细胞内转录因子高分辨率DNA结合靶点鉴定新技术的发展及其应用研究

Identification of the precise DNA targets and target genes of transcription factors in cells is important for elucidating the molecular mechanisms underline the physiological and pathological functions of transcription factors, and finding important DNA targets and target genes with a value of biomedical application. Chromatin immunoprecipitation-sequencing (ChIP-Seq) is the current most effective method for identifying the intracellular DNA-binding targets of transcription factors; however, this technique can determine the approximate binding location, but can’t identify the precise DNA binding targets of transcription factors in the genomic DNA. For this scientific problem, this project will develop a new ChIP-Seq technology based on exonuclease III (ExoIII), and identify the accurate DNA binding targets and build the high-resolution DNA binding profile of a important transcription factor, NF-κB, in the human hepatoma cells by using this technique. In combination with the genes whose expression can repond to the change of NF-κB activity in the human hepatoma cells identified by gene expression profiling techniques, this project will identified the target genes of the transcription factor in the human hepatoma cell. This project has the important significance in the construction of the target gene regulatory networks of NF-κB in the human hepatoma cells, elucidation of the molecular mechanisms underlying the pathological functions of NF-κB in the human hepatoma cells, and finding of the DNA targets and target genes with hepatoma diagnostic and therapeutic values.

鉴定转录因子在细胞中的精确DNA靶点及靶基因，对于解析转录因子发挥其生理及病理作用的分子机制、筛查具有生物医学应用价值的DNA靶点和靶基因具有重要意义。ChIP-Seq技术是目前鉴定转录因子在细胞内DNA结合靶点最有效的方法，但该技术只能确定转录因子在基因组DNA中的大致结合位置，而不能鉴定其精确的DNA结合靶点。针对这一问题，本项目将发展ExoIII-ChIP-Seq新技术，并用其鉴定转录因子NF-κB在人肝癌细胞中的精确DNA结合靶点、绘制其高分辨率DNA结合谱，进而鉴定NF-κB在人肝癌细胞中的靶基因。本研究在构建NF-κB在肝癌中细胞中的靶基因调控网络、阐明NF-κB在人肝癌细胞中发挥其病理作用的分子机制、发现具有肝癌诊断及治疗价值的DNA靶点及靶基因方面具有重要意义。

鉴定转录因子在细胞中的精确DNA靶点及靶基因，对于解析转录因子发挥其生理及病理作用的分子机制、筛查具有生物医学应用价值的DNA靶点和靶基因具有重要意义。ChIP-Seq技术是目前鉴定转录因子在细胞内DNA结合靶点最有效的方法，但该技术只能确定转录因子在基因组DNA中的大致结合位置，而不能鉴定其精确的DNA结合靶点。针对这一问题，本项目将发展ExoIII-ChIP-Seq新技术，并用其鉴定转录因子NF-κB在人肝癌细胞中的精确DNA结合靶点、绘制其高分辨率DNA结合谱，进而鉴定NF-κB在人肝癌细胞中的靶基因。本研究在构建NF-κB在肝癌中细胞中的靶基因调控网络、阐明NF-κB在人肝癌细胞中发挥其病理作用的分子机制、发现具有肝癌诊断及治疗价值的DNA靶点及靶基因方面具有重要意义。