Genome-wide binding sites of NF90/ILF3 in K562 erythroleukemia cells. Homo sapiens

NF90 and splice variant NF110 are DNA- and RNA-binding proteins encoded by the Interleukin enhancer-binding factor 3 (ILF3) gene that regulate RNA splicing, stabilization and export. The role of NF90 in regulating transcription as a DNA-binding protein has not been comprehensively characterized. Here, ENCODE ChIP-seq identified 9,081 genomic targets specifically bound by NF90/110 in K562 cells. One third of binding sites occurred at promoters of annotated genes. NF90/110 binding colocalized with chromatin marks associated with active promoters and strong enhancers. Analysis using ENCODE ChIP-seq experiments revealed that NF90 clustered with transcription factors exhibiting preference for promoters over enhancers (POLR2A, MYC, YY1). Differential gene expression analysis following shRNA knockdown of NF90 in K562 cells revealed that NF90 directly activates transcription factors that drive growth and proliferation (EGR1, MYC), while attenuating differentiation along erythroid lineage (KLF1). NF90/110 interacts with chromatin to hierarchically regulate transcription factors to promote proliferation and suppress differentiation. Overall design: NF90 ChIP-seq in K562 cells

NF90及其剪接变体NF110是由白细胞介素增强子结合因子3（Interleukin enhancer-binding factor 3, ILF3）基因编码的DNA结合蛋白与RNA结合蛋白，可调控RNA的剪接、稳定及出核转运。目前，NF90作为DNA结合蛋白调控转录的功能尚未得到全面阐释。本研究通过DNA元件百科全书（Encyclopedia of DNA Elements, ENCODE）的染色质免疫共沉淀测序（Chromatin Immunoprecipitation Sequencing, ChIP-seq）技术，在K562细胞中鉴定出9081个被NF90/110特异性结合的基因组靶位点。其中三分之一的结合位点位于已注释基因的启动子区域。NF90/110的结合位点与活跃启动子及强增强子相关的染色质标记共定位。通过分析ENCODE的ChIP-seq实验数据，研究发现NF90与偏好结合启动子而非增强子的转录因子（POLR2A、MYC、YY1）聚集成簇。在K562细胞中通过短发夹RNA（short hairpin RNA, shRNA）敲低NF90后的差异基因表达分析显示，NF90可直接激活驱动细胞生长与增殖的转录因子（EGR1、MYC），同时抑制红系谱系的分化（KLF1）。NF90/110可与染色质相互作用，通过层级调控转录因子以促进细胞增殖并抑制分化。实验整体设计：在K562细胞中开展NF90的ChIP-seq实验