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1.
Nat Commun ; 12(1): 5056, 2021 08 20.
Artigo em Inglês | MEDLINE | ID: mdl-34417458

RESUMO

Melanoma cells rely on developmental programs during tumor initiation and progression. Here we show that the embryonic stem cell (ESC) factor Sall4 is re-expressed in the Tyr::NrasQ61K; Cdkn2a-/- melanoma model and that its expression is necessary for primary melanoma formation. Surprisingly, while Sall4 loss prevents tumor formation, it promotes micrometastases to distant organs in this melanoma-prone mouse model. Transcriptional profiling and in vitro assays using human melanoma cells demonstrate that SALL4 loss induces a phenotype switch and the acquisition of an invasive phenotype. We show that SALL4 negatively regulates invasiveness through interaction with the histone deacetylase (HDAC) 2 and direct co-binding to a set of invasiveness genes. Consequently, SALL4 knock down, as well as HDAC inhibition, promote the expression of an invasive signature, while inhibition of histone acetylation partially reverts the invasiveness program induced by SALL4 loss. Thus, SALL4 appears to regulate phenotype switching in melanoma through an HDAC2-mediated mechanism.


Assuntos
Epigênese Genética , Melanoma/genética , Melanoma/patologia , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/patologia , Fator de Células-Tronco/metabolismo , Fatores de Transcrição/metabolismo , Acetilação , Animais , Sequência de Bases , Carcinogênese/genética , Carcinogênese/patologia , Adesão Celular/genética , Linhagem Celular Tumoral , Linhagem da Célula , Proliferação de Células , Proteínas de Ligação a DNA/metabolismo , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Histona Desacetilase 2/metabolismo , Histonas/metabolismo , Humanos , Melanócitos/metabolismo , Melanócitos/patologia , Camundongos Nus , Camundongos Transgênicos , Invasividade Neoplásica , Micrometástase de Neoplasia , Ligação Proteica , Carga Tumoral
2.
EMBO Rep ; : e53014, 2021 Aug 17.
Artigo em Inglês | MEDLINE | ID: mdl-34403195

RESUMO

Prostate cancer (PCa) is one of the most prevalent cancers in men. Cancer stem cells are thought to be associated with PCa relapse. Here, we show that BAZ2A is required for PCa cells with a cancer stem-like state. BAZ2A genomic occupancy in PCa cells coincides with H3K14ac-enriched chromatin regions. This association is mediated by BAZ2A-bromodomain (BAZ2A-BRD) that specifically binds H3K14ac. BAZ2A associates with inactive enhancers marked by H3K14ac and repressing transcription of genes frequently silenced in aggressive and poorly differentiated PCa. BAZ2A-mediated repression is also linked to EP300 that acetylates H3K14ac. BAZ2A-BRD mutations or treatment with inhibitors abrogating BAZ2A-BRD/H3K14ac interaction impair PCa stem cells. Furthermore, pharmacological inactivation of BAZ2A-BRD impairs Pten-loss oncogenic transformation of prostate organoids. Our findings indicate a role of BAZ2A-BRD in PCa stem cell features and suggest potential epigenetic-reader therapeutic strategies to target BAZ2A in aggressive PCa.

3.
Stem Cell Reports ; 15(6): 1206-1219, 2020 12 08.
Artigo em Inglês | MEDLINE | ID: mdl-32976768

RESUMO

The nucleolus is the largest compartment of the eukaryotic cell's nucleus. It acts as a ribosome factory, thereby sustaining the translation machinery. The nucleolus is also the subnuclear compartment with the highest transcriptional activity in the cell, where hundreds of ribosomal RNA (rRNA) genes transcribe the overwhelming majority of RNAs. The structure and composition of the nucleolus change according to the developmental state. For instance, in embryonic stem cells (ESCs), rRNA genes display a hyperactive transcriptional state and open chromatin structure compared with differentiated cells. Increasing evidence indicates that the role of the nucleolus and rRNA genes might go beyond the control of ribosome biogenesis. One such role is linked to the genome architecture, since repressive domains are often located close to the nucleolus. This review highlights recent findings describing how the nucleolus is regulated in ESCs and its role in regulating ribosome biogenesis and genome organization for the maintenance of stem cell identity.

