RESUMO
The estrogen receptor (ER), glucocorticoid receptor (GR), and forkhead box protein 1 (FoxA1) are significant factors in breast cancer progression. FoxA1 has been implicated in establishing ER-binding patterns though its unique ability to serve as a pioneer factor. However, the molecular interplay between ER, GR, and FoxA1 requires further investigation. Here we show that ER and GR both have the ability to alter the genomic distribution of the FoxA1 pioneer factor. Single-molecule tracking experiments in live cells reveal a highly dynamic interaction of FoxA1 with chromatin in vivo. Furthermore, the FoxA1 factor is not associated with detectable footprints at its binding sites throughout the genome. These findings support a model wherein interactions between transcription factors and pioneer factors are highly dynamic. Moreover, at a subset of genomic sites, the role of pioneer can be reversed, with the steroid receptors serving to enhance binding of FoxA1.
Assuntos
Fator 3-alfa Nuclear de Hepatócito/metabolismo , Cromatina/metabolismo , Desoxirribonucleases/metabolismo , Humanos , Células MCF-7 , Receptores de Estrogênio/genética , Receptores de Glucocorticoides/genética , Fatores de Transcrição/metabolismoRESUMO
Transcription factors (TFs) regulate gene expression by binding to specific consensus motifs within the local chromatin context. The mechanisms by which TFs navigate the nuclear environment as they search for binding sites remain unclear. Here, we used single-molecule tracking and machine-learning-based classification to directly measure the nuclear mobility of the glucocorticoid receptor (GR) in live cells. We revealed two distinct and dynamic low-mobility populations. One accounts for specific binding to chromatin, while the other represents a confinement state that requires an intrinsically disordered region (IDR), implicated in liquid-liquid condensate subdomains. Further analysis showed that the dwell times of both subpopulations follow a power-law distribution, consistent with a broad distribution of affinities on the GR cistrome and interactome. Together, our data link IDRs with a confinement state that is functionally distinct from specific chromatin binding and modulates the transcriptional output by increasing the local concentration of TFs at specific sites.
Assuntos
Proteínas Intrinsicamente Desordenadas/química , Receptores de Glucocorticoides/química , Fatores de Transcrição/química , Animais , Feminino , Proteínas Intrinsicamente Desordenadas/genética , Proteínas Intrinsicamente Desordenadas/metabolismo , Camundongos , Ratos , Receptores de Glucocorticoides/genética , Receptores de Glucocorticoides/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
Treatment of prostate cancer relies predominantly on the inhibition of androgen receptor (AR) signaling. Despite the initial effectiveness of the antiandrogen therapies, the cancer often develops resistance to the AR blockade. One mechanism of the resistance is glucocorticoid receptor (GR)-mediated replacement of AR function. Nevertheless, the mechanistic ways and means how the GR-mediated antiandrogen resistance occurs have remained elusive. Here, we have discovered several crucial features of GR action in prostate cancer cells through genome-wide techniques. We detected that the replacement of AR by GR in enzalutamide-exposed prostate cancer cells occurs almost exclusively at pre-accessible chromatin sites displaying FOXA1 occupancy. Counterintuitively to the classical pioneer factor model, silencing of FOXA1 potentiated the chromatin binding and transcriptional activity of GR. This was attributed to FOXA1-mediated repression of the NR3C1 (gene encoding GR) expression via the corepressor TLE3. Moreover, the small-molecule inhibition of coactivator p300's enzymatic activity efficiently restricted GR-mediated gene regulation and cell proliferation. Overall, we identified chromatin pre-accessibility and FOXA1-mediated repression as important regulators of GR action in prostate cancer, pointing out new avenues to oppose steroid receptor-mediated antiandrogen resistance.
