School of Medicine, Loma Linda University, Loma Linda, CA 92350, USA.
College of Medicine, The Pennsylvania State University, Hershey, PA 17033, USA.
Int J Mol Sci. 2020 Mar 3;21(5):1718. doi: 10.3390/ijms21051718.
Ikaros is a DNA-binding protein that regulates gene expression and functions as a tumor suppressor in B-cell acute lymphoblastic leukemia (B-ALL). The full cohort of Ikaros target genes have yet to be identified. Here, we demonstrate that Ikaros directly regulates expression of the small GTPase, Rab20. Using ChIP-seq and qChIP we assessed Ikaros binding and the epigenetic signature at the RAB20 promoter. Expression of Ikaros, CK2, and was determined by qRT-PCR. Overexpression of Ikaros was achieved by retroviral transduction, whereas shRNA was used to knockdown Ikaros and CK2. Regulation of transcription from the promoter was analyzed by luciferase reporter assay. The results showed that Ikaros binds the promoter in B-ALL. Gain-of-function and loss-of-function experiments demonstrated that Ikaros represses transcription via chromatin remodeling. Phosphorylation by CK2 kinase reduces Ikaros' affinity toward the promoter and abolishes its ability to repress transcription. Dephosphorylation by PP1 phosphatase enhances both Ikaros' DNA-binding affinity toward the promoter and repression. In conclusion, the results demonstrated opposing effects of CK2 and PP1 on expression of Rab20 via control of Ikaros' activity as a transcriptional regulator. A novel regulatory signaling network in B-cell leukemia that involves CK2, PP1, Ikaros, and Rab20 is identified.
Ikaros 是一种 DNA 结合蛋白，可调节基因表达，并在 B 细胞急性淋巴细胞白血病 (B-ALL) 中作为肿瘤抑制因子发挥作用。Ikaros 的全部靶基因尚未确定。在这里，我们证明 Ikaros 可直接调节小 GTPase Rab20 的表达。我们使用 ChIP-seq 和 qChIP 评估了 Ikaros 结合和 RAB20 启动子上的表观遗传特征。通过 qRT-PCR 测定 Ikaros、CK2 和 的表达。通过逆转录病毒转导实现 Ikaros 的过表达，而 shRNA 用于敲低 Ikaros 和 CK2。通过荧光素酶报告基因分析研究了 启动子转录的调节。结果表明，Ikaros 在 B-ALL 中结合 启动子。功能获得和功能丧失实验表明，Ikaros 通过染色质重塑抑制 转录。CK2 激酶的磷酸化降低了 Ikaros 对 启动子的亲和力，并使其丧失抑制 转录的能力。PP1 磷酸酶的去磷酸化增强了 Ikaros 与 启动子的 DNA 结合亲和力和 抑制作用。总之，结果表明 CK2 和 PP1 通过控制 Ikaros 作为转录调节剂的活性对 Rab20 的表达产生相反的影响。鉴定了涉及 CK2、PP1、Ikaros 和 Rab20 的 B 细胞白血病中的新型调节信号网络。
