RESUMEN
Mammalian SWI/SNF chromatin remodeling complexes move and evict nucleosomes at gene promoters and enhancers to modulate DNA access. Although SWI/SNF subunits are commonly mutated in disease, therapeutic options are limited by our inability to predict SWI/SNF gene targets and conflicting studies on functional significance. Here, we leverage a fast-acting inhibitor of SWI/SNF remodeling to elucidate direct targets and effects of SWI/SNF. Blocking SWI/SNF activity causes a rapid and global loss of chromatin accessibility and transcription. Whereas repression persists at most enhancers, we uncover a compensatory role for the EP400/TIP60 remodeler, which reestablishes accessibility at most promoters during prolonged loss of SWI/SNF. Indeed, we observe synthetic lethality between EP400 and SWI/SNF in cancer cell lines and human cancer patient data. Our data define a set of molecular genomic features that accurately predict gene sensitivity to SWI/SNF inhibition in diverse cancer cell lines, thereby improving the therapeutic potential of SWI/SNF inhibitors.
Asunto(s)
Proteínas Nucleares , Factores de Transcripción , Animales , Humanos , Cromatina , Ensamble y Desensamble de Cromatina , Proteínas Nucleares/metabolismo , Nucleosomas , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , RatonesRESUMEN
Cell fate decisions are governed by sequence-specific transcription factors (TFs) that act in small populations of cells within developing embryos. To understand their functions in vivo, it is important to identify TF binding sites in these cells. However, current methods cannot profile TFs genome-wide at or near the single-cell level. Here we adapt the cleavage under targets and release using nuclease (CUT&RUN) method to profile TFs in low cell numbers, including single cells and individual pre-implantation embryos. Single-cell experiments suggest that only a fraction of TF binding sites are occupied in most cells, in a manner broadly consistent with measurements of peak intensity from multi-cell studies. We further show that chromatin binding by the pluripotency TF NANOG is highly dependent on the SWI/SNF chromatin remodeling complex in individual blastocysts but not in cultured cells. Ultra-low input CUT&RUN (uliCUT&RUN) therefore enables interrogation of TF binding from rare cell populations of particular importance in development or disease.
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Blastocisto/metabolismo , Ensamble y Desensamble de Cromatina/fisiología , Cromatina/metabolismo , Perfilación de la Expresión Génica , Regulación del Desarrollo de la Expresión Génica/fisiología , Factores de Transcripción/metabolismo , Animales , Femenino , RatonesRESUMEN
Cyclin-dependent kinase 9 (CDK9) promotes transcriptional elongation through RNAPII pause release. We now report that CDK9 is also essential for maintaining gene silencing at heterochromatic loci. Through a live cell drug screen with genetic confirmation, we discovered that CDK9 inhibition reactivates epigenetically silenced genes in cancer, leading to restored tumor suppressor gene expression, cell differentiation, and activation of endogenous retrovirus genes. CDK9 inhibition dephosphorylates the SWI/SNF protein BRG1, which contributes to gene reactivation. By optimization through gene expression, we developed a highly selective CDK9 inhibitor (MC180295, IC50 = 5 nM) that has broad anti-cancer activity in vitro and is effective in in vivo cancer models. Additionally, CDK9 inhibition sensitizes to the immune checkpoint inhibitor α-PD-1 in vivo, making it an excellent target for epigenetic therapy of cancer.
