , Ratones , Ensayos Antitumor por Modelo de Xenoinjerto , Proteínas que Contienen Bromodominio , Proteína de Unión a CREBRESUMEN
Epithelial mesenchymal transition (EMT) is a critical process implicated in the pathogenesis of retinal fibrosis and the exacerbation of diabetic retinopathy (DR) within retinal pigment epithelium (RPE) cells. Apigenin (AP), a potential dietary supplement for managing diabetes and its associated complications, has demonstrated inhibitory effects on EMT in various diseases. However, the specific impact and underlying mechanisms of AP on EMT in RPE cells remain poorly understood. In this study, we have successfully validated the inhibitory effects of AP on high glucose-induced EMT in ARPE-19 cells and diabetic db/db mice. Notably, our findings have identified CBP/p300 as a potential therapeutic target for EMT in RPE cells and have further substantiated that AP effectively downregulates the expression of EMT-related genes by attenuating the activity of CBP/p300, consequently reducing histone acetylation alterations within the promoter region of these genes. Taken together, our results provide novel evidence supporting the inhibitory effect of AP on EMT in RPE cells, and highlight the potential of specifically targeting CBP/p300 as a strategy for inhibiting retinal fibrosis in the context of DR.
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Apigenina , Transición Epitelial-Mesenquimal , Glucosa , Histonas , Epitelio Pigmentado de la Retina , Transición Epitelial-Mesenquimal/efectos de los fármacos , Epitelio Pigmentado de la Retina/efectos de los fármacos , Epitelio Pigmentado de la Retina/metabolismo , Epitelio Pigmentado de la Retina/patología , Animales , Apigenina/farmacología , Acetilación/efectos de los fármacos , Humanos , Glucosa/metabolismo , Glucosa/toxicidad , Histonas/metabolismo , Línea Celular , Ratones , Factores de Transcripción p300-CBP/metabolismo , Factores de Transcripción p300-CBP/antagonistas & inhibidores , Ratones Endogámicos C57BL , Retinopatía Diabética/metabolismo , Retinopatía Diabética/patología , Retinopatía Diabética/tratamiento farmacológico , Proteína p300 Asociada a E1A/metabolismo , Masculino , Células Epiteliales/efectos de los fármacos , Células Epiteliales/metabolismo , Células Epiteliales/patología , Proteína de Unión a CREB/metabolismo , Proteína de Unión a CREB/genéticaRESUMEN
Natural products are often uniquely suited to modulate protein-protein interactions (PPIs) due to their architectural and functional group complexity relative to synthetic molecules. Here we demonstrate that the natural product garcinolic acid allosterically blocks the CBP/p300 KIX PPI network and displays excellent selectivity over related GACKIX motifs. It does so via a strong interaction (KD 1â µM) with a non-canonical binding site containing a structurally dynamic loop in CBP/p300 KIX. Garcinolic acid engages full-length CBP in the context of the proteome and in doing so effectively inhibits KIX-dependent transcription in a leukemia model. As the most potent small-molecule KIX inhibitor yet reported, garcinolic acid represents an important step forward in the therapeutic targeting of CBP/p300.