Assuntos
Cromatina , Neoplasias da Próstata , Receptores de Glucocorticoides , Humanos , Masculino , Antagonistas de Androgênios/farmacologia , Linhagem Celular Tumoral , Cromatina/genética , Regulação Neoplásica da Expressão Gênica , Fator 3-alfa Nuclear de Hepatócito/genética , Fator 3-alfa Nuclear de Hepatócito/metabolismo , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/genética , Neoplasias da Próstata/metabolismo , Receptores Androgênicos/genética , Receptores Androgênicos/metabolismo , Receptores de Glucocorticoides/genética , Receptores de Glucocorticoides/metabolismoRESUMO
The androgen receptor (AR) is pivotal in prostate cancer (PCa) progression and represents a critical therapeutic target. AR-mediated gene regulation involves intricate interactions with nuclear proteins, with many mediating and undergoing post-translational modifications that present alternative therapeutic avenues. Through chromatin proteomics in PCa cells, we identified SUMO ligases together with nuclear receptor coregulators and pioneer transcription factors within the AR's protein network. Intriguingly, this network displayed a significant association with SUMO2/3. To elucidate the influence of SUMOylation on AR chromatin interactions and subsequent gene regulation, we inhibited SUMOylation using ML-792 (SUMOi). While androgens generally facilitated the co-occupancy of SUMO2/3 and AR on chromatin, SUMOi induced divergent effects dependent on the type of AR-binding site (ARB). SUMOi augmented AR's pioneer-like binding on inaccessible chromatin regions abundant in androgen response elements (AREs) and diminished its interaction with accessible chromatin regions sparse in AREs yet rich in pioneer transcription factor motifs. The SUMOi-impacted ARBs divergently influenced AR-regulated genes; those associated with AR-mediated activation played roles in negative regulation of cell proliferation, while those with AR-mediated repression were involved in pattern formation. In conclusion, our findings underscore the pervasive influence of SUMOylation in shaping AR's role in PCa cells, potentially unveiling new therapeutic strategies.
RESUMO
The use of androgen receptor (AR) inhibitors in prostate cancer gives rise to increased cellular lineage plasticity resulting in resistance to AR-targeted therapies. In this study, we examined the chromatin landscape of AR-positive prostate cancer cells post-exposure to the AR inhibitor enzalutamide. We identified a novel regulator of cell plasticity, the homeobox transcription factor SIX2, whose motif is enriched in accessible chromatin regions after treatment. Depletion of SIX2 in androgen-independent PC-3 prostate cancer cells induced a switch from a stem-like to an epithelial state, resulting in reduced cancer-related properties such as proliferation, colony formation, and metastasis both in vitro and in vivo. These effects were mediated through the downregulation of the Wnt/ß-catenin signalling pathway and subsequent reduction of nuclear ß-catenin. Collectively, our findings provide compelling evidence that the depletion of SIX2 may represent a promising strategy for overcoming the cell plasticity mechanisms driving antiandrogen resistance in prostate cancer.
Assuntos
Benzamidas , Plasticidade Celular , Proteínas de Homeodomínio , Nitrilas , Feniltioidantoína , Neoplasias da Próstata , Receptores Androgênicos , Via de Sinalização Wnt , beta Catenina , Animais , Humanos , Masculino , Camundongos , Benzamidas/farmacologia , beta Catenina/metabolismo , beta Catenina/genética , Linhagem Celular Tumoral , Plasticidade Celular/genética , Proliferação de Células/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/genética , Regulação Neoplásica da Expressão Gênica , Proteínas de Homeodomínio/metabolismo , Proteínas de Homeodomínio/genética , Proteínas do Tecido Nervoso/metabolismo , Proteínas do Tecido Nervoso/genética , Nitrilas/farmacologia , Células PC-3 , Feniltioidantoína/farmacologia , Feniltioidantoína/análogos & derivados , Neoplasias da Próstata/genética , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Neoplasias da Próstata/tratamento farmacológico , Receptores Androgênicos/metabolismo , Receptores Androgênicos/genética , Via de Sinalização Wnt/efeitos dos fármacosRESUMO
The androgen receptor (AR) is a primary target for treating prostate cancer (PCa), forming the bedrock of its clinical management. Despite their efficacy, resistance often hampers AR-targeted therapies, necessitating new strategies against therapy-resistant PCa. These resistances involve various mechanisms, including AR splice variant overexpression and altered activities of transcription factors like the glucocorticoid receptor (GR) and FOXA1. These factors rely on common coregulators, such as EP300/CREBBP, suggesting a rationale for coregulator-targeted therapies. Our study explores EP300/CREBBP acetyltransferase inhibition's impact on steroid receptor and FOXA1 signaling in PCa cells using genome-wide techniques. Results reveal that EP300/CREBBP inhibition significantly disrupts the AR-regulated transcriptome and receptor chromatin binding by reducing the AR-gene expression. Similarly, GR's regulated transcriptome and receptor binding were hindered, not linked to reduced GR expression but to diminished FOXA1 chromatin binding, restricting GR signaling. Overall, our findings highlight how EP300/CREBBP inhibition distinctively curtails oncogenic transcription factors' signaling, suggesting the potential of coregulatory-targeted therapies in PCa.