Asunto(s)
Quinasa 9 Dependiente de la Ciclina/metabolismo , Animales , Línea Celular Tumoral , Quinasa 9 Dependiente de la Ciclina/antagonistas & inhibidores , Quinasa 9 Dependiente de la Ciclina/genética , ADN Helicasas/genética , ADN Helicasas/metabolismo , Metilación de ADN , Femenino , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Leucocitos Mononucleares/citología , Leucocitos Mononucleares/efectos de los fármacos , Leucocitos Mononucleares/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos NOD , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Proto-Oncogénicas c-myc/metabolismo , Interferencia de ARN , ARN Interferente Pequeño/metabolismo , Bibliotecas de Moléculas Pequeñas/química , Bibliotecas de Moléculas Pequeñas/farmacología , Relación Estructura-Actividad , Factores de Transcripción/genética , Factores de Transcripción/metabolismoRESUMEN
RNA transcribed from enhancers, i.e., eRNA, has been suggested to directly activate transcription by recruiting transcription factors and co-activators. Although there have been specific examples of eRNA functioning in this way, it is not clear how general this may be. We find that the AT-hook of SWI/SNF preferentially binds RNA and, as part of the esBAF complex, associates with eRNA transcribed from intronic and intergenic regions. Our data suggest that SWI/SNF is globally recruited in cis by eRNA to cell-type-specific enhancers, representative of two distinct stages that mimic early mammalian development, and not at enhancers that are shared between the two stages. In this manner, SWI/SNF facilitates recruitment and/or activation of MLL3/4, p300/CBP, and Mediator to stage-specific enhancers and super-enhancers that regulate the transcription of metabolic and cell lineage priming-related genes. These findings highlight a connection between ATP-dependent chromatin remodeling and eRNA in cell identity and typical- and super-enhancer activation.
Asunto(s)
Linaje de la Célula , ADN Helicasas , Elementos de Facilitación Genéticos , Proteínas Nucleares , Factores de Transcripción , Animales , Humanos , Linaje de la Célula/genética , Ensamble y Desensamble de Cromatina , Proteínas Cromosómicas no Histona/metabolismo , Proteínas Cromosómicas no Histona/genética , ADN Helicasas/metabolismo , ADN Helicasas/genética , Proteínas Nucleares/metabolismo , Proteínas Nucleares/genética , Factores de Transcripción/metabolismo , Factores de Transcripción/genéticaRESUMEN
B lineage priming by pioneer transcription factor EBF1 requires the function of an intrinsically disordered region (IDR). Here, we examine the role of regularly spaced tyrosines in the IDR as potential determinants of IDR function and activity of EBF1. We found that four Y > A mutations in EBF1 reduced the formation of condensates in vitro and subdiffractive clusters in vivo. Notably, Y > A mutant EBF1 was inefficient in promoting B cell differentiation and showed impaired chromatin binding, recruitment of BRG1, and activation of specific target genes. Thus, regularly spaced tyrosines in the IDR contribute to the biophysical and functional properties of EBF1.
Asunto(s)
Linfocitos B , Regulación de la Expresión Génica , Linaje de la Célula/genética , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Heterocromatina/metabolismoRESUMEN
Polycomb group (PcG) proteins are essential for post-implantation development by depositing repressive histone modifications at promoters, mainly CpG islands (CGIs), of developmental regulator genes. However, promoter PcG marks are erased after fertilization and de novo established in peri-implantation embryos, coinciding with the transition from naive to primed pluripotency. Nevertheless, the molecular basis for this establishment remains unknown. In this study, we show that the expression of the long KDM2B isoform (KDM2BLF), which contains the demethylase domain, is specifically induced at peri-implantation and that its H3K36me2 demethylase activity is required for PcG enrichment at CGIs. Moreover, KDM2BLF interacts with BRG1/BRM-associated factor (BAF) and stabilizes BAF occupancy at CGIs for subsequent gain of accessibility, which precedes PcG enrichment. Consistently, KDM2BLF inactivation results in significantly delayed post-implantation development. In summary, our data unveil dynamic chromatin configuration of CGIs during exit from naive pluripotency and provide a conceptual framework for the spatiotemporal establishment of PcG functions.
Asunto(s)
Cromatina , Proteínas de Drosophila , Islas de CpG , Proteínas de Drosophila/metabolismo , Código de Histonas , Proteínas del Grupo Polycomb/genética , Proteínas del Grupo Polycomb/metabolismo , Regiones Promotoras GenéticasRESUMEN
Establishment of B-lineage-specific gene expression requires the binding of transcription factors to inaccessible chromatin of progenitors. The transcription factor EBF1 can bind genomic regions prior to the detection of chromatin accessibility in a manner dependent on EBF1's C-terminal domain (CTD) and independent of cooperating transcription factors. Here, we studied the mechanism whereby the CTD enables this pioneering function. The CTD of EBF1 was dispensable for initial chromatin targeting but stabilized occupancy via recruitment of the chromatin remodeler Brg1. We found that the CTD harbors a prion-like domain (PLD) with an ability of liquid-liquid phase separation, which was enhanced by interaction of EBF1 with the RNA-binding protein FUS. Brg1 also partitioned into phase-separated FUS condensates and coincided with EBF1 and FUS foci in pro-B cells. Heterologous PLDs conferred pioneering function on EBF1ΔCTD. Thus, the phase separation ability of EBF1 facilitates Brg1-mediated chromatin opening and the transition of naive progenitor chromatin to B-lineage-committed chromatin.