4.
Proc Natl Acad Sci U S A ; 117(7): 3637-3647, 2020 02 18.
Artigo em Inglês | MEDLINE | ID: mdl-32024754

RESUMO

Prostate cancer (PCa) is the second leading cause of cancer death in men. Its clinical and molecular heterogeneities and the lack of in vitro models outline the complexity of PCa in the clinical and research settings. We established an in vitro mouse PCa model based on organoid technology that takes into account the cell of origin and the order of events. Primary PCa with deletion of the tumor suppressor gene PTEN (PTEN-del) can be modeled through Pten-down-regulation in mouse organoids. We used this system to elucidate the contribution of TIP5 in PCa initiation, a chromatin regulator that is implicated in aggressive PCa. High TIP5 expression correlates with primary PTEN-del PCa and this combination strongly associates with reduced prostate-specific antigen (PSA) recurrence-free survival. TIP5 is critical for the initiation of PCa of luminal origin mediated by Pten-loss whereas it is dispensable once Pten-loss mediated transformation is established. Cross-species analyses revealed a PTEN gene signature that identified a group of aggressive primary PCas characterized by PTEN-del, high-TIP5 expression, and a TIP5-regulated gene expression profile. The results highlight the modeling of PCa with organoids as a powerful tool to elucidate the role of genetic alterations found in recent studies in their time orders and cells of origin, thereby providing further optimization for tumor stratification to improve the clinical management of PCa.


Assuntos
Proteínas Cromossômicas não Histona/metabolismo , PTEN Fosfo-Hidrolase/genética , Neoplasias da Próstata/metabolismo , Animais , Carcinogênese , Transformação Celular Neoplásica , Proteínas Cromossômicas não Histona/genética , Deleção de Genes , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Camundongos , Camundongos Knockout , PTEN Fosfo-Hidrolase/metabolismo , Próstata/metabolismo , Neoplasias da Próstata/genética
5.
EMBO J ; 39(23): e105606, 2020 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-33433018

RESUMO

Chromosomes have an intrinsic tendency to segregate into compartments, forming long-distance contacts between loci of similar chromatin states. How genome compartmentalization is regulated remains elusive. Here, comparison of mouse ground-state embryonic stem cells (ESCs) characterized by open and active chromatin, and advanced serum ESCs with a more closed and repressed genome, reveals distinct regulation of their genome organization due to differential dependency on BAZ2A/TIP5, a component of the chromatin remodeling complex NoRC. On ESC chromatin, BAZ2A interacts with SNF2H, DNA topoisomerase 2A (TOP2A) and cohesin. BAZ2A associates with chromatin sub-domains within the active A compartment, which intersect through long-range contacts. We found that ground-state chromatin selectively requires BAZ2A to limit the invasion of active domains into repressive compartments. BAZ2A depletion increases chromatin accessibility at B compartments. Furthermore, BAZ2A regulates H3K27me3 genome occupancy in a TOP2A-dependent manner. Finally, ground-state ESCs require BAZ2A for growth, differentiation, and correct expression of developmental genes. Our results uncover the propensity of open chromatin domains to invade repressive domains, which is counteracted by chromatin remodeling to establish genome partitioning and preserve cell identity.


Assuntos
Proteínas Cromossômicas não Histona/genética , Proteínas Cromossômicas não Histona/metabolismo , Genoma , Células-Tronco Pluripotentes/metabolismo , Adenosina Trifosfatases/metabolismo , Animais , Proteínas de Ciclo Celular , Diferenciação Celular , Cromatina/metabolismo , Montagem e Desmontagem da Cromatina , DNA Topoisomerases Tipo II/metabolismo , Epigenômica , Regulação da Expressão Gênica , Histonas/metabolismo , Camundongos , Células-Tronco Embrionárias Murinas/citologia , Células-Tronco Pluripotentes/citologia , Proteínas de Ligação a Poli-ADP-Ribose/metabolismo
6.
Trends Genet ; 35(11): 868-879, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31327501

RESUMO

The nucleolus is the largest substructure in the nucleus and forms around the nucleolar organizer regions (NORs), which comprise hundreds of rRNA genes. Recent evidence highlights further functions of the nucleolus that go beyond ribosome biogenesis. Data indicate that the nucleolus acts as a compartment for the location and regulation of repressive genomic domains and, together with the nuclear lamina, represents the hub for the organization of the inactive heterochromatin. In this review, we discuss recent findings that have revealed how nucleolar structure and rRNA gene chromatin states are regulated during early mammalian development and their contribution to the higher-order spatial organization of the genome.


