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1.
Nature ; 619(7969): 385-393, 2023 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-37407816

RESUMO

The basic helix-loop-helix (bHLH) family of transcription factors recognizes DNA motifs known as E-boxes (CANNTG) and includes 108 members1. Here we investigate how chromatinized E-boxes are engaged by two structurally diverse bHLH proteins: the proto-oncogene MYC-MAX and the circadian transcription factor CLOCK-BMAL1 (refs. 2,3). Both transcription factors bind to E-boxes preferentially near the nucleosomal entry-exit sites. Structural studies with engineered or native nucleosome sequences show that MYC-MAX or CLOCK-BMAL1 triggers the release of DNA from histones to gain access. Atop the H2A-H2B acidic patch4, the CLOCK-BMAL1 Per-Arnt-Sim (PAS) dimerization domains engage the histone octamer disc. Binding of tandem E-boxes5-7 at endogenous DNA sequences occurs through direct interactions between two CLOCK-BMAL1 protomers and histones and is important for circadian cycling. At internal E-boxes, the MYC-MAX leucine zipper can also interact with histones H2B and H3, and its binding is indirectly enhanced by OCT4 elsewhere on the nucleosome. The nucleosomal E-box position and the type of bHLH dimerization domain jointly determine the histone contact, the affinity and the degree of competition and cooperativity with other nucleosome-bound factors.


Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos , DNA , Histonas , Fatores de Transcrição ARNTL/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , DNA/genética , DNA/metabolismo , Sequências Hélice-Alça-Hélice/genética , Histonas/química , Histonas/metabolismo , Nucleossomos/química , Nucleossomos/genética , Nucleossomos/metabolismo , Ligação Proteica , Proteínas CLOCK/química , Proteínas CLOCK/metabolismo , Proteínas Proto-Oncogênicas c-myc/química , Proteínas Proto-Oncogênicas c-myc/metabolismo , Regulação Alostérica , Zíper de Leucina , Fator 3 de Transcrição de Octâmero/metabolismo , Multimerização Proteica
2.
J Hepatol ; 79(1): 141-149, 2023 07.
Artigo em Inglês | MEDLINE | ID: mdl-36906109

RESUMO

BACKGROUND & AIMS: Primary liver cancer (PLC) comprises hepatocellular carcinoma (HCC) and intrahepatic cholangiocarcinoma (iCCA), two frequent and lethal tumour types that differ regarding their tumour biology and responses to cancer therapies. Liver cells harbour a high degree of cellular plasticity and can give rise to either HCC or iCCA. However, little is known about the cell-intrinsic mechanisms directing an oncogenically transformed liver cell to either HCC or iCCA. The scope of this study was to identify cell-intrinsic factors determining lineage commitment in PLC. METHODS: Cross-species transcriptomic and epigenetic profiling was applied to murine HCCs and iCCAs and to two human PLC cohorts. Integrative data analysis comprised epigenetic Landscape In Silico deletion Analysis (LISA) of transcriptomic data and Hypergeometric Optimization of Motif EnRichment (HOMER) analysis of chromatin accessibility data. Identified candidate genes were subjected to functional genetic testing in non-germline genetically engineered PLC mouse models (shRNAmir knockdown or overexpression of full-length cDNAs). RESULTS: Integrative bioinformatic analyses of transcriptomic and epigenetic data pinpointed the Forkhead-family transcription factors FOXA1 and FOXA2 as MYC-dependent determination factors of the HCC lineage. Conversely, the ETS family transcription factor ETS1 was identified as a determinant of the iCCA lineage, which was found to be suppressed by MYC during HCC development. Strikingly, shRNA-mediated suppression of FOXA1 and FOXA2 with concomitant ETS1 expression fully switched HCC to iCCA development in PLC mouse models. CONCLUSIONS: The herein reported data establish MYC as a key determinant of lineage commitment in PLC and provide a molecular explanation why common liver-damaging risk factors such as alcoholic or non-alcoholic steatohepatitis can lead to either HCC or iCCA. IMPACT AND IMPLICATIONS: Liver cancer is a major health problem and comprises hepatocellular carcinoma (HCC) and intrahepatic cholangiocarcinoma (iCCA), two frequent and lethal tumour types that differ regarding their morphology, tumour biology, and responses to cancer therapies. We identified the transcription factor and oncogenic master regulator MYC as a switch between HCC and iCCA development. When MYC levels are high at the time point when a hepatocyte becomes a tumour cell, an HCC is growing out. Conversely, if MYC levels are low at this time point, the result is the outgrowth of an iCCA. Our study provides a molecular explanation why common liver-damaging risk factors such as alcoholic or non-alcoholic steatohepatitis can lead to either HCC or iCCA. Furthermore, our data harbour potential for the development of better PLC therapies.