Assuntos
Próstata , Neoplasias da Próstata , Masculino , Humanos , Neoplasias da Próstata/genética , Receptores de Glucocorticoides/genética , Fatores de Transcrição , Cromatina , Acetiltransferases , Fator 3-alfa Nuclear de Hepatócito/genética , Proteína p300 Associada a E1A/genética , Proteína de Ligação a CREB/genéticaRESUMO
Here, we investigate the impact of hypoxia on the hepatic response of glucocorticoid receptor (GR) to dexamethasone (DEX) in mice via RNA-sequencing. Hypoxia causes three types of reprogramming of GR: (i) much weaker induction of classical GR-responsive genes by DEX in hypoxia, (ii) a number of genes is induced by DEX specifically in hypoxia, and (iii) hypoxia induces a group of genes via activation of the hypothalamic-pituitary-adrenal (HPA) axis. Transcriptional profiles are reflected by changed GR DNA-binding as measured by ChIP sequencing. The HPA axis is induced by hypothalamic HIF1α and HIF2α activation and leads to GR-dependent lipolysis and ketogenesis. Acute inflammation, induced by lipopolysaccharide, is prevented by DEX in normoxia but not during hypoxia, and this is attributed to HPA axis activation by hypoxia. We unfold new physiological pathways that have consequences for patients suffering from GC resistance.
Assuntos
Glucocorticoides , Receptores de Glucocorticoides , Animais , Dexametasona/metabolismo , Dexametasona/farmacologia , Glucocorticoides/metabolismo , Glucocorticoides/farmacologia , Humanos , Sistema Hipotálamo-Hipofisário/metabolismo , Hipóxia/genética , Hipóxia/metabolismo , Camundongos , Sistema Hipófise-Suprarrenal/metabolismo , Receptores de Glucocorticoides/genética , Receptores de Glucocorticoides/metabolismoRESUMO
Single-molecule tracking (SMT) allows the study of transcription factor (TF) dynamics in the nucleus, giving important information regarding the diffusion and binding behavior of these proteins in the nuclear environment. Dwell time distributions obtained by SMT for most TFs appear to follow bi-exponential behavior. This has been ascribed to two discrete populations of TFs-one non-specifically bound to chromatin and another specifically bound to target sites, as implied by decades of biochemical studies. However, emerging studies suggest alternate models for dwell-time distributions, indicating the existence of more than two populations of TFs (multi-exponential distribution), or even the absence of discrete states altogether (power-law distribution). Here, we present an analytical pipeline to evaluate which model best explains SMT data. We find that a broad spectrum of TFs (including glucocorticoid receptor, oestrogen receptor, FOXA1, CTCF) follow a power-law distribution of dwell-times, blurring the temporal line between non-specific and specific binding, suggesting that productive binding may involve longer binding events than previously believed. From these observations, we propose a continuum of affinities model to explain TF dynamics, that is consistent with complex interactions of TFs with multiple nuclear domains as well as binding and searching on the chromatin template.