Assuntos
Cromatina/genética , Desenvolvimento Embrionário/genética , Genes de RNAr , Animais , Diferenciação Celular/genética , Nucléolo Celular/genética , Núcleo Celular , Células-Tronco Embrionárias/metabolismo , Gametogênese/genética , Genoma , Mamíferos , Oócitos/metabolismo
7.
Cells ; 8(6)2019 06 12.
Artigo em Inglês | MEDLINE | ID: mdl-31212844

RESUMO

The nucleolus is the largest substructure in the nucleus, where ribosome biogenesis takes place, and forms around the nucleolar organizer regions (NORs) that comprise ribosomal RNA (rRNA) genes. Each cell contains hundreds of rRNA genes, which are organized in three distinct chromatin and transcriptional states-silent, inactive and active. Increasing evidence indicates that the role of the nucleolus and rRNA genes goes beyond the control of ribosome biogenesis. Recent results highlighted the nucleolus as a compartment for the location and regulation of repressive genomic domains and, together with the nuclear lamina, represents the hub for the organization of the inactive heterochromatin. In this review, we aim to describe the crosstalk between the nucleolus and the rest of the genome and how distinct rRNA gene chromatin states affect nucleolus structure and are implicated in genome stability, genome architecture, and cell fate decision.


Assuntos
Nucléolo Celular/genética , Genoma , Animais , Células-Tronco Embrionárias/metabolismo , Instabilidade Genômica , Humanos , RNA Ribossômico/genética
8.
Epigenetics Chromatin ; 11(1): 60, 2018 10 17.
Artigo em Inglês | MEDLINE | ID: mdl-30333056

RESUMO

BACKGROUND: Reactive oxygen species (ROS)-induced oxidative stress is well known to play a major role in male infertility. Sperm are sensitive to ROS damaging effects because as male germ cells form mature sperm they progressively lose the ability to repair DNA damage. However, how oxidative DNA lesions in sperm affect early embryonic development remains elusive. RESULTS: Using cattle as model, we show that fertilization using sperm exposed to oxidative stress caused a major developmental arrest at the time of embryonic genome activation. The levels of DNA damage response did not directly correlate with the degree of developmental defects. The early cellular response for DNA damage, γH2AX, is already present at high levels in zygotes that progress normally in development and did not significantly increase at the paternal genome containing oxidative DNA lesions. Moreover, XRCC1, a factor implicated in the last step of base excision repair (BER) pathway, was recruited to the damaged paternal genome, indicating that the maternal BER machinery can repair these DNA lesions induced in sperm. Remarkably, the paternal genome with oxidative DNA lesions showed an impairment of zygotic active DNA demethylation, a process that previous studies linked to BER. Quantitative immunofluorescence analysis and ultrasensitive LC-MS-based measurements revealed that oxidative DNA lesions in sperm impair active DNA demethylation at paternal pronuclei, without affecting 5-hydroxymethylcytosine (5hmC), a 5-methylcytosine modification that has been implicated in paternal active DNA demethylation in mouse zygotes. Thus, other 5hmC-independent processes are implicated in active DNA demethylation in bovine embryos. The recruitment of XRCC1 to damaged paternal pronuclei indicates that oxidative DNA lesions drive BER to repair DNA at the expense of DNA demethylation. Finally, this study highlighted striking differences in DNA methylation dynamics between bovine and mouse zygotes that will facilitate the understanding of the dynamics of DNA methylation in early development. CONCLUSIONS: The data demonstrate that oxidative stress in sperm has an impact not only on DNA integrity but also on the dynamics of epigenetic reprogramming, which may harm the paternal genetic and epigenetic contribution to the developing embryo and affect embryo development and embryo quality.