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Proteína de Unión a CREB , Estructura Terciaria de Proteína , Dominios Proteicos , Sitios de Unión , Unión Proteica , Proteína de Unión a CREB/químicaRESUMEN
BACKGROUND: As an anti-inflammatory cytokine, interleukin 10 (IL-10) plays a vital role in preventing inflammatory and autoimmune pathologies while also maintaining immune homeostasis. IL-10 production in macrophages is tightly regulated by multiple pathways. TRIM24, a member of the Transcriptional Intermediary Factor 1 (TIF1) family, contributes to antiviral immunity and macrophage M2 polarization. However, the role of TRIM24 in regulating IL-10 expression and its involvement in endotoxic shock remains unclear. METHODS: In vitro, bone marrow derived macrophages cultured with GM-CSF or M-CSF were stimulated with LPS (100ng/ml). Murine models of endotoxic shock were established by challenging the mice with different dose of LPS (i.p). RTPCR, RNA sequencing, ELISA and hematoxylin and eosin staining were performed to elucidate the role and mechanisms of TRIM24 in endotoxic shock. RESULTS: The expression of TRIM24 is downregulated in LPS-stimulated bone marrow-derived macrophages (BMDMs). Loss of TRIM24 boosted IL-10 expression during the late stage of LPS-stimulation in macrophages. RNA-seq analysis revealed the upregulation of IFNß1, an upstream regulator of IL-10, in TRIM24 knockout macrophages. Treatment with C646, a CBP/p300 inhibitor, diminished the difference in both IFNß1 and IL-10 expression between TRIM24 knockout and control macrophages. Loss of TRIM24 provided protection against LPS-induced endotoxic shock in mice. CONCLUSION: Our results demonstrated that inhibiting TRIM24 promoted the expression of IFNß1 and IL-10 during macrophage activation, therefore protecting mice from endotoxic shock. This study offers novel insights into the regulatory role of TRIM24 in IL-10 expression, making it a potentially attractive therapeutic target for inflammatory diseases.
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Interleucina-10 , Choque Séptico , Ratones , Animales , Interleucina-10/genética , Interleucina-10/metabolismo , Lipopolisacáridos/farmacología , Lipopolisacáridos/metabolismo , Activación de Macrófagos , Macrófagos , Citocinas/metabolismo , Choque Séptico/metabolismoRESUMEN
Ecdysone signaling in Drosophila remains a popular model for investigating the mechanisms of steroid action in eukaryotes. The ecdysone receptor EcR can effectively bind ecdysone-response elements with or without the presence of a hormone. For years, EcR enhancers were thought to respond to ecdysone via recruiting coactivator complexes, which replace corepressors and stimulate transcription. However, the exact mechanism of transcription activation by ecdysone remains unclear. Here, we present experimental data on 11 various coregulators at ecdysone-responsive loci of Drosophila S2 cells. We describe the regulatory elements where coregulators reside within these loci and assess changes in their binding levels following 20-hydroxyecdysone treatment. In the current study, we detected the presence of some coregulators at the TSSs (active and inactive) and boundaries marked with CP190 rather than enhancers of the ecdysone-responsive loci where EcR binds. We observed minor changes in the coregulators' binding level. Most were present at inducible loci before and after 20-hydroxyecdysone treatment. Our findings suggest that: (1) coregulators can activate a particular TSS operating from some distal region (which could be an enhancer, boundary regulatory region, or inactive TSS); (2) coregulators are not recruited after 20-hydroxyecdysone treatment to the responsive loci; rather, their functional activity changes (shown as an increase in H3K27 acetylation marks generated by CBP/p300/Nejire acetyltransferase). Taken together, our findings imply that the 20-hydroxyecdysone signal enhances the functional activity of coregulators rather than promoting their binding to regulatory regions during the ecdysone response.
Asunto(s)
Proteínas de Drosophila , Receptores de Esteroides , Animales , Drosophila/genética , Drosophila/metabolismo , Ecdisona , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Ecdisterona/farmacología , Ecdisterona/metabolismo , Receptores de Esteroides/genética , Receptores de Esteroides/metabolismo , Activación Transcripcional , Drosophila melanogaster/metabolismo , Proteínas Asociadas a Microtúbulos/metabolismo , Proteínas Nucleares/metabolismoRESUMEN
Histone acetyltransferases of the CBP/p300 family are involved in transcriptional regulation and many biological processes (cell proliferation and differentiation, development, and regulation of the stress response and metabolism). Overexpression and knockdown of the nejire (nej) gene (codes for an ortholog of human CBP/p300 proteins) in various tissues (the fat body, intestine, and nervous system) and at various stages of the life cycle (throughout all developmental stages or in adulthood only) were tested for effect on lifespan in the fruit fly Drosophila melanogaster. The activation of nej exerted a positive or a negative effect on the lifespan, depending on the induction mode and the sex. A 6-15% greater lifespan was observed in females with conditional overexpression of nej in the intestine and constitutive overexpression of nej in the nervous system. A decrease (to 44%) or lack of significant changes in lifespan was detected in all other cases observed. In addition, stress response genes (Sod1, Gadd45, Hsp27, Hsp68, and Hif1) were regulated by nej activation. nej knockdown caused a pronounced negative effect on the D. melanogaster lifespan in most variants of the experiment.