Assuntos
Neoplasias dos Ductos Biliares , Carcinoma Hepatocelular , Colangiocarcinoma , Fígado Gorduroso , Neoplasias Hepáticas , Humanos , Animais , Camundongos , Carcinoma Hepatocelular/patologia , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Proteínas Proto-Oncogênicas p21(ras)/genética , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Colangiocarcinoma/genética , Colangiocarcinoma/metabolismo , Fatores de Transcrição/metabolismo , Ductos Biliares Intra-Hepáticos/patologia , Neoplasias dos Ductos Biliares/patologia
3.
Blood ; 141(5): 453-466, 2023 02 02.
Artigo em Inglês | MEDLINE | ID: mdl-36095844

RESUMO

Chromosomal rearrangements involving the MDS1 and EVI1 complex locus (MECOM) on chromosome 3q26 define an aggressive subtype of acute myeloid leukemia (AML) that is associated with chemotherapy resistance and dismal prognosis. Established treatment regimens commonly fail in these patients, therefore, there is an urgent need for new therapeutic concepts that will require a better understanding of the molecular and cellular functions of the ecotropic viral integration site 1 (EVI1) oncogene. To characterize gene regulatory functions of EVI1 and associated dependencies in AML, we developed experimentally tractable human and murine disease models, investigated the transcriptional consequences of EVI1 withdrawal in vitro and in vivo, and performed the first genome-wide CRISPR screens in EVI1-dependent AML. By integrating conserved transcriptional targets with genetic dependency data, we identified and characterized the ETS transcription factor ERG as a direct transcriptional target of EVI1 that is aberrantly expressed and selectively required in both human and murine EVI1-driven AML. EVI1 controls the expression of ERG and occupies a conserved intragenic enhancer region in AML cell lines and samples from patients with primary AML. Suppression of ERG induces terminal differentiation of EVI1-driven AML cells, whereas ectopic expression of ERG abrogates their dependence on EVI1, indicating that the major oncogenic functions of EVI1 are mediated through aberrant transcriptional activation of ERG. Interfering with this regulatory axis may provide entry points for the development of rational targeted therapies.


Assuntos
Proteínas de Ligação a DNA , Leucemia Mieloide Aguda , Humanos , Animais , Camundongos , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Proteína do Locus do Complexo MDS1 e EVI1/genética , Proto-Oncogenes/genética , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/metabolismo , Carcinogênese/genética , Regulador Transcricional ERG/genética
4.
Sci Immunol ; 6(61)2021 07 23.
Artigo em Inglês | MEDLINE | ID: mdl-34301800

RESUMO

The transcription factor Pax5 controls B cell development, but its role in mature B cells is largely enigmatic. Here, we demonstrated that the loss of Pax5 by conditional mutagenesis in peripheral B lymphocytes led to the strong reduction of B-1a, marginal zone (MZ), and germinal center (GC) B cells as well as plasma cells. Follicular (FO) B cells tolerated the loss of Pax5 but had a shortened half-life. The Pax5-deficient FO B cells failed to proliferate upon B cell receptor or Toll-like receptor stimulation due to impaired PI3K-AKT signaling, which was caused by increased expression of PTEN, a negative regulator of the PI3K pathway. Pax5 restrained PTEN protein expression at the posttranscriptional level, likely involving Pten-targeting microRNAs. Additional PTEN loss in Pten,Pax5 double-mutant mice rescued FO B cell numbers and the development of MZ B cells but did not restore GC B cell formation. Hence, the posttranscriptional down-regulation of PTEN expression is an important function of Pax5 that facilitates the differentiation and survival of mature B cells, thereby promoting humoral immunity.


Assuntos
Linfócitos B/imunologia , Fator de Transcrição PAX5/imunologia , PTEN Fosfo-Hidrolase/imunologia , Fosfatidilinositol 3-Quinases/imunologia , Animais , Diferenciação Celular , Regulação para Baixo , Feminino , Masculino , Camundongos Transgênicos , Fator de Transcrição PAX5/genética , PTEN Fosfo-Hidrolase/genética , Receptores de Antígenos de Linfócitos B/imunologia , Transdução de Sinais , Receptores Toll-Like/imunologia
5.
Oncotarget ; 11(9): 875-890, 2020 Mar 03.
Artigo em Inglês | MEDLINE | ID: mdl-32180900

RESUMO

Diffuse large B-cell lymphoma (DLBCL) is the most common type of non-Hodgkin lymphomas worldwide and is characterized by a high diversity of genetic and molecular alterations. Chromosomal translocations and mutations leading to deregulated expression of the transcriptional repressor BCL6 occur in a significant fraction of DLBCL patients. An oncogenic role of BCL6 in the initiation of DLBCL has been shown as the constitutive expression of BCL6 in mice recapitulates the pathogenesis of human DLBCL. However, the role of BCL6 in tumor maintenance remains poorly investigated due to the absence of suitable genetic models and limitations of pharmacological inhibitors. Here, we have utilized tetracycline-inducible CRISPR/Cas9 mutagenesis to study the consequences of BCL6 deletion in established DLBCL models in culture and in vivo. We show that BCL6 knock-out in SU-DHL-4 cells in vitro results in an anti-proliferative response 4-7 days after Cas9 induction that was characterized by cell cycle (G1) arrest. Conditional BCL6 deletion in established DLBCL tumors in vivo induced a significant tumor growth inhibition with initial tumor stasis followed by slow tumor growth kinetics. Our findings support a role of BCL6 in the maintenance of lymphoma growth and showcase the utility of inducible CRISPR/Cas9 systems for probing oncogene addiction.