Assuntos
Desenvolvimento Embrionário/genética , Epigênese Genética , Estresse Oxidativo , Herança Paterna , Espermatozoides/metabolismo , Animais , Bovinos , Metilação de DNA , Reparo do DNA , Feminino , Masculino , Proteína 1 Complementadora Cruzada de Reparo de Raio-X/metabolismo
9.
Cancer Cell ; 34(1): 69-84.e14, 2018 07 09.
Artigo em Inglês | MEDLINE | ID: mdl-30008323

RESUMO

Human melanomas frequently harbor amplifications of EZH2. However, the contribution of EZH2 to melanoma formation has remained elusive. Taking advantage of murine melanoma models, we show that EZH2 drives tumorigenesis from benign BrafV600E- or NrasQ61K-expressing melanocytes by silencing of genes relevant for the integrity of the primary cilium, a signaling organelle projecting from the surface of vertebrate cells. Consequently, gain of EZH2 promotes loss of primary cilia in benign melanocytic lesions. In contrast, blockade of EZH2 activity evokes ciliogenesis and cilia-dependent growth inhibition in malignant melanoma. Finally, we demonstrate that loss of cilia enhances pro-tumorigenic WNT/ß-catenin signaling, and is itself sufficient to drive metastatic melanoma in benign cells. Thus, primary cilia deconstruction is a key process in EZH2-driven melanomagenesis.


Assuntos
Movimento Celular , Proliferação de Células , Cílios/metabolismo , Proteína Potenciadora do Homólogo 2 de Zeste/metabolismo , Melanócitos/metabolismo , Melanoma/metabolismo , Neoplasias Cutâneas/metabolismo , Animais , Linhagem Celular Tumoral , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/metabolismo , Transformação Celular Neoplásica/patologia , Cílios/genética , Cílios/patologia , Proteína Potenciadora do Homólogo 2 de Zeste/genética , Feminino , GTP Fosfo-Hidrolases/genética , Regulação Neoplásica da Expressão Gênica , Células HEK293 , Humanos , Metástase Linfática , Masculino , Melanócitos/patologia , Melanoma/genética , Melanoma/secundário , Proteínas de Membrana/genética , Camundongos Nus , Camundongos Transgênicos , Proteínas Proto-Oncogênicas B-raf/genética , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/patologia , Via de Sinalização Wnt , beta Catenina/genética , beta Catenina/metabolismo
11.
Nat Cell Biol ; 19(7): 763-773, 2017 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-28604677

RESUMO

Naive pluripotency is established in preimplantation epiblast. Embryonic stem cells (ESCs) represent the immortalization of naive pluripotency. 2i culture has optimized this state, leading to a gene signature and DNA hypomethylation closely comparable to preimplantation epiblast, the developmental ground state. Here we show that Pramel7 (PRAME-like 7), a protein highly expressed in the inner cell mass (ICM) but expressed at low levels in ESCs, targets for proteasomal degradation UHRF1, a key factor for DNA methylation maintenance. Increasing Pramel7 expression in serum-cultured ESCs promotes a preimplantation epiblast-like gene signature, reduces UHRF1 levels and causes global DNA hypomethylation. Pramel7 is required for blastocyst formation and its forced expression locks ESCs in pluripotency. Pramel7/UHRF1 expression is mutually exclusive in ICMs whereas Pramel7-knockout embryos express high levels of UHRF1. Our data reveal an as-yet-unappreciated dynamic nature of DNA methylation through proteasome pathways and offer insights that might help to improve ESC culture to reproduce in vitro the in vivo ground-state pluripotency.


Assuntos
Antígenos de Neoplasias/metabolismo , Blastocisto/enzimologia , Células-Tronco Embrionárias/enzimologia , Epigênese Genética , Proteínas de Neoplasias/metabolismo , Proteínas Nucleares/metabolismo , Células-Tronco Pluripotentes/enzimologia , Complexo de Endopeptidases do Proteassoma/metabolismo , Animais , Antígenos de Neoplasias/genética , Blastocisto/citologia , Proteínas Estimuladoras de Ligação a CCAAT , Proteínas Culina/metabolismo , Metilação de DNA , Regulação da Expressão Gênica no Desenvolvimento , Células HEK293 , Humanos , Camundongos Endogâmicos C57BL , Proteínas de Neoplasias/genética , Proteínas Nucleares/genética , Fenótipo , Domínios e Motivos de Interação entre Proteínas , Estabilidade Proteica , Proteólise , Interferência de RNA , Fatores de Tempo , Transcriptoma , Transfecção , Ubiquitina-Proteína Ligases
12.
EMBO Rep ; 18(7): 1248-1262, 2017 07.
Artigo em Inglês | MEDLINE | ID: mdl-28588071