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Proteínas de Drosophila , Drosophila melanogaster , Animales , Femenino , Humanos , Drosophila/genética , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Expresión Génica , Histona Acetiltransferasas/genética , Longevidad/genética , Sistema Nervioso/metabolismoRESUMEN
In developmental biology, the regulation of stem cell plasticity and differentiation remains an open question. CBP(CREB-binding protein)/p300 is a conserved gene family that functions as a transcriptional co-activator and plays important roles in a wide range of cellular processes, including cell death, the DNA damage response, and tumorigenesis. The acetyl transferase activity of CBPs is particularly important, as histone and non-histone acetylation results in changes in chromatin architecture and protein activity that affect gene expression. Many studies have described the conserved functions of CBP/p300 in stem cell proliferation and differentiation. The planarian Schmidtea mediterranea is an excellent model for the in vivo study of the molecular mechanisms underlying stem cell differentiation during regeneration. However, how this process is regulated genetically and epigenetically is not well-understood yet. We identified 5 distinct Smed-cbp genes in S. mediterranea that show different expression patterns. Functional analyses revealed that Smed-cbp-2 appears to be essential for stem cell maintenance. On the other hand, the silencing of Smed-cbp-3 resulted in the growth of blastemas that were apparently normal, but remained largely unpigmented and undifferentiated. Smed-cbp-3 silencing also affected the differentiation of several cell lineages including neural, epidermal, digestive, and excretory cell types. Finally, we analysed the predicted interactomes of CBP-2 and CBP-3 as an initial step to better understand their functions in planarian stem cell biology. Our results indicate that planarian cbp genes play key roles in stem cell maintenance and differentiation.
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Proteína de Unión a CREB/metabolismo , Diferenciación Celular/genética , Planarias/genética , Animales , Proteína de Unión a CREB/genética , Proteína de Unión a CREB/fisiología , Diferenciación Celular/fisiología , Proliferación Celular , Cromatina/metabolismo , Histonas/metabolismo , Planarias/metabolismo , Regeneración/genética , Células Madre/citología , Células Madre/metabolismo , Células Madre/fisiología , Factores de Transcripción/metabolismoRESUMEN
Acute lymphoblastic leukemia with chromosomal rearrangements involving the mixed-lineage leukemia (MLL) gene (MLL-r ALL) remains an incurable disease. Thus, development of a safe and effective therapeutic agent to treat this disease is crucial to address this unmet medical need. BRD4, a member of the bromodomain and extra-terminal domain (BET) protein family, and cyclic AMP response element binding protein binding protein (CBP) and p300, two paralogous histone acetyltransferases, are all considered cancer drug targets and simultaneous targeting of these proteins may have therapeutic advantages. Here, we demonstrate that a BET/CBP/p300 multi-bromodomain inhibitor, CN470, has anti-tumor activity against MLL-r ALL in vitro and in vivo. CN470, potently inhibited ligand binding to the bromodomains of BRD4, CBP, and p300 and suppressed the growth of MLL-r ALL cell lines and patient-derived cells with MLL rearrangements. CN470 suppressed mRNA and protein expression of MYC and induced apoptosis in MLL-r ALL cells, following a cell cycle arrest in the G1 phase. Moreover, CN470 reduced BRD4 binding to acetylated histone H3. The in vivo effects of CN470 were investigated using SEMLuc/GFP cells expressing luminescent markers in an orthotopic mouse model. Mice administered CN470 daily had prolonged survival compared to the vehicle group. Further, CN470 also showed anti-tumor effects against an MLL-r ALL patient-derived xenograft model. These findings suggest that inhibition of BET/CBP/p300 by the multi-bromodomain inhibitor, CN470, represents a promising therapeutic approach against MLL-r ALL.