6.
Nat Immunol ; 17(10): 1187-96, 2016 10.
Artigo em Inglês | MEDLINE | ID: mdl-27487330

RESUMO

During unresolved infections, some viruses escape immunological control and establish a persistant reservoir in certain cell types, such as human immunodeficiency virus (HIV), which persists in follicular helper T cells (TFH cells), and Epstein-Barr virus (EBV), which persists in B cells. Here we identified a specialized group of cytotoxic T cells (TC cells) that expressed the chemokine receptor CXCR5, selectively entered B cell follicles and eradicated infected TFH cells and B cells. The differentiation of these cells, which we have called 'follicular cytotoxic T cells' (TFC cells), required the transcription factors Bcl6, E2A and TCF-1 but was inhibited by the transcriptional regulators Blimp1, Id2 and Id3. Blimp1 and E2A directly regulated Cxcr5 expression and, together with Bcl6 and TCF-1, formed a transcriptional circuit that guided TFC cell development. The identification of TFC cells has far-reaching implications for the development of strategies to control infections that target B cells and TFH cells and to treat B cell-derived malignancies.


Assuntos
Infecções por Arenaviridae/imunologia , Linfócitos B/imunologia , Infecções por Vírus Epstein-Barr/imunologia , HIV/imunologia , Vírus da Coriomeningite Linfocítica/imunologia , Linfócitos T Citotóxicos/imunologia , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Diferenciação Celular , Células Cultivadas , Regulação da Expressão Gênica , Centro Germinativo/patologia , Centro Germinativo/virologia , Fator 1-alfa Nuclear de Hepatócito/genética , Fator 1-alfa Nuclear de Hepatócito/metabolismo , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Fator 1 de Ligação ao Domínio I Regulador Positivo , Proteínas Proto-Oncogênicas c-bcl-6/genética , Proteínas Proto-Oncogênicas c-bcl-6/metabolismo , Receptores CXCR5/genética , Receptores CXCR5/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo
7.
J Immunol ; 192(6): 2667-76, 2014 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-24532575

RESUMO

NK cells can be grouped into distinct subsets that are localized to different organs and exhibit a different capacity to secrete cytokines and mediate cytotoxicity. Despite these hallmarks that reflect tissue-specific specialization in NK cells, little is known about the factors that control the development of these distinct subsets. The basic leucine zipper transcription factor Nfil3 (E4bp4) is essential for bone marrow-derived NK cell development, but it is not clear whether Nfil3 is equally important for all NK cell subsets or how it induces NK lineage commitment. In this article, we show that Nfil3 is required for the formation of Eomes-expressing NK cells, including conventional medullary and thymic NK cells, whereas TRAIL(+) Eomes(-) NK cells develop independently of Nfil3. Loss of Nfil3 during the development of bone marrow-derived NK cells resulted in reduced expression of Eomes and, conversely, restoration of Eomes expression in Nfil3(-/-) progenitors rescued NK cell development and maturation. Collectively, these findings demonstrate that Nfil3 drives the formation of mature NK cells by inducing Eomes expression and reveal the differential requirements of NK cell subsets for Nfil3.


Assuntos
Fatores de Transcrição de Zíper de Leucina Básica/imunologia , Diferenciação Celular/imunologia , Linhagem da Célula/imunologia , Células Matadoras Naturais/imunologia , Animais , Animais Recém-Nascidos , Fatores de Transcrição de Zíper de Leucina Básica/genética , Fatores de Transcrição de Zíper de Leucina Básica/metabolismo , Diferenciação Celular/genética , Linhagem da Célula/genética , Células Cultivadas , Citometria de Fluxo , Expressão Gênica/imunologia , Células Matadoras Naturais/metabolismo , Fígado/citologia , Fígado/imunologia , Fígado/metabolismo , Camundongos , Camundongos Knockout , Camundongos Transgênicos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Proteínas com Domínio T/genética , Proteínas com Domínio T/imunologia , Proteínas com Domínio T/metabolismo , Ligante Indutor de Apoptose Relacionado a TNF/genética , Ligante Indutor de Apoptose Relacionado a TNF/imunologia , Ligante Indutor de Apoptose Relacionado a TNF/metabolismo , Timo/citologia , Timo/imunologia , Timo/metabolismo
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