RESUMO

Long non-coding RNAs (lncRNAs) have been implicated in the regulation of chromatin conformation and epigenetic patterns. lncRNA expression levels are widely taken as an indicator for functional properties. However, the role of RNA processing in modulating distinct features of the same lncRNA is less understood. The establishment of heterochromatin at rRNA genes depends on the processing of IGS-rRNA into pRNA, a reaction that is impaired in embryonic stem cells (ESCs) and activated only upon differentiation. The production of mature pRNA is essential since it guides the repressor TIP5 to rRNA genes, and IGS-rRNA abolishes this process. Through screening for IGS-rRNA-binding proteins, we here identify the RNA helicase DHX9 as a regulator of pRNA processing. DHX9 binds to rRNA genes only upon ESC differentiation and its activity guides TIP5 to rRNA genes and establishes heterochromatin. Remarkably, ESCs depleted of DHX9 are unable to differentiate and this phenotype is reverted by the addition of pRNA, whereas providing IGS-rRNA and pRNA mutants deficient for TIP5 binding are not sufficient. Our results reveal insights into lncRNA biogenesis during development and support a model in which the state of rRNA gene chromatin is part of the regulatory network that controls exit from pluripotency and initiation of differentiation pathways.


Assuntos
Diferenciação Celular , RNA Helicases DEAD-box/metabolismo , Células-Tronco Embrionárias/fisiologia , Heterocromatina/metabolismo , Proteínas de Neoplasias/metabolismo , Animais , Proteínas Cromossômicas não Histona , RNA Helicases DEAD-box/genética , DNA Ribossômico , Epigênese Genética , Genes de RNAr , Células HEK293 , Humanos , Camundongos , Células NIH 3T3 , Proteínas de Neoplasias/genética , RNA Helicases/metabolismo , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo
13.
Parasitol Res ; 116(6): 1745-1753, 2017 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-28466247

RESUMO

Bioaccumulation of 13 trace elements in the livers of 38 Pelophylax sinkl. hispanicus (Ranidae) and its helminth communities were studied and compared among three sites, each with a different degree of pollution along River Neto (south Italy) during September, 2014. Trace element concentrations in water and liver were measured using inductively coupled plasma mass spectrometry. For most elements, the highest concentration was recorded in the frogs inhabiting the third site, the one with the highest degree of pollution. The trend of trace element concentration in the liver can be represented as follows: Cu > Zn > Mn > Se > Cr. Concentrations of some elements in water and liver samples were significantly different among the three sites and this is evidenced by the bioaccumulation in the frogs. Four species of helminths, all belonging to Nematoda, were found: Rhabdias sp., Oswaldocruzia filiformis (Goeze, 1782), Cosmocerca ornata (Dujarden, 1845), Seuratascaris numidica (Seurat, 1917). The parasite survey presents an important difference of prevalence and average number of helminths in frogs between the three sites. Correlating parasitological and ecotoxicological data showed a strong positive correlation between prevalence and number of parasites with some trace elements such as Mn, Co, Ni, As, Se, and Cd.


Assuntos
Monitoramento Ambiental , Nematoides/isolamento & purificação , Infecções por Nematoides/veterinária , Oligoelementos , Animais , Itália/epidemiologia , Fígado/química , Nematoides/classificação , Infecções por Nematoides/epidemiologia , Infecções por Nematoides/parasitologia , Ranidae/parasitologia , Fatores de Risco , Rios
14.
J Clin Invest ; 126(11): 4237-4249, 2016 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-27760049