Asunto(s)
Antineoplásicos/farmacología , Proteína p300 Asociada a E1A/antagonistas & inhibidores , Reordenamiento Génico , N-Metiltransferasa de Histona-Lisina/genética , Proteína de la Leucemia Mieloide-Linfoide/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Animales , Apoptosis/efectos de los fármacos , Apoptosis/genética , Puntos de Control del Ciclo Celular/efectos de los fármacos , Puntos de Control del Ciclo Celular/genética , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Proliferación Celular/genética , Proteína p300 Asociada a E1A/metabolismo , Regulación Leucémica de la Expresión Génica/efectos de los fármacos , Reordenamiento Génico/efectos de los fármacos , Humanos , Ratones , Leucemia-Linfoma Linfoblástico de Células Precursoras/patología , ARN Mensajero/genética , ARN Mensajero/metabolismo , Análisis de Supervivencia , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
We previously reported that the cell cycle-related cyclin-dependent kinase 4-retinoblastoma (RB) transcriptional corepressor pathway is essential for stroke-induced cell death both in vitro and in vivo However, how this signaling pathway induces cell death is unclear. Previously, we found that the cyclin-dependent kinase 4 pathway activates the pro-apoptotic transcriptional co-regulator Cited2 in vitro after DNA damage. In the present study, we report that Cited2 protein expression is also dramatically increased following stroke/ischemic insult. Critically, utilizing conditional knockout mice, we show that Cited2 is required for neuronal cell death, both in culture and in mice after ischemic insult. Importantly, determining the mechanism by which Cited2 levels are regulated, we found that E2F transcription factor (E2F) family members participate in Cited2 regulation. First, E2F1 expression induced Cited2 transcription, and E2F1 deficiency reduced Cited2 expression. Moreover, determining the potential E2F-binding regions on the Cited2 gene regulatory sequence by ChIP analysis, we provide evidence that E2F1/4 proteins bind to this DNA region. A luciferase reporter assay to probe the functional outcomes of this interaction revealed that E2F1 activates and E2F4 inhibits Cited2 transcription. Moreover, we identified the functional binding motif for E2F1 in the Cited2 gene promoter by demonstrating that mutation of this site dramatically reduces E2F1-mediated Cited2 transcription. Finally, E2F1 and E2F4 regulated Cited2 expression in neurons after stroke-related insults. Taken together, these results indicate that the E2F-Cited2 regulatory pathway is critically involved in stroke injury.
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Factor de Transcripción E2F1/metabolismo , Factor de Transcripción E2F4/metabolismo , Regulación de la Expresión Génica , Neuronas/metabolismo , Proteínas Represoras/biosíntesis , Accidente Cerebrovascular/metabolismo , Transactivadores/biosíntesis , Secuencias de Aminoácidos , Animales , Muerte Celular , Factor de Transcripción E2F1/genética , Factor de Transcripción E2F4/genética , Ratones , Ratones Transgénicos , Neuronas/patología , Proteínas Represoras/genética , Accidente Cerebrovascular/genética , Accidente Cerebrovascular/patología , Transactivadores/genéticaRESUMEN
Homeostasis of adult tissues is maintained by a small number of stem cells, which are sustained by their niches. In the Drosophila female germline stem cell (GSC) niche, Decapentaplegic (Dpp) is the primary factor that promotes GSC self-renewal. However, the mechanism regulating dpp expression in the niche is largely unknown. Here, we identify a 2.0â kb fragment located in a 5' cis-regulatory region of the dpp locus containing enhancer activity that drives its expression in the niche. This region is distinct from a previously characterized 3' cis-regulatory enhancer responsible for dpp expression in imaginal discs. Our data demonstrate that Engrailed, a homeodomain-containing transcription factor that serves as a cap cell marker, binds to this region and regulates dpp expression in cap cells. Further data suggest that En forms a complex with Nejire (Nej), the Drosophila ortholog of histone acetyltransferase CBP/p300, and directs Nej to this cis-regulatory region where Nej functions as the co-activator for dpp expression. Therefore, our study defines the molecular pathway controlling dpp expression in the Drosophila ovarian stem cell niche.