Assuntos
Fatores de Transcrição/metabolismo , Animais , Linhagem Celular Tumoral , Cinética , Camundongos , Modelos Biológicos , Fotodegradação , Ligação Proteica , Receptores de Glucocorticoides/metabolismo , Imagem Individual de MoléculaRESUMO
Glucocorticoid receptor (GR) is an essential transcription factor (TF), controlling metabolism, development and immune responses. SUMOylation regulates chromatin occupancy and target gene expression of GR in a locus-selective manner, but the mechanism of regulation has remained elusive. Here, we identify the protein network around chromatin-bound GR by using selective isolation of chromatin-associated proteins and show that the network is affected by receptor SUMOylation, with several nuclear receptor coregulators and chromatin modifiers preferring interaction with SUMOylation-deficient GR and proteins implicated in transcriptional repression preferring interaction with SUMOylation-competent GR. This difference is reflected in our chromatin binding, chromatin accessibility and gene expression data, showing that the SUMOylation-deficient GR is more potent in binding and opening chromatin at glucocorticoid-regulated enhancers and inducing expression of target loci. Blockage of SUMOylation by a SUMO-activating enzyme inhibitor (ML-792) phenocopied to a large extent the consequences of GR SUMOylation deficiency on chromatin binding and target gene expression. Our results thus show that SUMOylation modulates the specificity of GR by regulating its chromatin protein network and accessibility at GR-bound enhancers. We speculate that many other SUMOylated TFs utilize a similar regulatory mechanism.
Assuntos
Cromatina/metabolismo , Receptores de Glucocorticoides/metabolismo , Sumoilação , Sítios de Ligação , Regulação da Expressão Gênica , Células HEK293 , Humanos , Correpressor 1 de Receptor Nuclear/metabolismo , Coativador 1 de Receptor Nuclear , Mapeamento de Interação de Proteínas , Proteínas Modificadoras Pequenas Relacionadas à Ubiquitina/metabolismo , Sumoilação/efeitos dos fármacosRESUMO
Most transcription factors, including nuclear receptors, are widely modeled as binding regulatory elements as monomers, homodimers, or heterodimers. Recent findings in live cells show that the glucocorticoid receptor NR3C1 (also known as GR) forms tetramers on enhancers, owing to an allosteric alteration induced by DNA binding, and suggest that higher oligomerization states are important for the gene regulatory responses of GR. By using a variant (GRtetra) that mimics this allosteric transition, we performed genome-wide studies using a GR knockout cell line with reintroduced wild-type GR or reintroduced GRtetra. GRtetra acts as a super receptor by binding to response elements not accessible to the wild-type receptor and both induces and represses more genes than GRwt. These results argue that DNA binding induces a structural transition to the tetrameric state, forming a transient higher-order structure that drives both the activating and repressive actions of glucocorticoids.
Assuntos
Cromatina/ultraestrutura , Células Epiteliais/efeitos dos fármacos , Genoma , Glucocorticoides/farmacologia , RNA Mensageiro/genética , Receptores de Glucocorticoides/química , Animais , Sequência de Bases , Sistemas CRISPR-Cas , Linhagem Celular Tumoral , Cromatina/química , DNA/genética , DNA/metabolismo , Elementos Facilitadores Genéticos , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Feminino , Edição de Genes/métodos , Glucocorticoides/metabolismo , Sequenciamento de Nucleotídeos em Larga Escala , Glândulas Mamárias Animais/metabolismo , Glândulas Mamárias Animais/patologia , Camundongos , Ligação Proteica , Estrutura Quaternária de Proteína , RNA Mensageiro/metabolismo , Ratos , Receptores de Glucocorticoides/genética , Receptores de Glucocorticoides/metabolismo , Ativação TranscricionalRESUMO
Glucocorticoid resistance (GCR) is defined as an unresponsiveness to the therapeutic effects, including the antiinflammatory ones of glucocorticoids (GCs) and their receptor, the glucocorticoid receptor (GR). It is a problem in the management of inflammatory diseases and can be congenital as well as acquired. The strong proinflammatory cytokine TNF-alpha (TNF) induces an acute form of GCR, not only in mice, but also in several cell lines: e.g., in the hepatoma cell line BWTG3, as evidenced by impaired Dexamethasone (Dex)-stimulated direct GR-dependent gene up- and down-regulation. We report that TNF has a significant and broad impact on this transcriptional performance of GR, but no impact on nuclear translocation, dimerization, or DNA binding capacity of GR. Proteome-wide proximity-mapping (BioID), however, revealed that the GR interactome was strongly modulated by TNF. One GR cofactor that interacted significantly less with the receptor under GCR conditions is p300. NFκB activation and p300 knockdown both reduced direct transcriptional output of GR whereas p300 overexpression and NFκB inhibition reverted TNF-induced GCR, which is in support of a cofactor reshuffle model. This hypothesis was supported by FRET studies. This mechanism of GCR opens avenues for therapeutic interventions in GCR diseases.