RESUMO

A vast number of cancer genes are transcription factors that drive tumorigenesis as oncogenic fusion proteins. Although the direct targeting of transcription factors remains challenging, therapies aimed at oncogenic fusion proteins are attractive as potential treatments for cancer. There is particular interest in targeting the oncogenic PAX3-FOXO1 fusion transcription factor, which induces alveolar rhabdomyosarcoma (aRMS), an aggressive cancer of skeletal muscle cells for which patient outcomes remain dismal. In this work, we have defined the interactome of PAX3-FOXO1 and screened 60 candidate interactors using siRNA-mediated depletion to identify candidates that affect fusion protein activity in aRMS cells. We report that chromodomain helicase DNA binding protein 4 (CHD4), an ATP-dependent chromatin remodeler, acts as crucial coregulator of PAX3-FOXO1 activity. CHD4 interacts with PAX3-FOXO1 via short DNA fragments. Together, they bind to regulatory regions of PAX3-FOXO1 target genes. Gene expression analysis suggested that CHD4 coregulatory activity is essential for a subset of PAX3-FOXO1 target genes. Depletion of CHD4 reduced cell viability of fusion-positive but not of fusion-negative RMS in vitro, which resembled loss of PAX3-FOXO1. It also caused specific regression of fusion-positive xenograft tumors in vivo. Therefore, this work identifies CHD4 as an epigenetic coregulator of PAX3-FOXO1 activity, providing rational evidence for CHD4 as a potential therapeutic target in aRMS.


Assuntos
Autoantígenos/metabolismo , Epigênese Genética , Regulação Neoplásica da Expressão Gênica , Complexo Mi-2 de Remodelação de Nucleossomo e Desacetilase/metabolismo , Proteínas de Fusão Oncogênica/metabolismo , Fatores de Transcrição Box Pareados/metabolismo , Rabdomiossarcoma Alveolar/metabolismo , Animais , Autoantígenos/genética , Linhagem Celular Tumoral , Feminino , Xenoenxertos , Humanos , Complexo Mi-2 de Remodelação de Nucleossomo e Desacetilase/genética , Camundongos Endogâmicos NOD , Camundongos SCID , Transplante de Neoplasias , Proteínas de Fusão Oncogênica/genética , Fatores de Transcrição Box Pareados/genética , Rabdomiossarcoma Alveolar/genética , Rabdomiossarcoma Alveolar/patologia
15.
FEBS Lett ; 590(15): 2342-53, 2016 08.
Artigo em Inglês | MEDLINE | ID: mdl-27417130

RESUMO

Long noncoding RNA (lncRNA) are emerging as important regulators of diverse biological functions. Although mechanistic models are starting to emerge, it is also clear that the lncRNA field needs appropriate model systems in order to better elucidate the functions of lncRNA and their roles in both physiological and pathological conditions. The field of lncRNA is new, and the biochemical and genetic methods used to address function and mechanisms of lncRNA have only recently been developed or adapted from techniques used to investigate protein-coding genes. In this review, we discuss the strengths and weaknesses of available techniques for the analysis of chromatin-associated lncRNA and emerging models for the recruitment to specific genomic sites such as triple-helix, RNA-protein-DNA recognition and proximity-guided search models.


Assuntos
Cromatina/genética , RNA Longo não Codificante/genética , Regulação da Expressão Gênica , Genoma , Humanos
16.
Oncotarget ; 7(16): 21272-86, 2016 Apr 19.
Artigo em Inglês | MEDLINE | ID: mdl-26848772

RESUMO

Calretinin (CALB2) is a diagnostic marker for epithelioid mesothelioma. It is also a prognostic marker since patients with tumors expressing high calretinin levels have better overall survival. Silencing of calretinin decreases viability of epithelioid mesothelioma cells. Our aim was to elucidate mechanisms regulating calretinin expression in mesothelioma. Analysis of calretinin transcript and protein suggested a control at the mRNA level. Treatment with 5-aza-2'-deoxycytidine and analysis of TCGA data indicated that promoter methylation is not likely to be involved. Therefore, we investigated CALB2 promoter by analyzing ~1kb of genomic sequence surrounding the transcription start site (TSS) + 1 using promoter reporter assay. Deletion analysis of CALB2 proximal promoter showed that sequence spanning the -161/+80bp region sustained transcriptional activity. Site-directed analysis identified important cis-regulatory elements within this -161/+80bp CALB2 promoter. EMSA and ChIP assays confirmed binding of NRF-1 and E2F2 to the CALB2 promoter and siRNA knockdown of NRF-1 led to decreased expression of calretinin. Cell synchronization experiment showed that calretinin expression was cell cycle regulated with a peak of expression at G1/S phase. This study provides the first insight in the regulation of CALB2 expression in mesothelioma cells.


