Effect of knockdown PHF8 on the transcriptional regulation of BCR-ABL1 fusion gene on CML cell line K562

Epigenetic dysregulation plays a vital role in the pathogenesis of hematological malignancies. Histone demethylation is a key mechanism for epigenetic modification, of which dysregulation has an immeasurable effect on both TKI resistance and the transformation from CP CML to AP/BC CML. Here we report that PHF8 regulate the transcription of the driven fusion gene BCR-ABL1 in CML cells. The depletion of PHF8 promotes differentiation and inhibits proliferation. Furthermore, the proliferation-inhibited function of PHF8-knockdown have stronger effect on imatinib mesylate (IM)-resistant CML cells. We identified that PHF8 as a transcriptional modulator interacted with the promoter of the BCR::ABL1 fusion gene and alters the methylation levels of H3K9me1, H3K9me2 and H3K27me1, thereby promoting BCR::ABL1 transcription. Taken together, PHF8, as an oncogenic protein, was abnormally expressed in CML, altered the histone methylation levels of BCR::ABL1 promoter, and upregulated BCR::ABL1. Our study indicated that regulating histone demethylation is essential for BCR::ABL1 expression. Targeting PHF8 can be a promising therapeutic option for CML patients with TKI resistance. Overall design: To investigate the mechanism and function of PHF8 in CML, we constructed the PHF8 knockdown CML cell lines with shRNA in K562. Than we performed gene expression profiling analysis using data obtained from RNA-seq of K562 with shNC and shPHF8.

表观遗传失调在血液系统恶性肿瘤的发病机制中发挥关键作用。组蛋白去甲基化是表观遗传修饰的核心机制之一，其失调对酪氨酸激酶抑制剂（TKI）耐药以及慢性期慢性髓系白血病（CP CML）向加速期/急变期慢性髓系白血病（AP/BC CML）的转化均具有不可估量的影响。本研究证实，PHF8可调控慢性髓系白血病（CML）细胞中驱动性融合基因BCR-ABL1的转录。敲低PHF8可促进细胞分化并抑制增殖。进一步研究发现，PHF8敲低的增殖抑制作用对甲磺酸伊马替尼（IM）耐药的CML细胞效果更为显著。我们明确，PHF8作为转录调控因子，可与BCR::ABL1融合基因的启动子区域结合，并改变H3K9me1、H3K9me2及H3K27me1的甲基化水平，进而促进BCR-ABL1的转录。综上，PHF8作为致癌蛋白，在慢性髓系白血病中异常表达，通过改变BCR-ABL1基因启动子的组蛋白甲基化水平，上调BCR-ABL1的表达。本研究表明，调控组蛋白去甲基化对于BCR-ABL1的表达至关重要，靶向PHF8可为存在TKI耐药的CML患者提供极具潜力的治疗策略。整体实验设计：为探究PHF8在慢性髓系白血病中的作用机制与功能，我们在K562细胞中通过短发夹RNA（shRNA）构建了PHF8敲低的CML细胞系。随后，利用阴性对照短发夹RNA（shNC）与shPHF8处理的K562细胞的RNA测序（RNA-seq）数据，开展基因表达谱分析。