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Proteínas de Drosophila/metabolismo , Drosophila melanogaster/citología , Drosophila melanogaster/metabolismo , Proteínas de Homeodominio/metabolismo , Ovario/citología , Ovario/metabolismo , Nicho de Células Madre , Factores de Transcripción/metabolismo , Factores de Transcripción p300-CBP/metabolismo , Animales , Proteínas de Drosophila/genética , Femenino , Regulación de la Expresión Génica , Genes Reporteros , Unión Proteica , Secuencias Reguladoras de Ácidos Nucleicos/genética , Nicho de Células Madre/genética , Células Madre/citología , Células Madre/metabolismoRESUMEN
Histone acetylation has been recognized as an important post-translational modification of core nucleosomal histones that changes access to the chromatin to allow gene transcription, DNA replication, and repair. Histone acetyltransferases were initially identified as co-activators that link DNA-binding transcription factors to the general transcriptional machinery. Over the years, more chromatin-binding modes have been discovered suggesting direct interaction of histone acetyltransferases and their protein complex partners with histone proteins. While much progress has been made in characterizing histone acetyltransferase complexes biochemically, cell-free activity assay results are often at odds with in-cell histone acetyltransferase activities. In-cell studies suggest specific histone lysine targets, but broad recruitment modes, apparently not relying on specific DNA sequences, but on chromatin of a specific functional state. Here we review the evidence for general versus specific roles of individual nuclear lysine acetyltransferases in light of in vivo and in vitro data in the mammalian system.
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Genoma , Histona Acetiltransferasas/metabolismo , Histonas/metabolismo , Lisina/metabolismo , Acetilación , Animales , Cromatina/metabolismo , Eliminación de Gen , Histona Acetiltransferasas/genética , Humanos , Lisina Acetiltransferasas/genética , Lisina Acetiltransferasas/metabolismo , Mamíferos , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Procesamiento Proteico-Postraduccional , ARN Interferente PequeñoRESUMEN
The canonical Wnt signaling pathway is a master cell regulator involved in CD8+ T cell proliferation and differentiation. In human CD8+ T cells, this pathway induces differentiation into memory cells or a "stem cell memory like" population, which is preferentially present in cord blood. To better understand the role of canonical Wnt signals in neonatal or adult blood, we compared the proteins associated with ß-catenin, in nonstimulated and Wnt3a-stimulated human neonatal and adult naive CD8+ T cells. Differentially recruited proteins established different complexes in adult and neonatal cells. In the former, ß-catenin-associated proteins were linked to cell signaling and immunological functions, whereas those of neonates were linked to proliferation and metabolism. Wnt3a stimulation led to the recruitment and overexpression of Wnt11 in adult cells and Wnt5a in neonatal cells, suggesting a differential connexion with planar polarity and Wnt/Ca2+ noncanonical pathways, respectively. The chromatin immunoprecipitation polymerase chain reaction ß-catenin was recruited to a higher level on the promoters of cell renewal genes in neonatal cells and of differentiation genes in those of adults. We found a preferential association of ß-catenin with CBP in neonatal cells and with p300 in the adult samples, which could be involved in a higher self-renewal capacity of the neonatal cells and memory commitment in those of adults. Altogether, our results show that different proteins associated with ß-catenin during Wnt3a activation mediate a differential response of neonatal and adult human CD8+ T cells.
