The RUNX1-ETO fusion protein trans-activates c-KIT expression by recruiting histone acetyltransferase P300 on its promoter.

The oncoprotein RUNX1-ETO is the fusion product of t(8;21)(q22;q22) and constitutes one of the most common genetic alterations in acute myeloid leukemia (AML). Abnormal c-KIT overexpression is considered an independent negative prognostic factor for relapse and survival in t(8;21) AML patients. However, the molecular mechanism of high c-KIT expression in t(8;21) AML remains unknown. In this study, we detected RUNX1-ETO and c-KIT gene expression in AML-M2 patients and verified the overexpression of c-KIT in t(8;21) AML patients. We also found that c-KIT overexpression was a poor prognostic indicator in RUNX1-ETO positive AML patients, but not in RUNX1-ETO negative AML patients. We used the dual-luciferase and ChIP assays to demonstrate that the RUNX1-ETO protein epigenetically trans-activates c-KIT by binding to the c-KIT promoter and recruiting the histone acetyltransferase P300 to the c-KIT promoter, elucidating the mechanism of the abnormally increased c-KIT expression in t(8;21) AML patients. Moreover, pharmacological studies revealed that C646, a P300 inhibitor, could inhibit proliferation, induce apoptosis and arrest the cell cycle more effectively in RUNX1-ETO positive cells than in negative ones. The levels of c-KIT and RUNX1-ETO proteins were also decreased with C646 treatment in RUNX1-ETO positive cells. These findings suggested that P300 could be a therapeutic target and that C646 could be used as a potential treatment for RUNX1-ETO positive AML patients. Interestingly, using the dual-luciferase assay, we also found that the binding capacity of RUNX1-ETO9a, a truncated RUNX1-ETO isoform, to the c-KIT promoter was stronger than that of RUNX1-ETO, suggesting RUNX1-ETO9a as another valuable therapeutic target in t(8;21) AML.

Chen G ，Liu A ，Xu Y ，Gao L ，Jiang M ，Li Y ，Lv N ，Zhou L ，Wang L ，Yu L ，Li Y ... -  《-》

A histone acetyltransferase p300 inhibitor C646 induces cell cycle arrest and apoptosis selectively in AML1-ETO-positive AML cells.

Gao XN ，Lin J ，Ning QY ，Gao L ，Yao YS ，Zhou JH ，Li YH ，Wang LL ，Yu L ... -  《PLoS One》

AML1-ETO triggers epigenetic activation of early growth response gene l, inducing apoptosis in t(8;21) acute myeloid leukemia.

The t(8;21)(q22;q22) translocation is the most common chromosomal translocation in acute myeloid leukemia (AML), and it gives rise to acute myeloid gene 1 (AML1)-myeloid transforming gene 8 (ETO)-positive AML, which has a relatively favorable prognosis. However, the molecular mechanism related to a favorable prognosis in AML1-ETO-positive AML is still not fully understood. Our results show that the AML1-ETO fusion protein triggered activation of early growth response gene l (EGR1) by binding at AML1-binding sites on the EGR1 promoter and, subsequently, recruiting acetyltransferase P300, which is known to acetylate histones. However, AML1-ETO could not recruit DNA methyltransferases and histone deacetylases; therefore, EGR1 expression was affected by histone acetylation but not by DNA methylation. Both transcription and translation of EGR1 were higher in AML1-ETO-positive AML cell lines than in AML1-ETO-negative AML cell lines, owing to acetylation. Furthermore, when AML1-ETO-positive AML cell lines were treated with C646 (P300 inhibitor) and trichostatin A (histone deacetylase inhibitor), EGR1 expression was significantly decreased and increased, respectively. In addition, treatment with 5-azacytidine (methyltransferase inhibitor) did not cause any significant change in EGR1 expression. Overexpression of EGR1 inhibited cell proliferation and promoted apoptosis, and EGR1 knockout promoted cell proliferation. Thus, EGR1 could be a novel prognostic factor for a favorable outcome in AML1-ETO-positive AML. The results of our study may explain the molecular mechanisms underlying the favorable prognosis in AML1-ETO-positive AML.

Fu L ，Huang W ，Jing Y ，Jiang M ，Zhao Y ，Shi J ，Huang S ，Xue X ，Zhang Q ，Tang J ，Dou L ，Wang L ，Nervi C ，Li Y ，Yu L ... -  《-》

High expression of c-kit mRNA predicts unfavorable outcome in adult patients with t(8;21) acute myeloid leukemia.

Gao X ，Lin J ，Gao L ，Deng A ，Lu X ，Li Y ，Wang L ，Yu L ... -  《PLoS One》

RUNX1-ETO Depletion in t(8;21) AML Leads to C/EBPα- and AP-1-Mediated Alterations in Enhancer-Promoter Interaction.

Acute myeloid leukemia (AML) is associated with mutations in transcriptional and epigenetic regulator genes impairing myeloid differentiation. The t(8;21)(q22;q22) translocation generates the RUNX1-ETO fusion protein, which interferes with the hematopoietic master regulator RUNX1. We previously showed that the maintenance of t(8;21) AML is dependent on RUNX1-ETO expression. Its depletion causes extensive changes in transcription factor binding, as well as gene expression, and initiates myeloid differentiation. However, how these processes are connected within a gene regulatory network is unclear. To address this question, we performed Promoter-Capture Hi-C assays, with or without RUNX1-ETO depletion and assigned interacting cis-regulatory elements to their respective genes. To construct a RUNX1-ETO-dependent gene regulatory network maintaining AML, we integrated cis-regulatory element interactions with gene expression and transcription factor binding data. This analysis shows that RUNX1-ETO participates in cis-regulatory element interactions. However, differential interactions following RUNX1-ETO depletion are driven by alterations in the binding of RUNX1-ETO-regulated transcription factors.

Ptasinska A ，Pickin A ，Assi SA ，Chin PS ，Ames L ，Avellino R ，Gröschel S ，Delwel R ，Cockerill PN ，Osborne CS ，Bonifer C ... -  《Cell Reports》