Asunto(s)
Linfocitos B/metabolismo , Cromatina/metabolismo , Priones/química , Transactivadores/metabolismo , Secuencia de Aminoácidos , Linfocitos B/citología , ADN Helicasas/metabolismo , Humanos , Proteínas Nucleares/metabolismo , Transición de Fase , Células Precursoras de Linfocitos B/citología , Células Precursoras de Linfocitos B/metabolismo , Dominios Proteicos , Proteína FUS de Unión a ARN/metabolismo , Proteínas de Unión al ARN/metabolismo , Transactivadores/química , Factores de Transcripción/metabolismoRESUMEN
SWI/SNF-family remodelers (BAF/PBAF in mammals) are essential chromatin regulators, and mutations in human BAF/PBAF components are associated with â¼20% of cancers. Cancer-associated missense mutations in human BRG1 (encoding the catalytic ATPase) have been characterized previously as conferring loss-of-function. Here, we show that cancer-associated missense mutations in BRG1, when placed into the orthologous Sth1 ATPase of the yeast RSC remodeler, separate into two categories: loss-of-function enzymes, or instead, gain-of-function enzymes that greatly improve DNA translocation efficiency and nucleosome remodeling in vitro. Our work identifies a structural "hub," formed by the association of several Sth1 domains, that regulates ATPase activity and DNA translocation efficiency. Remarkably, all gain-of-function cancer-associated mutations and all loss-of-function mutations physically localize to distinct adjacent regions in the hub, which specifically regulate and implement DNA translocation, respectively. In vivo, only gain-of-function cancer-associated mutations conferred precocious chromatin accessibility. Taken together, we provide a structure-function mechanistic basis for cancer-associated hyperactivity.
Asunto(s)
Proteínas de Ciclo Celular/metabolismo , Ensamble y Desensamble de Cromatina , Proteínas Cromosómicas no Histona/metabolismo , Mutación con Ganancia de Función , Neoplasias/patología , Proteínas Nucleares/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas Cromosómicas no Histona/genética , ADN Helicasas/genética , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Humanos , Neoplasias/genética , Proteínas Nucleares/genética , Nucleosomas , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/crecimiento & desarrollo , Proteínas de Saccharomyces cerevisiae/genética , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Translocación GenéticaRESUMEN
BET proteins commonly activate cellular gene expression, yet inhibiting their recruitment paradoxically reactivates latent HIV-1 transcription. Here we identify the short isoform of BET family member BRD4 (BRD4S) as a corepressor of HIV-1 transcription. We found that BRD4S was enriched in chromatin fractions of latently infected T cells, and it was more rapidly displaced from chromatin upon BET inhibition than the long isoform. BET inhibition induced marked nucleosome remodeling at the latent HIV-1 promoter, which was dependent on the activity of BRG1-associated factors (BAF), an SWI/SNF chromatin-remodeling complex with known repressive functions in HIV-1 transcription. BRD4S directly bound BRG1, a catalytic subunit of BAF, via its bromodomain and extraterminal (ET) domain, and this isoform was necessary for BRG1 recruitment to latent HIV-1 chromatin. Using chromatin immunoprecipitation sequencing (ChIP-seq) combined with assay for transposase-accessible chromatin coupled to high-throughput sequencing (ATAC-seq) data, we found that the latent HIV-1 promoter phenotypically resembles endogenous long terminal repeat (LTR) sequences, pointing to a select role of BRD4S-BRG1 complexes in genomic silencing of invasive retroelements.