Assuntos
Calbindina 2/genética , Regulação Neoplásica da Expressão Gênica , Mesotelioma/genética , Regiões Promotoras Genéticas/genética , Sequências Reguladoras de Ácido Nucleico , Sequência de Bases , Calbindina 2/metabolismo , Células Cultivadas , Fator de Transcrição E2F2/genética , Fator de Transcrição E2F2/metabolismo , Humanos , Mesotelioma/metabolismo , Mesotelioma/patologia , Fator 1 Nuclear Respiratório/genética , Fator 1 Nuclear Respiratório/metabolismo
17.
Genome Biol ; 16: 42, 2015 Feb 22.
Artigo em Inglês | MEDLINE | ID: mdl-25885555

RESUMO

BACKGROUND: Melanoma is the most fatal skin cancer displaying a high degree of molecular heterogeneity. Phenotype switching is a mechanism that contributes to melanoma heterogeneity by altering transcription profiles for the transition between states of proliferation/differentiation and invasion/stemness. As phenotype switching is reversible, epigenetic mechanisms, like DNA methylation, could contribute to the changes in gene expression. RESULTS: Integrative analysis of methylation and gene expression datasets of five proliferative and five invasion melanoma cell cultures reveal two distinct clusters. SOX9 is methylated and lowly expressed in the highly proliferative group. SOX9 overexpression results in decreased proliferation but increased invasion in vitro. In a B16 mouse model, sox9 overexpression increases the number of lung metastases. Transcriptional analysis of SOX9-overexpressing melanoma cells reveals enrichment in epithelial to mesenchymal transition (EMT) pathways. Survival analysis of The Cancer Genome Atlas melanoma dataset shows that metastatic patients with high expression levels of SOX9 have significantly worse survival rates. Additional survival analysis on the targets of SOX9 reveals that most SOX9 downregulated genes have survival benefit for metastatic patients. CONCLUSIONS: Our genome-wide DNA methylation and gene expression study of 10 early passage melanoma cell cultures reveals two phenotypically distinct groups. One of the genes regulated by DNA methylation between the two groups is SOX9. SOX9 induces melanoma cell invasion and metastasis and decreases patient survival. A number of genes downregulated by SOX9 have a negative impact on patient survival. In conclusion, SOX9 is an important gene involved in melanoma invasion and negatively impacts melanoma patient survival.


Assuntos
Melanoma/genética , Invasividade Neoplásica/genética , Fatores de Transcrição SOX9/biossíntese , Neoplasias Cutâneas/genética , Idoso , Animais , Linhagem Celular Tumoral , Proliferação de Células/genética , Metilação de DNA/genética , Transição Epitelial-Mesenquimal/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Melanoma/patologia , Camundongos , Pessoa de Meia-Idade , Invasividade Neoplásica/patologia , Estadiamento de Neoplasias , Prognóstico , Fatores de Transcrição SOX9/genética , Transdução de Sinais , Neoplasias Cutâneas/patologia , Análise de Sobrevida
18.
Nat Commun ; 6: 6051, 2015 Jan 22.
Artigo em Inglês | MEDLINE | ID: mdl-25609585

RESUMO

Increased activity of the epigenetic modifier EZH2 has been associated with different cancers. However, evidence for a functional role of EZH2 in tumorigenesis in vivo remains poor, in particular in metastasizing solid cancers. Here we reveal central roles of EZH2 in promoting growth and metastasis of cutaneous melanoma. In a melanoma mouse model, conditional Ezh2 ablation as much as treatment with the preclinical EZH2 inhibitor GSK503 stabilizes the disease through inhibition of growth and virtually abolishes metastases formation without affecting normal melanocyte biology. Comparably, in human melanoma cells, EZH2 inactivation impairs proliferation and invasiveness, accompanied by re-expression of tumour suppressors connected to increased patient survival. These EZH2 target genes suppress either melanoma growth or metastasis in vivo, revealing the dual function of EZH2 in promoting tumour progression. Thus, EZH2-mediated epigenetic repression is highly relevant especially during advanced melanoma progression, which makes EZH2 a promising target for novel melanoma therapies.