Assuntos
Resistência a Medicamentos/genética , Proteína p300 Associada a E1A/metabolismo , Glucocorticoides/farmacologia , Inflamação/tratamento farmacológico , Receptores de Glucocorticoides/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Células A549 , Animais , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Dexametasona/farmacologia , Dexametasona/uso terapêutico , Regulação para Baixo/efeitos dos fármacos , Regulação para Baixo/imunologia , Proteína p300 Associada a E1A/genética , Feminino , Técnicas de Silenciamento de Genes , Glucocorticoides/uso terapêutico , Células HEK293 , Humanos , Inflamação/imunologia , Camundongos , NF-kappa B/metabolismo , Mapeamento de Interação de Proteínas , Mapas de Interação de Proteínas/efeitos dos fármacos , Mapas de Interação de Proteínas/imunologia , RNA Interferente Pequeno/metabolismo , RNA-Seq , Receptores de Glucocorticoides/imunologia , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/imunologiaRESUMO
Fasting elicits transcriptional programs in hepatocytes leading to glucose and ketone production. This transcriptional program is regulated by many transcription factors (TFs). To understand how this complex network regulates the metabolic response to fasting, we aimed at isolating the enhancers and TFs dictating it. Measuring chromatin accessibility revealed that fasting massively reorganizes liver chromatin, exposing numerous fasting-induced enhancers. By utilizing computational methods in combination with dissecting enhancer features and TF cistromes, we implicated four key TFs regulating the fasting response: glucocorticoid receptor (GR), cAMP responsive element binding protein 1 (CREB1), peroxisome proliferator activated receptor alpha (PPARA), and CCAAT/enhancer binding protein beta (CEBPB). These TFs regulate fuel production by two distinctly operating modules, each controlling a separate metabolic pathway. The gluconeogenic module operates through assisted loading, whereby GR doubles the number of sites occupied by CREB1 as well as enhances CREB1 binding intensity and increases accessibility of CREB1 binding sites. Importantly, this GR-assisted CREB1 binding was enhancer-selective and did not affect all CREB1-bound enhancers. Single-molecule tracking revealed that GR increases the number and DNA residence time of a portion of chromatin-bound CREB1 molecules. These events collectively result in rapid synergistic gene expression and higher hepatic glucose production. Conversely, the ketogenic module operates via a GR-induced TF cascade, whereby PPARA levels are increased following GR activation, facilitating gradual enhancer maturation next to PPARA target genes and delayed ketogenic gene expression. Our findings reveal a complex network of enhancers and TFs that dynamically cooperate to restore homeostasis upon fasting.
Assuntos
Proteína beta Intensificadora de Ligação a CCAAT/metabolismo , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Elementos Facilitadores Genéticos , Jejum/metabolismo , Hepatócitos/metabolismo , PPAR alfa/metabolismo , Receptores de Glucocorticoides/metabolismo , Animais , Sítios de Ligação , Proteína beta Intensificadora de Ligação a CCAAT/genética , Cromatina/genética , Cromatina/metabolismo , Montagem e Desmontagem da Cromatina , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/genética , Glucose/metabolismo , Cetonas/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , PPAR alfa/genética , Receptores de Glucocorticoides/genética , Ativação TranscricionalRESUMO
Progressive, technological achievements in the quantitative fluorescence microscopy field are allowing researches from many different areas to start unraveling the dynamic intricacies of biological processes inside living cells. From super-resolution microscopy techniques to tracking of individual proteins, fluorescence microscopy is changing our perspective on how the cell works. Fortunately, a growing number of research groups are exploring single-molecule studies in living cells. However, no clear consensus exists on several key aspects of the technique such as image acquisition conditions, or analysis of the obtained data. Here, we describe a detailed approach to perform single-molecule tracking (SMT) of transcription factors in living cells to obtain key binding characteristics, namely their residence time and bound fractions. We discuss different types of fluorophores, labeling density, microscope, cameras, data acquisition, and data analysis. Using the glucocorticoid receptor as a model transcription factor, we compared alternate tags (GFP, mEOS, HaloTag, SNAP-tag, CLIP-tag) for potential multicolor applications. We also examine different methods to extract the dissociation rates and compare them with simulated data. Finally, we discuss several challenges that this exciting technique still faces.