Asunto(s)
Ensamble y Desensamble de Cromatina , Cromatina/metabolismo , Proteínas Cromosómicas no Histona/metabolismo , ADN Viral/metabolismo , VIH-1/metabolismo , Proteínas Nucleares/metabolismo , Linfocitos T/metabolismo , Factores de Transcripción/metabolismo , Transcripción Genética , Latencia del Virus , Azepinas/farmacología , Proteínas de Ciclo Celular , Cromatina/genética , Ensamble y Desensamble de Cromatina/efectos de los fármacos , Inmunoprecipitación de Cromatina , Proteínas Cromosómicas no Histona/efectos de los fármacos , Proteínas Cromosómicas no Histona/genética , ADN Helicasas/genética , ADN Helicasas/metabolismo , ADN Viral/genética , Relación Dosis-Respuesta a Droga , Regulación hacia Abajo , Regulación Viral de la Expresión Génica , Células HEK293 , VIH-1/efectos de los fármacos , VIH-1/genética , VIH-1/inmunología , Secuenciación de Nucleótidos de Alto Rendimiento , Interacciones Huésped-Patógeno , Humanos , Células Jurkat , Proteínas Nucleares/genética , Regiones Promotoras Genéticas , Unión Proteica , Isoformas de Proteínas , Interferencia de ARN , Retroelementos , Linfocitos T/efectos de los fármacos , Linfocitos T/inmunología , Linfocitos T/virología , Factores de Tiempo , Factores de Transcripción/efectos de los fármacos , Factores de Transcripción/genética , Transcripción Genética/efectos de los fármacos , Transfección , Triazoles/farmacología , Latencia del Virus/efectos de los fármacosRESUMEN
The establishment of de novo chromatin accessibility in lymphoid progenitors requires the "pioneering" function of transcription factor (TF) early B cell factor 1 (EBF1), which binds to naïve chromatin and induces accessibility by recruiting the BRG1 chromatin remodeler subunit. However, it remains unclear whether the function of EBF1 is continuously required for stabilizing local chromatin accessibility. To this end, we replaced EBF1 by EBF1-FKBPF36V in pro-B cells, allowing the rapid degradation by adding the degradation TAG13 (dTAG13) dimerizer. EBF1 degradation results in a loss of genome-wide EBF1 occupancy and EBF1-targeted BRG1 binding. Chromatin accessibility was rapidly diminished at EBF1-binding sites with a preference for sites whose occupancy requires the pioneering activity of the C-terminal domain of EBF1. Diminished chromatin accessibility correlated with altered gene expression. Thus, continuous activity of EBF1 is required for the stable maintenance of the transcriptional and epigenetic state of pro-B cells.
Asunto(s)
Cromatina , Células Precursoras de Linfocitos B , Cromatina/genética , Epigenómica , Sitios de Unión , Recuento de CélulasRESUMEN
Strong evidence has indicated that upregulation of chemokine (CC motif) ligand-2 (CCL2) expression and the presence of an inflammatory tumor microenvironment significantly contribute to the migratory and invasive properties of oral squamous cell carcinoma, specifically oral tongue squamous cell carcinoma (OTSCC). However, the precise epigenetic mechanism responsible for enhanced CCL2 expression in response to the inflammatory mediator tumor necrosis factor alpha (TNF-α) in OTSCC remains inadequately elucidated. We have demonstrated that the production of CCL2 can be induced by TNF-α, and this induction is mediated by the chromatin remodel protein BRG1. Through the use of a chromatin immunoprecipitation (ChIP) assay, we have found that BRG1 was involved in the recruitment of acetylated histones H3 and H4 at the CCL2 promoter, thereby activating TNF-α-induced CCL2 transcription. Furthermore, we have observed that recruitment of NF-κB p65 to the CCL2 promoter was increased following BRG1 overexpression and decreased after BRG1 knockdown in OTSCC cells. Our Re-ChIP assay has shown that BRG1 knockdown completely inhibits the recruitment of both acetylated histone H3 or H4 and NF-κB to the CCL2 promoter. In summary, the findings of our study demonstrate that BRG1 plays a significant role in mediating the production of CCL2 in OTSCC cells in response to TNF-α stimulation. This process involves the cooperative action of acetylated histone and NF-κB recruitment to the CCL2 promoter site. Our data suggest that BRG1 serves as a critical epigenetic mediator in the regulation of TNF-α-induced CCL2 transcription in OTSCC cells.
