RESUMO
The last three decades of cancer research were guided by the hypothesis that cancer cells evolve due to the accumulation of many genetic aberrations over time. While this is still true for most solid cancers, it might be different in haemato-malignant diseases, which are mostly characterized by chromosomal translocations that exhibit only few additional mutations. Some of the resulting fusion gene products functionally interfer with epigenetic mechanisms. Recent findings of mutated IDH1, IDH2, DNMT3A or TET2 in myelodysplastic syndrome/acute myeloid leukaemia patients underscore this notion, and point to the importance of epigenetic changes for developing tumour cells. This review aims (i) to give an overview about the different components of the epigenetic system, (ii) to describe the functions of different proteins or complexes that are involved in setting-up the epigenetic layer, (iii) to highlight some recent findings, and (iv) to describe the failures and successes when using drugs that are targeting epigenetic components.
Assuntos
Doenças Hematológicas/genética , Epigênese Genética , HumanosRESUMO
AF4/AFF1 and AF5/AFF4 are the molecular backbone to assemble "super-elongation complexes" (SECs) that have two main functions: (1) control of transcriptional elongation by recruiting the positive transcription elongation factor b (P-TEFb = CyclinT1/CDK9) that is usually stored in inhibitory 7SK RNPs; (2) binding of different histone methyltransferases, like DOT1L, NSD1 and CARM1. This way, transcribed genes obtain specific histone signatures (e.g. H3K79me2/3, H3K36me2) to generate a transcriptional memory system. Here we addressed several questions: how is P-TEFb recruited into SEC, how is the AF4 interactome composed, and what is the function of the naturally occuring AF4N protein variant which exhibits only the first 360 amino acids of the AF4 full-length protein. Noteworthy, shorter protein variants are a specific feature of all AFF protein family members. Here, we demonstrate that full-length AF4 and AF4N are both catalyzing the transition of P-TEFb from 7SK RNP to their N-terminal domain. We have also mapped the protein-protein interaction network within both complexes. In addition, we have first evidence that the AF4N protein also recruits TFIIH and the tumor suppressor MEN1. This indicate that AF4N may have additional functions in transcriptional initiation and in MEN1-dependend transcriptional processes.
RESUMO
The human 5-lipoxygenase (5-LO), encoded by the ALOX5 gene, is the key enzyme in the formation of pro-inflammatory leukotrienes. ALOX5 gene transcription is strongly stimulated by calcitriol (1α, 25-dihydroxyvitamin D3) and TGFß (transforming growth factor-ß). Here, we investigated the influence of MLL (activator of transcript initiation), AF4 (activator of transcriptional elongation) as well as of the leukemogenic fusion proteins MLL-AF4 (ectopic activator of transcript initiation) and AF4-MLL (ectopic activator of transcriptional elongation) on calcitriol/TGFß-dependent 5-LO transcript elongation. We present evidence that the AF4 complex directly interacts with the vitamin D receptor (VDR) and promotes calcitriol-dependent ALOX5 transcript elongation. Activation of transcript elongation was strongly enhanced by the AF4-MLL fusion protein but was sensitive to Flavopiridol. By contrast, MLL-AF4 displayed no effect on transcriptional elongation. Furthermore, HDAC class I inhibitors inhibited the ectopic effects caused by AF4-MLL on transcriptional elongation, suggesting that HDAC class I inhibitors are potential therapeutics for the treatment of t(4;11)(q21;q23) leukemia.
Assuntos
Araquidonato 5-Lipoxigenase/biossíntese , Calcitriol/farmacologia , Proteínas de Ligação a DNA/metabolismo , Histona-Lisina N-Metiltransferase/metabolismo , Leucemia/enzimologia , Proteína de Leucina Linfoide-Mieloide/metabolismo , Proteínas Nucleares/metabolismo , Proteínas de Fusão Oncogênica/metabolismo , RNA Mensageiro/biossíntese , Elongação da Transcrição Genética/efeitos dos fármacos , Araquidonato 5-Lipoxigenase/genética , Quinase 9 Dependente de Ciclina/antagonistas & inibidores , Quinase 9 Dependente de Ciclina/metabolismo , Proteínas de Ligação a DNA/genética , Indução Enzimática , Células HEK293 , Células HeLa , Inibidores de Histona Desacetilases/farmacologia , Histona-Lisina N-Metiltransferase/genética , Humanos , Leucemia/tratamento farmacológico , Leucemia/genética , Ligantes , Proteína de Leucina Linfoide-Mieloide/genética , Proteínas Nucleares/genética , Proteínas de Fusão Oncogênica/genética , Regiões Promotoras Genéticas , Complexo de Endopeptidases do Proteassoma/metabolismo , Inibidores de Proteassoma/farmacologia , Ligação Proteica , Inibidores de Proteínas Quinases/farmacologia , Proteólise , Interferência de RNA , RNA Mensageiro/genética , Receptores de Calcitriol/agonistas , Receptores de Calcitriol/metabolismo , Fatores de Elongação da Transcrição , TransfecçãoRESUMO
MLL gene aberrations are frequently diagnosed in infant acute myeloid leukemia (AML). We previously described the MLL-NEBL and NEBL-MLL genomic fusions in an infant AML patient with a chromosomal translocation t(10;11)(p12;q23). NEBL was the second Nebulin family member (LASP1, NEBL) which was found to be involved in MLL rearrangements. Here, we report on our attempts to unravel the oncogenic properties of both fusion genes. First, RT-PCR analyses revealed the presence of the MLL-NEBL and NEBL-MLL mRNAs in the diagnostic sample of the patient. Next, expression cassettes for MLL-NEBL and NEBL-MLL were cloned into a sleeping beauty vector backbone. After stable transfection, the biological effects of MLL-NEBL, NEBL-MLL or the combination of both fusion proteins were investigated in a conditional cell culture model. NEBL-MLL but also co-transfected cells displayed significantly higher growth rates according to the data obtained by cell proliferation assay. The focus formation experiments revealed differences in the shape and number of colonies when comparing MLL-NEBL, NEBL-MLL- and co-transfected cells. The results obtained in this study suggest that the reciprocal fusion genes of the Nebulin gene family might be of biological importance.
Assuntos
Proteínas de Transporte/genética , Proteínas do Citoesqueleto/genética , Fusão Gênica , Proteínas com Domínio LIM/genética , Leucemia Mieloide Aguda/genética , Proteína de Leucina Linfoide-Mieloide/genética , Proteínas de Fusão Oncogênica/genética , Animais , Proteínas de Transporte/metabolismo , Proliferação de Células , Forma Celular , Proteínas do Citoesqueleto/metabolismo , Regulação Neoplásica da Expressão Gênica , Genótipo , Células HEK293 , Histona-Lisina N-Metiltransferase , Humanos , Lactente , Proteínas com Domínio LIM/metabolismo , Leucemia Mieloide Aguda/metabolismo , Leucemia Mieloide Aguda/patologia , Camundongos , Proteína de Leucina Linfoide-Mieloide/metabolismo , Proteínas de Fusão Oncogênica/metabolismo , Fenótipo , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , TransfecçãoRESUMO
Inspired by nature: Natural product isopropylstilbene was identified as an inhibitor of soluble epoxide hydrolase exhibiting antiproliferative properties. Following the natural product inspired design approach, a library of (E)-styryl-1H-benzo[d]imidazoles was synthesized and evaluated with recombinant enzyme and on several cancer cell lines.