Liu Shujun, Shen Tiansheng, Huynh Lenguyen, Klisovic Marko I, Rush Laura J, Ford Jamie L, Yu Jianhua, Becknell Brian, Li Yu, Liu Chunhui, Vukosavljevic Tamara, Whitman Susan P, Chang Kun-Sang, Byrd John C, Perrotti Danilo, Plass Christoph, Marcucci Guido
Divisions of Hematology-Oncology, Department of Internal Medicine and the Comprehensive Cancer Center, Ohio State University, Columbus, Ohio, USA.
Cancer Res. 2005 Feb 15;65(4):1277-84. doi: 10.1158/0008-5472.CAN-04-4532.
The translocation t(8;21)(q22;q22) in acute myeloid leukemia (AML) results in the expression of the fusion protein RUNX1/MTG8, which in turn recruits histone deacetylases (HDAC) to silence RUNX1 target genes [e.g., interleukin-3 (IL-3)]. We previously reported that expression of the RUNX1/MTG8 target gene IL-3 is synergistically restored by the combination of inhibitors of HDACs (i.e., depsipeptide) and DNA methyltransferases (DNMT; i.e., decitabine) in RUNX1/MTG8-positive Kasumi-1 cells. Thus, we hypothesized that DNMT1 is also part of the transcriptional repressor complex recruited by RUNX1/MTG8. By a chromatin immunoprecipitation assay, we identified a RUNX1/MTG8-DNMT1 complex on the IL-3 promoter in Kasumi-1 cells and in primary RUNX1/MTG8-positive AML blasts. The physical association of RUNX1/MTG8 with DNMT1 was shown by coimmunoprecipitation experiments. Furthermore, RUNX1/MTG8 and DNMT1 were concurrently released from the IL-3 promoter by exposure to depsipeptide or stabilized on the promoter by decitabine treatment. Finally, we proved that RUNX1/MTG8 and DNMT1 were functionally interrelated by showing an enhanced repression of IL-3 after coexpression in 293T cells. These results suggest a novel mechanism for gene silencing mediated by RUNX1/MTG8 and support the combination of HDAC and DNMT inhibitors as a novel therapeutic approach for t(8;21) AML.
急性髓系白血病（AML）中的t(8;21)(q22;q22)易位导致融合蛋白RUNX1/MTG8的表达，进而募集组蛋白脱乙酰酶（HDAC）使RUNX1靶基因沉默[如白细胞介素-3（IL-3）]。我们之前报道，在RUNX1/MTG8阳性的Kasumi-1细胞中，HDAC抑制剂（即缩肽）和DNA甲基转移酶（DNMT；即地西他滨）联合使用可协同恢复RUNX1/MTG8靶基因IL-3的表达。因此，我们推测DNMT1也是RUNX1/MTG8募集的转录抑制复合物的一部分。通过染色质免疫沉淀试验，我们在Kasumi-1细胞和原发性RUNX1/MTG8阳性AML母细胞的IL-3启动子上鉴定出RUNX1/MTG8-DNMT1复合物。免疫共沉淀实验表明RUNX1/MTG8与DNMT1存在物理关联。此外，通过缩肽处理可使RUNX1/MTG8和DNMT1同时从IL-3启动子上释放，而地西他滨处理则可使其稳定结合在启动子上。最后，我们通过在293T细胞中共表达后显示出IL-3抑制增强，证明RUNX1/MTG8和DNMT1在功能上相互关联。这些结果提示了RUNX1/MTG8介导的基因沉默的新机制，并支持将HDAC和DNMT抑制剂联合使用作为t(8;21) AML的一种新治疗方法。