Asunto(s)
Carcinoma de Células Escamosas , Neoplasias de Cabeza y Cuello , Neoplasias de la Lengua , Factor de Necrosis Tumoral alfa , Humanos , Carcinoma de Células Escamosas/genética , Quimiocina CCL2/metabolismo , Epigénesis Genética , Histonas/metabolismo , Neoplasias de la Boca , FN-kappa B/metabolismo , Carcinoma de Células Escamosas de Cabeza y Cuello , Neoplasias de la Lengua/genética , Microambiente Tumoral , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
During development, gene expression is tightly controlled to facilitate the generation of the diverse cell types that form the central nervous system. Brahma-related gene 1 (Brg1, also known as Smarca4) is the catalytic subunit of the SWItch/sucrose nonfermentable (SWI/SNF) chromatin remodeling complex that regulates transcription. We investigated the role of Brg1 between embryonic day 6.5 (E6.5) and E14.5 in Sox2-positive neural stem cells (NSCs). Being without major consequences at E6.5 and E14.5, loss of Brg1 between E7.5 and E12.5 resulted in the formation of rosette-like structures in the subventricular zone, as well as morphological alterations and enlargement of neural retina (NR). Additionally, Brg1-deficient cells showed decreased survival in vitro and in vivo. Furthermore, we uncovered distinct changes in gene expression upon Brg1 loss, pointing towards impaired neuron functions, especially those involving synaptic communication and altered composition of the extracellular matrix. Comparison with mice deficient for integrase interactor 1 (Ini1, also known as Smarcb1) revealed that the enlarged NR was Brg1 specific and was not caused by a general dysfunction of the SWI/SNF complex. These results suggest a crucial role for Brg1 in NSCs during brain and eye development.
Asunto(s)
Encéfalo/embriología , ADN Helicasas/genética , Ojo/embriología , Regulación del Desarrollo de la Expresión Génica/genética , Proteínas Nucleares/genética , Proteína SMARCB1/genética , Factores de Transcripción/genética , Animales , Apoptosis/genética , ADN Helicasas/metabolismo , Matriz Extracelular/genética , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Células-Madre Neurales/citología , Proteínas Nucleares/metabolismo , Factores de Transcripción/metabolismoRESUMEN
Inactivating alterations in the SWItch/Sucrose Non-Fermentable (SWI/SNF) Chromatin Remodeling Complex subunits have been described in multiple tumor types. Recent studies focused on SMARC subunits of this complex to understand their relationship with tumor characteristics and therapeutic opportunities. To date, pancreatic cancer with these alterations has not been well-studied, although isolated cases of undifferentiated carcinomas have been reported. Herein, we screened 59 pancreatic undifferentiated carcinomas for alterations in SWI/SNF complex-related [SMARCB1 (BAF47/INI1), SMARCA4 (BRG1), SMARCA2 (BRM)] proteins and/or genes using immunohistochemistry (IHC) and/or next-generation sequencing (NGS). Cases with alterations in SWI/SNF complex-related proteins/genes were compared to cases without alterations, as well as to 96 conventional pancreatic ductal adenocarcinomas (PDAC). In all tumor groups, MMR and PD-L1 protein expression were also evaluated. Thirty of 59 (51%) undifferentiated carcinomas had a loss of SWI/SNF complex-related protein expression or gene alteration. Twenty-seven of 30 (90%) SWI/SNF-deficient undifferentiated carcinomas had rhabdoid morphology [vs. 9/29 (31%) SWI/SNF-retained undifferentiated carcinomas; p < 0.001] and all expressed cytokeratin, at least focally. Immunohistochemically, SMARCB1 protein expression was absent in 16/30 (53%) cases, SMARCA2 in 4/30 (13%), and SMARCA4 in 4/30 (13%); both SMARCB1 and SMARCA2 protein expressions were absent in 1/30 (3%). Five of 8 (62.5%) SWI/SNF-deficient undifferentiated carcinomas that displayed loss of SMARCB1 protein expression by IHC were found to have corresponding SMARCB1 deletions by NGS. Analysis of canonical driver mutations for PDAC in these cases showed KRAS (2/5) and TP53 (2/5) abnormalities. Median CPS for PD-L1 (E1L3N) was significantly higher in the undifferentiated carcinomas with/without SWI/SNF deficiency compared to the conventional PDACs (p < 0.001). SWI/SNF-deficient undifferentiated carcinomas were larger (p < 0.001) and occurred in younger patients (p < 0.001). Patients with SWI/SNF-deficient undifferentiated carcinoma had worse overall survival compared to patients with SWI/SNF-retained undifferentiated carcinoma (p = 0.004) and PDAC (p < 0.001). Our findings demonstrate that SWI/SNF-deficient pancreatic undifferentiated carcinomas are frequently characterized by rhabdoid morphology, exhibit highly aggressive behavior, and have a negative prognostic impact. The ones with SMARCB1 deletions appear to be frequently KRAS wild-type. Innovative developmental therapeutic strategies targeting this genomic basis of the SWI/SNF complex and the therapeutic implications of EZH2 inhibition (NCT03213665), SMARCA2 degrader (NCT05639751), or immunotherapy are currently under investigation.
