RESUMEN
The transcription factor PLZF (promyelocytic leukemia zinc finger protein) acts as an epigenetic regulator balancing self-renewal and differentiation of hematopoietic cells through binding to various chromatin-modifying factors. First described as a transcriptional repressor, PLZF is also associated with active transcription, although the molecular bases underlying the differences are unknown. Here, we reveal that in a hematopoietic cell line, PLZF is predominantly associated with transcribed genes. Additionally, we identify a new association between PLZF and the histone methyltransferase, EZH2 at the genomic level. We find that co-occupancy of PLZF and EZH2 on chromatin at PLZF target genes is not associated with SUZ12 or trimethylated lysine 27 of histone H3 (H3K27me3) but with the active histone mark H3K4me3 and active transcription. Removal of EZH2 leads to an increase of PLZF binding and increased gene expression. Our results suggest a new role of EZH2 in restricting PLZF positive transcriptional activity independently of its canonical PRC2 activity.
Asunto(s)
Proteína Potenciadora del Homólogo Zeste 2/genética , Complejo Represivo Polycomb 2/genética , Proteína de la Leucemia Promielocítica con Dedos de Zinc/genética , Transcripción Genética , Sitios de Unión/genética , Diferenciación Celular/genética , Línea Celular Tumoral , Autorrenovación de las Células/genética , Cromatina/genética , Regulación de la Expresión Génica/genética , Células Madre Hematopoyéticas/metabolismo , Histona Metiltransferasas/genética , Histonas/genética , Humanos , Proteínas de Neoplasias , Unión Proteica/genética , Factores de TranscripciónRESUMEN
Termination of transcription is important for establishing gene punctuation marks. It is also critical for suppressing many of the pervasive transcription events occurring throughout eukaryotic genomes and coupling their RNA products to efficient decay. In human cells, the ARS2 protein has been implicated in such function as its depletion causes transcriptional read-through of selected gene terminators and because it physically interacts with the ribonucleolytic nuclear RNA exosome. Here, we study the role of ARS2 on transcription and RNA metabolism genome wide. We show that ARS2 depletion negatively impacts levels of promoter-proximal RNA polymerase II at protein-coding (pc) genes. Moreover, our results reveal a general role of ARS2 in transcription termination-coupled RNA turnover at short transcription units like snRNA-, replication-dependent histone-, promoter upstream transcript- and enhancer RNA-loci. Depletion of the ARS2 interaction partner ZC3H18 mimics the ARS2 depletion, although to a milder extent, whereas depletion of the exosome core subunit RRP40 only impacts RNA abundance post-transcriptionally. Interestingly, ARS2 is also involved in transcription termination events within first introns of pc genes. Our work therefore establishes ARS2 as a general suppressor of pervasive transcription with the potential to regulate pc gene expression.
Asunto(s)
Complejo Multienzimático de Ribonucleasas del Exosoma/metabolismo , Regulación de la Expresión Génica/fisiología , Proteínas Nucleares/fisiología , ARN Polimerasa II/metabolismo , Terminación de la Transcripción Genética , Inmunoprecipitación de Cromatina , Complejo Multienzimático de Ribonucleasas del Exosoma/fisiología , Células HeLa , Humanos , Intrones , Interferencia de ARN , ARN Mensajero/genética , ARN Interferente Pequeño/genética , ARN Nuclear Pequeño/genética , Proteínas de Unión al ARN/fisiologíaRESUMEN
Ets1 is a sequence-specific transcription factor that plays an important role during hematopoiesis, and is essential for the transition of CD4(-)/CD8(-) double negative (DN) to CD4(+)/CD8(+) double positive (DP) thymocytes. Using genome-wide and functional approaches, we investigated the binding properties, transcriptional role and chromatin environment of Ets1 during this transition. We found that while Ets1 binding at distal sites was associated with active genes at both DN and DP stages, its enhancer activity was attained at the DP stage, as reflected by levels of the core transcriptional hallmarks H3K4me1/3, RNA Polymerase II and eRNA. This dual, stage-specific ability reflected a switch from non-T hematopoietic toward T-cell specific gene expression programs during the DN-to-DP transition, as indicated by transcriptome analyses of Ets1(-/-) thymic cells. Coincidentally, Ets1 associates more specifically with Runx1 in DN and with TCF1 in DP cells. We also provide evidence that Ets1 predominantly binds distal nucleosome-occupied regions in DN and nucleosome-depleted regions in DP. Finally and importantly, we demonstrate that Ets1 induces chromatin remodeling by displacing H3K4me1-marked nucleosomes. Our results thus provide an original model whereby the ability of a transcription factor to bind nucleosomal DNA changes during differentiation with consequences on its cognate enhancer activity.