Assuntos
Inativação Gênica , Melanoma/metabolismo , Complexo Repressor Polycomb 2/fisiologia , Neoplasias Cutâneas/metabolismo , Adenosilmetionina Descarboxilase/metabolismo , Animais , Proliferação de Células , Proteína Potenciadora do Homólogo 2 de Zeste , Epigênese Genética , Transição Epitelial-Mesenquimal , Feminino , Regulação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Genótipo , Homeostase , Humanos , Melanócitos/metabolismo , Melanoma Experimental/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Invasividade Neoplásica , Metástase Neoplásica , Complexo Repressor Polycomb 2/genética , Resultado do Tratamento
19.
PLoS Genet ; 11(1): e1004877, 2015 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-25629959

RESUMO

Melanoma is the most fatal skin cancer, but the etiology of this devastating disease is still poorly understood. Recently, the transcription factor Sox10 has been shown to promote both melanoma initiation and progression. Reducing SOX10 expression levels in human melanoma cells and in a genetic melanoma mouse model, efficiently abolishes tumorigenesis by inducing cell cycle exit and apoptosis. Here, we show that this anti-tumorigenic effect functionally involves SOX9, a factor related to SOX10 and upregulated in melanoma cells upon loss of SOX10. Unlike SOX10, SOX9 is not required for normal melanocyte stem cell function, the formation of hyperplastic lesions, and melanoma initiation. To the contrary, SOX9 overexpression results in cell cycle arrest, apoptosis, and a gene expression profile shared by melanoma cells with reduced SOX10 expression. Moreover, SOX9 binds to the SOX10 promoter and induces downregulation of SOX10 expression, revealing a feedback loop reinforcing the SOX10 low/SOX9 high ant,m/ii-tumorigenic program. Finally, SOX9 is required in vitro and in vivo for the anti-tumorigenic effect achieved by reducing SOX10 expression. Thus, SOX10 and SOX9 are functionally antagonistic regulators of melanoma development.


Assuntos
Carcinogênese/genética , Melanoma/genética , Fatores de Transcrição SOX9/genética , Fatores de Transcrição SOXE/genética , Animais , Apoptose/genética , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica , Folículo Piloso , Humanos , Melanócitos/patologia , Melanoma/patologia , Camundongos , RNA Interferente Pequeno , Fatores de Transcrição SOX9/biossíntese , Fatores de Transcrição SOXE/biossíntese
20.
Nat Genet ; 47(1): 22-30, 2015 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-25485837

RESUMO

Prostate cancer is driven by a combination of genetic and/or epigenetic alterations. Epigenetic alterations are frequently observed in all human cancers, yet how aberrant epigenetic signatures are established is poorly understood. Here we show that the gene encoding BAZ2A (TIP5), a factor previously implicated in epigenetic rRNA gene silencing, is overexpressed in prostate cancer and is paradoxically involved in maintaining prostate cancer cell growth, a feature specific to cancer cells. BAZ2A regulates numerous protein-coding genes and directly interacts with EZH2 to maintain epigenetic silencing at genes repressed in metastasis. BAZ2A overexpression is tightly associated with a molecular subtype displaying a CpG island methylator phenotype (CIMP). Finally, high BAZ2A levels serve as an independent predictor of biochemical recurrence in a cohort of 7,682 individuals with prostate cancer. This work identifies a new aberrant role for the epigenetic regulator BAZ2A, which can also serve as a useful marker for metastatic potential in prostate cancer.


Assuntos
Adenocarcinoma/genética , Proteínas Cromossômicas não Histona/fisiologia , Repressão Epigenética , Regulação Neoplásica da Expressão Gênica , Metástase Neoplásica/genética , Proteínas de Neoplasias/fisiologia , Neoplasias da Próstata/genética , Adenocarcinoma/metabolismo , Adenocarcinoma/patologia , Biomarcadores Tumorais/genética , Divisão Celular , Linhagem Celular Tumoral , Proteínas Cromossômicas não Histona/biossíntese , Proteínas Cromossômicas não Histona/genética , Ilhas de CpG , Metilação de DNA , Proteína Potenciadora do Homólogo 2 de Zeste , Seguimentos , Humanos , Masculino , Invasividade Neoplásica/genética , Proteínas de Neoplasias/biossíntese , Proteínas de Neoplasias/genética , Complexo Repressor Polycomb 2/fisiologia , Prognóstico , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Mapeamento de Interação de Proteínas , RNA Neoplásico/biossíntese , RNA Ribossômico/biossíntese , Regulação para Cima
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