RESUMEN
SMARCA4 is a gene traditionally considered a tumor suppressor. Recent research has however found that SMARCA4 likely promotes cancer growth and is a good target for cancer treatment. The drug carbamazepine, an autophagy inducer, was used on colorectal cancer cell lines, HCT1116 and Hke3 (KRAS mutant and wildtype). Our study finds that Carbamazepine affects SMARCA4 levels and that this effect is different depending on the KRAS mutation status. This study analyzes the effect of carbamazepine on early-stage autophagy via ULK1 as well as simulates the docking of carbamazepine on KRAS, depending on the mutation status. Our study highlights the therapeutic uses of carbamazepine on cancer, and we propose that carbamazepine in conjunction with other chemotherapies may prove useful in targeting KRAS-mutated colorectal cancer.
Asunto(s)
Neoplasias Colorrectales , Proteínas Proto-Oncogénicas p21(ras) , Humanos , Proteínas Proto-Oncogénicas p21(ras)/genética , Mutación , Línea Celular , Neoplasias Colorrectales/tratamiento farmacológico , Neoplasias Colorrectales/genética , ADN Helicasas/genética , Proteínas Nucleares/genética , Factores de Transcripción/genéticaRESUMEN
Skin wound healing is driven by proliferation, migration and differentiation of several cell types that are controlled by the alterations in the gene expression programmes. Brahma Gene 1 (BRG1) (also known as SMARCA4) is a core ATPase in the BRG1 Associated Factors (BAF) ATP-dependent chromatin remodelling complexes that alter DNA-histone interaction in chromatin at the specific gene regulatory elements resulting in increase or decrease of the target gene transcription. Using siRNA mediated suppression of BRG1 during wound healing in a human ex vivo and in vitro (scratch assay) models, we demonstrated that BRG1 is essential for efficient skin wound healing by promoting epidermal keratinocytes migration, but not their proliferation or survival. BRG1 controls changes in the expression of genes associated with gene transcription, response to wounding, cell migration and cell signalling. Altogether, our data revealed that BRG1 play positive role in skin repair by promoting keratinocyte migration and impacting the genes expression programmes associated with cell migration and cellular signalling.