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Linfocitos T CD8-positivos/citología , Linfocitos T CD8-positivos/metabolismo , Diferenciación Celular , Complejos Multiproteicos/metabolismo , beta Catenina/metabolismo , Adulto , Regulación de la Expresión Génica , Humanos , Recién Nacido , Regiones Promotoras Genéticas/genética , Unión Proteica , Mapeo de Interacción de Proteínas , Vía de Señalización WntRESUMEN
BACKGROUND: HIV-associated neurocognitive disorder (HAND) is a neurodegenerative disease associated with persistent neuroinflammation and subsequent neuron damage. Pro-inflammatory factors and neurotoxins from activated astrocytes by HIV-1 itself and its encoded proteins, including the negative factor (Nef), are involved in the pathogenesis of HAND. This study was designed to find potential lncRNAs that regulate astrocyte functions and inflammation process. METHODS: We performed microarray analysis of lncRNAs from primary mouse astrocytes treated with Nef protein. Top ten lncRNAs were validated through real-time PCR analysis. Gene ontology (GO) and KEGG pathway analysis were applied to explore the potential functions of lncRNAs. RIP and ChIP assays were performed to demonstrate the mechanism of lncRNA regulating gene expression. RESULTS: There were 638 co-upregulated lncRNAs and 372 co-downregulated lncRNAs in primary astrocytes treated with Nef protein for both 6 h and 12 h. GO and KEGG pathway analysis showed that the biological functions of top differential-expressed mRNAs were associated with inflammatory cytokines and chemokine. Knockdown of lncRNA AK006025, not AK138360, inhibited significantly CXCL9, CXCL10 (IP-10), and CXCL11 expression in astrocytes treated with Nef protein. Mechanism study showed that AK006025 associated with CBP/P300 was enriched in the promoter of CXCL9, CXCL10, and CXCL11 genes. CONCLUSIONS: Our findings uncovered the expression profiles of lncRNAs and mRNAs in vitro, which might help to understand the pathways that regulate astrocyte activation during the process of HAND.
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Astrocitos/efectos de los fármacos , Quimiocina CXCL1/metabolismo , Regulación de la Expresión Génica/efectos de los fármacos , Proteínas de la Membrana/metabolismo , Fosfoproteínas/metabolismo , ARN Largo no Codificante/metabolismo , Productos del Gen nef del Virus de la Inmunodeficiencia Humana/farmacología , Animales , Animales Recién Nacidos , Astrocitos/metabolismo , Proteínas de Unión al Calcio/metabolismo , Células Cultivadas , Corteza Cerebral/citología , Regulación de la Expresión Génica/genética , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Humanos , Ratones , Ratones Endogámicos C57BL , Proteínas de Microfilamentos/metabolismo , ARN Mensajero/metabolismo , ARN Interferente Pequeño/genética , ARN Interferente Pequeño/metabolismo , Transducción de Señal/efectos de los fármacos , Transducción de Señal/genética , Factores de Tiempo , Transfección , Productos del Gen nef del Virus de la Inmunodeficiencia Humana/metabolismoRESUMEN
Th17 responses are critical to a variety of human autoimmune diseases, and therapeutic targeting with monoclonal antibodies against IL-17 and IL-23 has shown considerable promise. Here, we report data to support selective bromodomain blockade of the transcriptional coactivators CBP (CREB binding protein) and p300 as an alternative approach to inhibit human Th17 responses. We show that CBP30 has marked molecular specificity for the bromodomains of CBP and p300, compared with 43 other bromodomains. In unbiased cellular testing on a diverse panel of cultured primary human cells, CBP30 reduced immune cell production of IL-17A and other proinflammatory cytokines. CBP30 also inhibited IL-17A secretion by Th17 cells from healthy donors and patients with ankylosing spondylitis and psoriatic arthritis. Transcriptional profiling of human T cells after CBP30 treatment showed a much more restricted effect on gene expression than that observed with the pan-BET (bromo and extraterminal domain protein family) bromodomain inhibitor JQ1. This selective targeting of the CBP/p300 bromodomain by CBP30 will potentially lead to fewer side effects than with the broadly acting epigenetic inhibitors currently in clinical trials.