Assuntos
Células Epiteliais/metabolismo , Processamento de Imagem Assistida por Computador/estatística & dados numéricos , Receptores de Glucocorticoides/genética , Imagem Individual de Molécula/métodos , Animais , Antígenos de Diferenciação de Linfócitos B/genética , Antígenos de Diferenciação de Linfócitos B/metabolismo , Linhagem Celular Tumoral , Células Epiteliais/ultraestrutura , Regulação da Expressão Gênica , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Células Hep G2 , Antígenos de Histocompatibilidade Classe II/genética , Antígenos de Histocompatibilidade Classe II/metabolismo , Humanos , Cinética , Células MCF-7 , Camundongos , Ligação Proteica , Receptores de Glucocorticoides/metabolismo , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismoRESUMO
Transcription factor (TF) signaling regulates gene transcription and requires a complex network of proteins. This network includes co-activators, co-repressors, multiple TFs, histone-modifying complexes, and the basal transcription machinery. It has been widely appreciated that pioneer factors, such as FoxA1 and GATA1, play an important role in opening closed chromatin regions, thereby allowing binding of a secondary factor. In this review we will focus on a newly proposed model wherein multiple TFs, such as steroid receptors (SRs), can function in a pioneering role. This model, termed dynamic assisted loading, integrates data from widely divergent methodologies, including genome wide ChIP-Seq, digital genomic footprinting, DHS-Seq, live cell protein dynamics, and biochemical studies of ATP-dependent remodeling complexes, to present a real time view of TF chromatin interactions. Under this view, many TFs can act as initiating factors for chromatin landscape programming. Furthermore, enhancer and promoter states are more accurately described as energy-dependent, non-equilibrium steady states.
Assuntos
Trifosfato de Adenosina/metabolismo , Montagem e Desmontagem da Cromatina , Modelos Genéticos , Fatores de Transcrição/metabolismo , Animais , Elementos Facilitadores Genéticos , Humanos , Regiões Promotoras GenéticasRESUMO
In addition to the glucocorticoids, the glucocorticoid receptor (GR) is regulated by post-translational modifications, including SUMOylation. We have analyzed how SUMOylation influences the activity of endogenous GR target genes and the receptor chromatin binding by using isogenic HEK293 cells expressing wild-type GR (wtGR) or SUMOylation-defective GR (GR3KR). Gene expression profiling revealed that both dexamethasone up- and downregulated genes are affected by the GR SUMOylation and that the affected genes are significantly associated with pathways of cellular proliferation and survival. The GR3KR-expressing cells proliferated more rapidly, and their anti-proliferative response to dexamethasone was less pronounced than in the wtGR-expressing cells. ChIP-seq analyses indicated that the SUMOylation modulates the chromatin occupancy of GR on several loci associated with cellular growth in a fashion that parallels with their differential dexamethasone-regulated expression between the two cell lines. Moreover, chromatin SUMO-2/3 marks, which were associated with active GR-binding sites, showed markedly higher overlap with the wtGR cistrome than with the GR3KR cistrome. In sum, our results indicate that the SUMOylation does not simply repress the GR activity, but regulates the activity of the receptor in a target locus selective fashion, playing an important role in controlling the GR activity on genes influencing cell growth.