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Movimiento Celular , ADN Helicasas , Queratinocitos , Proteínas Nucleares , Transducción de Señal , Factores de Transcripción , Cicatrización de Heridas , Humanos , Queratinocitos/metabolismo , ADN Helicasas/metabolismo , ADN Helicasas/genética , Factores de Transcripción/metabolismo , Factores de Transcripción/genética , Proteínas Nucleares/metabolismo , Proteínas Nucleares/genética , Piel/metabolismo , Proliferación Celular , ARN Interferente PequeñoRESUMEN
BRG1 (Brahma-related gene 1) is a member of the SWI/SNF (switch/sucrose nonfermentable) chromatin remodeling complex which utilizes the energy from ATP hydrolysis for its activity. In addition to its role of regulating the expression of a vast array of genes, BRG1 mediates DNA repair upon genotoxic stress and regulates senescence. During organismal ageing, there is accumulation of unrepaired/unrepairable DNA damage due to progressive breakdown of the DNA repair machinery. The present study investigates the expression level of BRG1 as a function of age in the liver of 5- and 21-month-old female mice. It also explores the impact of dietary restriction on BRG1 expression in the old (21-month) mice. Salient findings of the study are: Real-time PCR and Western blot analyses reveal that BRG1 levels are higher in 5-month-old mice but decrease significantly with age. Dietary restriction increases BRG1 expression in the 21-month-old mice, nearly restoring it to the level observed in the younger group. Similar expression patterns are observed for DNA damage response genes ATM (Ataxia Telangiectasia Mutated) and ATR (Ataxia Telangiectasia and Rad3-related) with the advancement in age and which appears to be modulated by dietary restriction. BRG1 transcriptionally regulates ATM as a function of age and dietary restriction. These results suggest that BRG1, ATM and ATR are downregulated as mice age, and dietary restriction can restore their expression. This implies that dietary restriction may play a crucial role in regulating BRG1 and related gene expression, potentially maintaining liver repair and metabolic processes as mice age.
RESUMEN
Primary cutaneous SMARCA4-deficient undifferentiated malignant neoplasm (SD-UMN) is a rare and recently described entity characterized by the loss of expression of the SMARCA4 (BRG1) protein, which is involved in chromatin remodeling. SD-UMN presents a diagnostic challenge due to its rarity and unique histopathological and immunohistochemical features. In this report, we present a case of primary cutaneous SD-UMN in a 67-year-old man who presented with a rapidly growing, ulcerated, and bleeding nodule on his right cheek. Histopathological examination revealed a highly cellular dermal tumor consisting of pleomorphic epithelioid cells with prominent mitotic figures and necrosis, lacking any morphological evidence of differentiation. Immunohistochemical analysis showed a complete loss of SMARCA4 and SMARCA2 expression, while INI-1 expression remained intact. p53 was diffusely expressed, and p16 was completely absent. In addition, a range of markers, including high-molecular-weight cytokeratin, p63, SOX10, INSM1, MCPyV, NKX2.2, CD99, CDX2, CD56, ERG, NUT, desmin, androgen receptor, chromogranin, CD34, and CD43 were all negative. To date, only two cases of primary cutaneous SMARCA4-deficient undifferentiated tumors have been reported in the literature. Therefore, this case report adds to the limited body of knowledge on the clinical and histopathological features of this novel entity. The report highlights the importance of considering SD-UMN in the differential diagnosis of undifferentiated cutaneous tumors.
Asunto(s)
Carcinoma , Sarcoma , Masculino , Humanos , Anciano , Sarcoma/patología , Carcinoma/patología , Biomarcadores de Tumor/análisis , ADN Helicasas , Proteínas Nucleares , Factores de Transcripción , Proteínas RepresorasRESUMEN
Malignant ventricular arrhythmia (VA) after myocardial infarction (MI) is mainly caused by myocardial electrophysiological remodeling. Brahma-related gene 1 (BRG1) is an ATPase catalytic subunit that belongs to a family of chromatin remodeling complexes called Switch/Sucrose Non-Fermentable Chromatin (SWI/SNF). BRG1 has been reported as a molecular chaperone, interacting with various transcription factors or proteins to regulate transcription in cardiac diseases. In this study, we investigated the potential role of BRG1 in ion channel remodeling and VA after ischemic infarction. Myocardial infarction (MI) mice were established by ligating the left anterior descending (LAD) coronary artery, and electrocardiogram (ECG) was monitored. Epicardial conduction of MI mouse heart was characterized in Langendorff-perfused hearts using epicardial optical voltage mapping. Patch-clamping analysis was conducted in single ventricular cardiomyocytes isolated from the mice. We showed that BRG1 expression in the border zone was progressively increased in the first week following MI. Cardiac-specific deletion of BRG1 by tail vein injection of AAV9-BRG1-shRNA significantly ameliorated susceptibility to electrical-induced VA and shortened QTc intervals in MI mice. BRG1 knockdown significantly enhanced conduction velocity (CV) and reversed the prolonged action potential duration in MI mouse heart. Moreover, BRG1 knockdown improved the decreased densities of Na+ current (INa) and transient outward potassium current (Ito), as well as the expression of Nav1.5 and Kv4.3 in the border zone of MI mouse hearts and in hypoxia-treated neonatal mouse ventricular cardiomyocytes. We revealed that MI increased the binding among BRG1, T-cell factor 4 (TCF4) and ß-catenin, forming a transcription complex, which suppressed the transcription activity of SCN5A and KCND3, thereby influencing the incidence of VA post-MI.
