Dai Wei, Wu Jian, Zhang Shuyan, Shi Bingjie, Xu Xinhui, Wang Danyang, Wang Jinke
State Key Laboratory of Bioelectronics, Southeast University, Nanjing 210096, China.
State Key Laboratory of Bioelectronics, Southeast University, Nanjing 210096, China.
Int J Biochem Cell Biol. 2017 Aug;89:157-170. doi: 10.1016/j.biocel.2017.05.031. Epub 2017 Jun 1.
It has been well-known that over activation of NF-κB has close relationship with hepatitis and hepatocellular carcinoma (HCC). However, the complete and exact underlying molecular pathways and mechanisms still remain not fully understood. By manipulating NF-κB activity with its recognized activator TNFα and using ChIP-seq and RNA-seq techniques, this study identified 699 NF-κB direct target genes (DTGs) in a widely used HCC cell line, HepG2, including 399 activated and 300 repressed genes. In these NF-κB DTGs, 216 genes (126 activated and 90 repressed genes) are among the current HCC gene signature. In comparison with NF-κB target genes identified in LPS-induced THP-1 and TNFα-induced HeLa cells, only limited numbers (24-46) of genes were shared by the two cell lines, indicating the HCC specificity of identified genes. Functional annotation revealed that NF-κB DTGs in HepG2 cell are mainly related with many typical NF-κB-related biological processes including immune system process, response to stress, response to stimulus, defense response, and cell death, and signaling pathways of MAPK, TNF, TGF-beta, Chemokine, NF-kappa B, and Toll-like receptor. Some NF-κB DTGs are also involved in Hepatitis C and B pathways. It was found that 82 NF-κB DTGs code secretory proteins, which include CCL2 and DKK1 that have already been used as HCC markers. Finally, the NF-κB DTGs were further confirmed by detecting the NF-κB binding and expression of 14 genes with ChIP-PCR and RT-PCR. This study thus provides a useful NF-κB DTG list for future studies of NF-κB-related molecular mechanisms and theranostic biomarkers of HCC.
众所周知，NF-κB的过度激活与肝炎和肝细胞癌（HCC）密切相关。然而，其完整且确切的潜在分子途径和机制仍未完全清楚。通过用其公认的激活剂TNFα操纵NF-κB活性，并使用ChIP-seq和RNA-seq技术，本研究在广泛使用的HCC细胞系HepG2中鉴定出699个NF-κB直接靶基因（DTG），其中包括399个激活基因和300个抑制基因。在这些NF-κB DTG中，有216个基因（126个激活基因和90个抑制基因）属于当前的HCC基因特征。与在LPS诱导的THP-1和TNFα诱导的HeLa细胞中鉴定出的NF-κB靶基因相比，这两种细胞系仅共享有限数量（24 - 46个）的基因，表明所鉴定基因的HCC特异性。功能注释显示，HepG2细胞中的NF-κB DTG主要与许多典型的NF-κB相关生物学过程有关，包括免疫系统过程、应激反应、刺激反应、防御反应和细胞死亡，以及MAPK、TNF、TGF-β、趋化因子、NF-κB和Toll样受体的信号通路。一些NF-κB DTG也参与丙型肝炎和乙型肝炎途径。研究发现，82个NF-κB DTG编码分泌蛋白，其中包括已被用作HCC标志物的CCL2和DKK1。最后，通过ChIP-PCR和RT-PCR检测14个基因的NF-κB结合和表达，进一步证实了NF-κB DTG。因此，本研究为未来研究NF-κB相关分子机制和HCC的诊疗生物标志物提供了一份有用的NF-κB DTG清单。