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Bencimidazoles/farmacología , Inmunosupresores/farmacología , Interleucina-17/metabolismo , Isoxazoles/farmacología , Células Th17/efectos de los fármacos , Factores de Transcripción p300-CBP/antagonistas & inhibidores , Adulto , Anciano , Artritis Psoriásica/metabolismo , Artritis Psoriásica/patología , Azepinas/farmacología , Bencimidazoles/química , Linfocitos T CD4-Positivos/efectos de los fármacos , Linfocitos T CD4-Positivos/metabolismo , Calorimetría , Células Cultivadas , Cristalografía por Rayos X , Evaluación Preclínica de Medicamentos , Femenino , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Inmunosupresores/química , Interleucina-17/biosíntesis , Interleucina-17/genética , Isoxazoles/química , Cinética , Masculino , Persona de Mediana Edad , Modelos Moleculares , Estructura Molecular , Conformación Proteica , Estructura Terciaria de Proteína/efectos de los fármacos , Proteínas Recombinantes/metabolismo , Espondilitis Anquilosante/metabolismo , Espondilitis Anquilosante/patología , Relación Estructura-Actividad , Células Th17/inmunología , Triazoles/farmacologíaRESUMEN
The teratogenicity of the antiepileptic drug valproic acid (VPA) is well established and its inhibition of histone deacetylases (HDAC) is proposed as an initiating factor. Recently, VPA-mediated HDAC inhibition was demonstrated to involve transcriptional downregulation of histone acetyltransferases (HATs), which was proposed to compensate for the increased acetylation resulting from HDAC inhibition. Cbp and p300 are HATs required for embryonic development and deficiencies in either are associated with congenital malformations and embryolethality. The objective of the present study was to characterize Cbp/p300 following VPA exposure in P19 cells. Consistent with previous studies, exposure to 5mM VPA over 24h induced a moderate decrease in Cbp/p300 mRNA, which preceded a strong decrease in total cellular protein mediated by ubiquitin-proteasome degradation. Nuclear Cbp/p300 protein was also decreased following VPA exposure, although to a lesser extent. Total cellular and nuclear p300 HAT activity was reduced proportionately to p300 protein levels, however while total cellular HAT activity also decreased, nuclear HAT activity was unaffected. Using the Cbp/p300 HAT inhibitor C646, we demonstrated that HAT inhibition similarly affected many of the same endpoints as VPA, including increased reactive oxygen species and caspase-3 cleavage, the latter of which could be attenuated by pre-treatment with the antioxidant catalase. C646 exposure also decreased NF-κB/p65 protein, which was not due to reduced mRNA and was not attenuated with catalase pre-treatment. This study provides support for an adaptive HAT response following VPA exposure and suggests that reduced Cbp/p300 HAT activity could contribute to VPA-mediated alterations.
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Anticonvulsivantes/farmacología , Histona Acetiltransferasas/metabolismo , Ácido Valproico/farmacología , Factores de Transcripción p300-CBP/metabolismo , Animales , Apoptosis/efectos de los fármacos , Benzoatos/farmacología , Caspasa 3/metabolismo , Línea Celular , Regulación hacia Abajo , Ratones , Nitrobencenos , Pirazoles/farmacología , Pirazolonas , ARN Mensajero/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Factor de Transcripción ReIA/genética , Factor de Transcripción ReIA/metabolismo , Factores de Transcripción p300-CBP/antagonistas & inhibidores , Factores de Transcripción p300-CBP/genéticaRESUMEN
CBP and p300 are homologous proteins exhibiting remarkable structural and functional similarity. Both proteins function as acetyltransferase and coactivator, underscoring their significant roles in cellular processes. The function of histone acetyltransferases is to facilitate the release of DNA from nucleosomes and act as transcriptional co-activators to promote gene transcription. Transcription factors recruit CBP/p300 by co-condensation and induce transcriptional bursting. Disruption of CBP or p300 functions is associated with different diseases, especially cancer, which can result from either loss of function or gain of function. CBP and p300 are multidomain proteins containing HAT (histone acetyltransferase) and BRD (bromodomain) domains, which perform acetyltransferase activity and maintenance of HAT signaling, respectively. Inhibitors targeting HAT and BRD have been explored for decades, and some BRD inhibitors have been evaluated in clinical trials for treating hematologic malignancies or advanced solid tumors. Here, we review the development and application of CBP/p300 inhibitors. Several inhibitors have been evaluated in vivo, exhibiting notable potency but limited selectivity. Exploring these inhibitors emphasizes the promise of targeting CBP and p300 with small molecules in cancer therapy.