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Infarto del Miocardio , Ratones , Animales , Infarto del Miocardio/metabolismo , Arritmias Cardíacas/genética , Miocardio/patología , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Miocitos Cardíacos/metabolismoRESUMEN
An abnormal differentiation state is common in BRCA1-deficient mammary epithelial cells, but the underlying mechanism is unclear. Here, we report a convergence between DNA repair and normal, cultured human mammary epithelial (HME) cell differentiation. Surprisingly, depleting BRCA1 or FANCD2 (Fanconi anemia [FA] proteins) or BRG1, a mSWI/SNF subunit, caused HME cells to undergo spontaneous epithelial-to-mesenchymal transition (EMT) and aberrant differentiation. This also occurred when wild-type HMEs were exposed to chemicals that generate DNA interstrand crosslinks (repaired by FA proteins), but not in response to double-strand breaks. Suppressed expression of ΔNP63 also occurred in each of these settings, an effect that links DNA damage to the aberrant differentiation outcome. Taken together with somatic breast cancer genome data, these results point to a breakdown in a BRCA/FA-mSWI/SNF-ΔNP63-mediated DNA repair and differentiation maintenance process in mammary epithelial cells that may contribute to sporadic breast cancer development.
Asunto(s)
Proteína BRCA1/metabolismo , Neoplasias de la Mama/prevención & control , Diferenciación Celular , Daño del ADN , ADN Helicasas/metabolismo , Reparación del ADN , Células Epiteliales/metabolismo , Proteína del Grupo de Complementación D2 de la Anemia de Fanconi/metabolismo , Glándulas Mamarias Humanas/metabolismo , Proteínas Nucleares/metabolismo , Factores de Transcripción/metabolismo , Acetaldehído/farmacología , Proteína BRCA1/genética , Neoplasias de la Mama/genética , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Línea Celular , Transformación Celular Neoplásica/genética , Transformación Celular Neoplásica/metabolismo , Transformación Celular Neoplásica/patología , Cisplatino/farmacología , ADN Helicasas/genética , Células Epiteliales/efectos de los fármacos , Células Epiteliales/patología , Proteína del Grupo de Complementación D2 de la Anemia de Fanconi/genética , Femenino , Formaldehído/farmacología , Humanos , Glándulas Mamarias Humanas/efectos de los fármacos , Glándulas Mamarias Humanas/patología , Mutación , Proteínas Nucleares/genética , Fenotipo , Interferencia de ARN , Transducción de Señal , Factores de Transcripción/genética , Transfección , Proteínas Supresoras de Tumor/genética , Proteínas Supresoras de Tumor/metabolismoRESUMEN
Epigenetics is the study of heritable changes to the genome and gene expression patterns that are not caused by direct changes to the DNA sequence. Examples of these changes include posttranslational modifications to DNA-bound histone proteins, DNA methylation, and remodeling of nuclear architecture. Collectively, epigenetic changes provide a layer of regulation that affects transcriptional activity of genes while leaving DNA sequences unaltered. Sequence variants or mutations affecting enzymes responsible for modifying or sensing epigenetic marks have been identified in patients with congenital heart disease (CHD), and small-molecule inhibitors of epigenetic complexes have shown promise as therapies for adult heart diseases. Additionally, transgenic mice harboring mutations or deletions of genes encoding epigenetic enzymes recapitulate aspects of human cardiac disease. Taken together, these findings suggest that the evolving field of epigenetics will inform our understanding of congenital and adult cardiac disease and offer new therapeutic opportunities.