Assuntos
Proliferação de Células , Cromatina/metabolismo , Regulação da Expressão Gênica , Receptores de Glucocorticoides/metabolismo , Sumoilação , Proliferação de Células/efeitos dos fármacos , Dexametasona/farmacologia , Células HEK293 , Humanos , Proteínas Modificadoras Pequenas Relacionadas à Ubiquitina/metabolismoRESUMO
Prostate cancer is one of the most prevalent malignancies and is primarily driven by aberrant androgen receptor (AR) signaling. While AR-targeted therapies form the cornerstone of prostate cancer treatment, they often inadvertently activate compensatory pathways, leading to therapy resistance. This resistance is frequently mediated through changes in transcription factor (TF) crosstalk, reshaping gene regulatory programs and ultimately weakening treatment efficacy. Consequently, investigating TF interactions has become crucial for understanding the mechanisms driving therapy-resistant cancers. Recent evidence has highlighted the crosstalk between the glucocorticoid receptor (GR) and AR, demonstrating that GR can induce prostate cancer therapy resistance by replacing the inactivated AR, thereby becoming a driver of the disease. In addition to this oncogenic role, GR has also been shown to act as a tumor suppressor in prostate cancer. Owing to this dual role and the widespread use of glucocorticoids as adjuvant therapy, it is essential to understand GR's actions across different stages of prostate cancer development. In this review, we explore the current knowledge of GR in prostate cancer, with a specific focus on its crosstalk with other TFs. GR can directly and indirectly interact with a variety of TFs, and these interactions vary significantly depending on the type of prostate cancer cells. By highlighting these crosstalk interactions, we aim to provide insights that can guide the research and development of new GR-targeted therapies to mitigate its harmful effects in prostate cancer.
Assuntos
Neoplasias da Próstata , Receptores de Glucocorticoides , Fatores de Transcrição , Humanos , Receptores de Glucocorticoides/metabolismo , Receptores de Glucocorticoides/genética , Masculino , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/genética , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/patologia , Fatores de Transcrição/metabolismo , Fatores de Transcrição/genética , Receptores Androgênicos/metabolismo , Receptores Androgênicos/genética , Transdução de Sinais , Animais , Receptor Cross-Talk/fisiologia , Regulação Neoplásica da Expressão GênicaRESUMO
The inflammatory tumor microenvironment (TME) is a key driver for tumor-promoting processes. Tumor-associated macrophages are one of the main immune cell types in the TME and their increased density is related to poor prognosis in prostate cancer. Here, we investigated the influence of pro-inflammatory (M1) and immunosuppressive (M2) macrophages on prostate cancer lineage plasticity. Our findings reveal that M1 macrophage secreted factors upregulate genes related to stemness while downregulating genes associated with androgen response in prostate cancer cells. The expression of cancer stem cell (CSC) plasticity markers NANOG, KLF4, SOX2, OCT4, and CD44 was stimulated by the secreted factors from M1 macrophages. Moreover, AR and its target gene PSA were observed to be suppressed in LNCaP cells treated with secreted factors from M1 macrophages. Inhibition of NFκB signaling using the IKK16 inhibitor resulted in downregulation of NANOG, SOX2, and CD44 and CSC plasticity. Our study highlights that the secreted factors from M1 macrophages drive prostate cancer cell plasticity by upregulating the expression of CSC plasticity markers through NFκB signaling pathway.
