Indiana University Melvin and Bren Simon Cancer Center, Departments of Medicine (Hematology/Oncology Division), Indiana University School of Medicine, Indianapolis, IN 46202.
Veterans Affairs Medical Center, Indianapolis, IN 46202.
Clin Cancer Res. 2012 Jan 15;18(2):360-369. doi: 10.1158/1078-0432.CCR-10-3022. Epub 2011 Nov 17.
Death-associated protein kinase 1 (DAPK1), a tumor suppressor, is a rate-limiting effector in an endoplasmic reticulum (ER) stress-dependent apoptotic pathway. Its expression is epigenetically suppressed in several tumors. A mechanistic basis for epigenetic/transcriptional repression of DAPK1 was investigated in certain forms of acute myeloid leukemia (AML) with poor prognosis, which lacked ER stress-induced apoptosis.
Heterogeneous primary AMLs were screened to identify a subgroup with Flt3ITD in which repression of DAPK1, among NF-κB-and c-Jun-responsive genes, was studied. RNA interference knockdown studies were carried out in an Flt3ITD(+) cell line, MV-4-11, to establish genetic epistasis in the pathway Flt3ITD-TAK1-DAPK1 repression, and chromatin immunoprecipitations were carried out to identify proximate effector proteins, including TAK1-activated p52NF-κB, at the DAPK1 locus.
AMLs characterized by normal karyotype with Flt3ITD were found to have 10- to 100-fold lower DAPK1 transcripts normalized to the expression of c-Jun, a transcriptional activator of DAPK1, as compared with a heterogeneous cytogenetic category. In addition, Meis1, a c-Jun-responsive adverse AML prognostic gene signature was measured as control. These Flt3ITD(+) AMLs overexpress relB, a transcriptional repressor, which forms active heterodimers with p52NF-κB. Chromatin immunoprecipitation assays identified p52NF-κB binding to the DAPK1 promoter together with histone deacetylase 2 (HDAC2) and HDAC6 in the Flt3ITD(+) human AML cell line MV-4-11. Knockdown of p52NF-κB or its upstream regulator, NF-κB-inducing kinase (NIK), de-repressed DAPK1. DAPK1-repressed primary Flt3ITD(+) AMLs had selective nuclear activation of p52NF-κB.
Flt3ITD promotes a noncanonical pathway via TAK1 and p52NF-κB to suppress DAPK1 in association with HDACs, which explains DAPK1 repression in Flt3ITD(+) AML.
死亡相关蛋白激酶 1（DAPK1）是一种肿瘤抑制因子，是内质网（ER）应激依赖性凋亡途径中的限速效应因子。其在几种肿瘤中存在表观遗传抑制。在预后不良的某些形式的急性髓系白血病（AML）中，我们研究了导致 DAPK1 发生表观遗传/转录抑制的机制基础，这些 AML 缺乏 ER 应激诱导的凋亡。
对异质性原发性 AML 进行筛选，以确定一个亚组存在 Flt3ITD，在该亚组中，DAPK1 在 NF-κB 和 c-Jun 反应基因中受到抑制。在 Flt3ITD(+)细胞系 MV-4-11 中进行 RNA 干扰敲低研究，以确定 Flt3ITD-TAK1-DAPK1 抑制途径中的遗传上位效应，并进行染色质免疫沉淀实验以鉴定邻近的效应蛋白，包括 DAPK1 基因座上 TAK1 激活的 p52NF-κB。
与异质性细胞遗传学分类相比，具有正常核型且存在 Flt3ITD 的 AML 发现 DAPK1 转录本降低了 10 至 100 倍，相对于 c-Jun 的表达归一化，c-Jun 是 DAPK1 的转录激活剂。此外，还测量了 Meis1，这是一种与预后不良相关的 AML 基因标志，作为对照。这些 Flt3ITD(+)AML 过度表达 relB，这是一种转录抑制物，它与 p52NF-κB 形成活性异二聚体。染色质免疫沉淀实验鉴定出 p52NF-κB 与组蛋白去乙酰化酶 2（HDAC2）和 HDAC6 一起结合到 Flt3ITD(+)人 AML 细胞系 MV-4-11 中的 DAPK1 启动子上。敲低 p52NF-κB 或其上游调节因子 NF-κB 诱导激酶（NIK）可使 DAPK1 去抑制。Flt3ITD(+)原发性 AML 中 DAPK1 被选择性地激活。
Flt3ITD 通过 TAK1 和 p52NF-κB 促进非典型途径，与 HDACs 一起抑制 DAPK1，这解释了 Flt3ITD(+)AML 中 DAPK1 的抑制。
