Zhou Hua-Rong, Fu Hai-Ying, Wu Dan-Sen, Zhang Yuan-Yuan, Huang Si-Han, Chen Cong-Jie, Yan Jian-Guo, Huang Jin-Long, Shen Jian-Zhen
Fujian Institute of Hematology, Fujian Provincial Key Laboratory of Hematology, Fujian Medical University Union Hospital, Fuzhou, Fujian 350001, P.R. China.
Oncol Rep. 2017 May;37(5):2663-2671. doi: 10.3892/or.2017.5509. Epub 2017 Mar 15.
The present study was designed to investigate the relationship among epigenetic changes in Wnt antagonists, histone H4K20me1 and the expression of tumor-suppressor genes in acute leukemia (AL) to better understand the pathogenesis of leukemia. Quantitative reverse transcription polymerase chain reaction (qRT-PCR) was performed to detect messenger RNA (mRNA) expression levels of Wnt antagonists (Wnt5a, HDPR1, DKK1 and DKK3) in patients with AL and in normal controls; pyrophosphate sequencing was performed to detect the methylation status of the Wnt5a promoter; and western blotting was performed to detect the overall expression levels of Wnt5a protein and histone H4K20me1 in patients with acute myeloid leukemia (AML) and in normal controls. The relationship between Wnt5a protein expression and histone H4K20me1 was analyzed. Chromatin immunoprecipitation-qPCR (ChIP-qPCR) was performed to investigate the recruitment of H4K20me1 and SET8 to the Wnt5a promoter and coding regions. Our results demonstrated that the expression levels of Wnt antagonists were generally low in AML, but showed differential expression in acute lymphocytic leukemia (ALL). In most cases of AML, methylation of the Wnt5a promoter was observed and Wnt5a protein expression was low. In some cases of AML, the overall level of H4K20me1 protein was higher than that in normal controls. In addition, Wnt5a expression was positively correlated with H4K20me1 expression and was unrelated to the methylation status of its promoter. Moreover, H4K20me1 and SET8 were enriched in the Wnt5a promoter region and coding region. By contrast, Wnt5a expression was unrelated to H4K20me1 expression in normal controls. Moreover, we observed that the methylation of Wnt antagonists was often found in patients with AL, particularly those with AML, whereas the extent of methylation was variable in ALL patients. Wnt5a expression was positively correlated with the enrichment of H4K20me1 and SET8 at the Wnt5a promoter and coding regions. H4K20me1 increased Wnt5a expression by promoting transcription initiation and elongation.
本研究旨在探讨急性白血病（AL）中Wnt拮抗剂的表观遗传变化、组蛋白H4K20me1与肿瘤抑制基因表达之间的关系，以更好地理解白血病的发病机制。采用定量逆转录聚合酶链反应（qRT-PCR）检测AL患者和正常对照中Wnt拮抗剂（Wnt5a、HDPR1、DKK1和DKK3）的信使核糖核酸（mRNA）表达水平；采用焦磷酸测序检测Wnt5a启动子的甲基化状态；采用蛋白质免疫印迹法检测急性髓系白血病（AML）患者和正常对照中Wnt5a蛋白和组蛋白H4K20me1的整体表达水平。分析Wnt5a蛋白表达与组蛋白H4K20me1之间的关系。采用染色质免疫沉淀-qPCR（ChIP-qPCR）研究H4K20me1和SET8在Wnt5a启动子和编码区的募集情况。我们的结果表明，Wnt拮抗剂的表达水平在AML中普遍较低，但在急性淋巴细胞白血病（ALL）中表现出差异表达。在大多数AML病例中，观察到Wnt5a启动子甲基化且Wnt5a蛋白表达较低。在一些AML病例中，H4K20me1蛋白的整体水平高于正常对照。此外，Wnt5a表达与H4K20me1表达呈正相关，与其启动子的甲基化状态无关。而且，H4K20me1和SET8在Wnt5a启动子区域和编码区富集。相比之下，正常对照中Wnt5a表达与H4K20me1表达无关。此外，我们观察到Wnt拮抗剂的甲基化在AL患者中经常出现，尤其是AML患者，而ALL患者的甲基化程度则有所不同。Wnt5a表达与H4K20me1和SET8在Wnt5a启动子和编码区的富集呈正相关。H4K20me1通过促进转录起始和延伸增加Wnt5a表达。
