PU.1 activation relieves GATA-1-mediated repression of Cebpa and Cbfb during leukemia differentiation.
Hematopoietic transcription factors GATA-1 and PU.1 bind each other on DNA to block transcriptional programs of undesired lineage during hematopoietic commitment. Murine erythroleukemia (MEL) cells that coexpress GATA-1 and PU.1 are blocked at the blast stage but respond to molecular removal (downregulation) of PU.1 or addition (upregulation) of GATA-1 by inducing terminal erythroid differentiation. To test whether GATA-1 blocks PU.1 in MEL cells, we have conditionally activated a transgenic PU.1 protein fused with the estrogen receptor ligand-binding domain (PUER), resulting in activation of a myeloid transcriptional program. Gene expression arrays identified components of the PU.1-dependent transcriptome negatively regulated by GATA-1 in MEL cells, including CCAAT/enhancer binding protein alpha (Cebpa) and core-binding factor, beta subunit (Cbfb), which encode two key hematopoietic transcription factors. Inhibition of GATA-1 by small interfering RNA resulted in derepression of PU.1 target genes. Chromatin immunoprecipitation and reporter assays identified PU.1 motif sequences near Cebpa and Cbfb that are co-occupied by PU.1 and GATA-1 in the leukemic blasts. Significant derepression of Cebpa and Cbfb is achieved in MEL cells by either activation of PU.1 or knockdown of GATA-1. Furthermore, transcriptional regulation of these loci by manipulating the levels of PU.1 and GATA-1 involves quantitative increases in a transcriptionally active chromatin mark: acetylation of histone H3K9. Collectively, we show that either activation of PU.1 or inhibition of GATA-1 efficiently reverses the transcriptional block imposed by GATA-1 and leads to the activation of a myeloid transcriptional program directed by PU.1.
Burda
Pavel
P
Institute of Pathological Physiology and Center of Experimental Hematology, First Faculty of Medicine, Charles University in Prague, Prague, Czech Republic.
Curik
Nikola
N
Kokavec
Juraj
J
Basova
Petra
P
Mikulenkova
Dana
D
Skoultchi
Arthur I
AI
Zavadil
Jiri
J
Stopka
Tomas
T
D022762:CCAAT-Enhancer-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D002454:Cell Differentiation / Q000235:genetics; D002471:Cell Transformation, Neoplastic / Q000235:genetics* / Q000378:metabolism; D050658:Core Binding Factor beta Subunit / Q000235:genetics* / Q000378:metabolism; D050982:GATA1 Transcription Factor / Q000235:genetics* / Q000378:metabolism; D015972:Gene Expression Regulation, Neoplastic / Q000235:genetics; D006367:HeLa Cells; D006657:Histones / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007938:Leukemia / Q000235:genetics* / Q000378:metabolism / Q000503:physiopathology; D022423:Myeloid Cells / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000235:genetics*; D034622:RNA Interference; D034741:RNA, Small Interfering; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D050436:Regulatory Elements, Transcriptional / Q000235:genetics; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism; D015534:Trans-Activators / Q000235:genetics*; D015533:Transcriptional Activation / Q000235:genetics; D000099232:Proto-Oncogene Protein Spi-1
D022762:CCAAT-Enhancer-Binding Proteins; D050658:Core Binding Factor beta Subunit; D050982:GATA1 Transcription Factor; D006657:Histones; D011518:Proto-Oncogene Proteins; D034741:RNA, Small Interfering; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; D015534:Trans-Activators; D000099232:Proto-Oncogene Protein Spi-1; C493692:CBFB protein, human; C515848:CEBPA protein, human; C494689:Gata1 protein, mouse