Assuntos
Receptores de Hialuronatos , Fator 4 Semelhante a Kruppel , Macrófagos , NF-kappa B , Proteína Homeobox Nanog , Células-Tronco Neoplásicas , Neoplasias da Próstata , Fatores de Transcrição SOXB1 , Transdução de Sinais , Masculino , Humanos , Neoplasias da Próstata/patologia , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/genética , Proteína Homeobox Nanog/metabolismo , Proteína Homeobox Nanog/genética , Fatores de Transcrição SOXB1/metabolismo , Fatores de Transcrição SOXB1/genética , Receptores de Hialuronatos/metabolismo , Receptores de Hialuronatos/genética , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Fator 4 Semelhante a Kruppel/metabolismo , NF-kappa B/metabolismo , Linhagem Celular Tumoral , Macrófagos/metabolismo , Regulação para Cima , Microambiente Tumoral/imunologia , Plasticidade Celular/genética , Regulação Neoplásica da Expressão Gênica , Macrófagos Associados a Tumor/metabolismo , Macrófagos Associados a Tumor/imunologia , Animais , CamundongosRESUMO
Treatment-induced neuroendocrine prostate cancer (t-NEPC) is a lethal subtype of castration-resistant prostate cancer resistant to androgen receptor (AR) inhibitors. Our study unveils that AR suppresses the neuronal development protein dihydropyrimidinase-related protein 5 (DPYSL5), providing a mechanism for neuroendocrine transformation under androgen deprivation therapy. Our unique CRPC-NEPC cohort, comprising 135 patient tumor samples, including 55 t-NEPC patient samples, exhibits a high expression of DPYSL5 in t-NEPC patient tumors. DPYSL5 correlates with neuroendocrine-related markers and inversely with AR and PSA. DPYSL5 overexpression in prostate cancer cells induces a neuron-like phenotype, enhances invasion, proliferation, and upregulates stemness and neuroendocrine-related markers. Mechanistically, DPYSL5 promotes prostate cancer cell plasticity via EZH2-mediated PRC2 activation. Depletion of DPYSL5 decreases proliferation, induces G1 phase cell cycle arrest, reverses neuroendocrine phenotype, and upregulates luminal genes. In conclusion, DPYSL5 plays a critical role in regulating prostate cancer cell plasticity, and we propose the AR/DPYSL5/EZH2/PRC2 axis as a driver of t-NEPC progression.
Assuntos
Neoplasias da Próstata , Masculino , Humanos , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/genética , Neoplasias da Próstata/metabolismo , Antagonistas de Androgênios , Próstata/patologia , Hidrolases , Proteínas Associadas aos Microtúbulos , Proteína Potenciadora do Homólogo 2 de Zeste/genéticaRESUMO
Prostate cancer is one of the leading causes of death among men worldwide, and thus, research on the genetic factors enabling the formation of treatment-resistant cancer cells is crucial for improving patient outcomes. Here, we report a cell line-specific dependence on FANCI and related signaling pathways to counteract the effects of DNA-damaging chemotherapy in prostate cancer. Our results reveal that FANCI depletion results in significant downregulation of Fanconi anemia (FA) pathway members in prostate cancer cells, indicating that FANCI is an important regulator of the FA pathway. Furthermore, we found that FANCI silencing reduces proliferation in p53-expressing prostate cancer cells. This extends the evidence that inactivation of FANCI may convert cancer cells from a resistant state to an eradicable state under the stress of DNA-damaging chemotherapy. Our results also indicate that high expression of FA pathway genes correlates with poorer survival in prostate cancer patients. Moreover, genomic alterations of FA pathway members are prevalent in prostate adenocarcinoma patients; mutation and copy number information for the FA pathway genes in seven patient cohorts (N = 1,732 total tumor samples) reveals that 1,025 (59.2%) tumor samples have an alteration in at least one of the FA pathway genes, suggesting that genomic alteration of the pathway is a prominent feature in patients with the disease.
RESUMO
How chromatin dynamics relate to transcriptional activity remains poorly understood. Using single-molecule tracking, coupled with machine learning, we show that histone H2B and multiple chromatin-bound transcriptional regulators display two distinct low-mobility states. Ligand activation results in a marked increase in the propensity of steroid receptors to bind in the lowest-mobility state. Mutational analysis revealed that interactions with chromatin in the lowest-mobility state require an intact DNA binding domain and oligomerization domains. These states are not spatially separated as previously believed, but individual H2B and bound-TF molecules can dynamically switch between them on time scales of seconds. Single bound-TF molecules with different mobilities exhibit different dwell time distributions, suggesting that the mobility of TFs is intimately coupled with their binding dynamics. Together, our results identify two unique and distinct low-mobility states that appear to represent common pathways for transcription activation in mammalian cells.