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The promyelocytic leukemia (PML)/retinoic acid receptor-alpha (RARα) onco-fusion protein that is generated from t(15;17) chromosome translocation is crucial for the leukemogenesis of acute promyelocytic leukemia (APL) and is well documented as a transcriptional repressor. To understand the relationship between PML/RARα and the oncogene in the development of APL, we investigate the regulation mechanism of PML/RARα to MYB proto-oncogene and the role of this regulation on the proliferation and differentiation of APL cells. Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) assays show that MYB expression was significantly higher in PML/RARα positive cell lines. Microarray data verify that the MYB expression was significantly higher in APL patient samples than in normal promyelocyte samples. Further expression analysis from RT-qPCR and microarray data verifies that the expression of MYB is upregulated by PML/RARα. Transcriptional factor binding analysis shows that MYB is directly bound by PML/RARα and its cofactors. Luciferase assays show that PML/RARα transactivated MYB promoter activity through the RARα binding site and the coexistence of CCAAT enhancer binding protein ε. We also find that PML/RARα increases the acetylation level of the promoter region of MYB. Further evidence demonstrates that PML/RARα regulates MYB expression through long-range interaction. Functionally, PML/RARα increases the cell proliferation and blocks the differentiation through activating MYB expression. Collectively, this study uncovers a novel mechanism of PML/RARα-mediated transcriptional activation and enriches our knowledge of the onco-fusion protein-mediated transcription activation.

The AML1/ETO onco-fusion protein is crucial for the genesis of t(8;21) acute myeloid leukemia (AML) and is well documented as a transcriptional repressor through dominant-negative effect. However, little is known about the transactivation mechanism of AML1/ETO. Through large cohort of patient's expression level data analysis and a series of experimental validation, we report here that AML1/ETO transactivates c-KIT expression through directly binding to and mediating the long-range interaction between the promoter and intronic enhancer regions of c-KIT. Gene expression analyses verify that c-KIT expression is significantly high in t(8;21) AML. Further ChIP-seq analysis and motif scanning identify two regulatory regions located in the promoter and intronic enhancer region of c-KIT, respectively. Both regions are enriched by co-factors of AML1/ETO, such as AML1, CEBPe, c-Jun, and c-Fos. Further luciferase reporter assays show that AML1/ETO trans-activates c-KIT promoter activity through directly recognizing the AML1 motif and the co-existence of co-factors. The induction of c-KIT promoter activity is reinforced with the existence of intronic enhancer region. Furthermore, ChIP-3C-qPCR assays verify that AML1/ETO mediates the formation of DNA-looping between the c-KIT promoter and intronic enhancer region through the long-range interaction. Collectively, our data uncover a novel transcriptional activity mechanism of AML1/ETO and enrich our knowledge of the onco-fusion protein mediated transcription regulation.