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
Hematopoietic stem cells (HSCs) give rise to all blood populations due to their long-term self-renewal and multipotent differentiation capacities. Because they have to persist throughout an organism's life span, HSCs tightly regulate the balance between proliferation and quiescence. Here, we investigated the role of the transcription factor promyelocytic leukemia zinc finger (plzf) in HSC fate using the Zbtb16(lu/lu)mouse model, which harbors a natural spontaneous mutation that inactivates plzf. Regenerative stress revealed that Zbtb16(lu/lu)HSCs had a lineage-skewing potential from lymphopoiesis toward myelopoiesis, an increase in the long-term-HSC pool, and a decreased repopulation potential. Furthermore, oldplzf-mutant HSCs present an amplified aging phenotype, suggesting that plzf controls age-related pathway. We found that Zbtb16(lu/lu)HSCs harbor a transcriptional signature associated with a loss of stemness and cell cycle deregulation. Lastly, cell cycle analyses revealed an important role for plzf in the regulation of the G1-S transition of HSCs. Our study reveals a new role for plzf in regulating HSC function that is linked to cell cycle regulation, and positions plzf as a key player in controlling HSC homeostasis.
Asunto(s)
Regulación del Desarrollo de la Expresión Génica , Células Madre Hematopoyéticas/citología , Factores de Transcripción de Tipo Kruppel/genética , Factores de Transcripción de Tipo Kruppel/fisiología , Mutación , Animales , Apoptosis , Ciclo Celular , Diferenciación Celular , Linaje de la Célula , Senescencia Celular , Epigénesis Genética , Perfilación de la Expresión Génica , Homeostasis , Linfopoyesis , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Mielopoyesis , Análisis de Secuencia por Matrices de Oligonucleótidos , Fenotipo , Proteína de la Leucemia Promielocítica con Dedos de ZincRESUMEN
The PLZF/RARA fusion protein generated by the t(11;17)(q23;q21) translocation in acute promyelocytic leukaemia (APL) is believed to act as an oncogenic transcriptional regulator recruiting epigenetic factors to genes important for its transforming potential. However, molecular mechanisms associated with PLZF/RARA-dependent leukaemogenesis still remain unclear.We searched for specific PLZF/RARA target genes by ChIP-on-chip in the haematopoietic cell line U937 conditionally expressing PLZF/RARA. By comparing bound regions found in U937 cells expressing endogenous PLZF with PLZF/RARA-induced U937 cells, we isolated specific PLZF/RARA target gene promoters. We next analysed gene expression profiles of our identified target genes in PLZF/RARA APL patients and analysed DNA sequences and epigenetic modification at PLZF/RARA binding sites. We identify 413 specific PLZF/RARA target genes including a number encoding transcription factors involved in the regulation of haematopoiesis. Among these genes, 22 were significantly down regulated in primary PLZF/RARA APL cells. In addition, repressed PLZF/RARA target genes were associated with increased levels of H3K27me3 and decreased levels of H3K9K14ac. Finally, sequence analysis of PLZF/RARA bound sequences reveals the presence of both consensus and degenerated RAREs as well as enrichment for tissue-specific transcription factor motifs, highlighting the complexity of targeting fusion protein to chromatin. Our study suggests that PLZF/RARA directly targets genes important for haematopoietic development and supports the notion that PLZF/RARA acts mainly as an epigenetic regulator of its direct target genes.
