染色质三维结构对RNAa在CHO细胞中激活表达效率的影响

RNAa(RNA activation)通过小RNA靶向启动子激活基因表达，在疾病治疗和高效表达等领域中具有应用潜力，但其激活机制仍未完全清楚。本项目用小RNA靶向染色质调节序列UCOE和MAR调节的含增强子或CpG岛的启动子，以构建可致染色质三维结构变化的载体模型。拟采用ChIA-PET和4C等染色质三维分析技术，在CHO细胞中开展该模型上MAR与宿主染色质形成的染色质LOOP、UCOE与染色质的相互作用、以及激活小RNA靶向后的启动子与全基因组中其它启动子的相互作用研究，探讨染色质三维结构对RNAa激活效率的影响。本项目有助于认识染色质三维结构在RNAa机制中的作用，为高效表达载体的构建等应用奠定基础。

RNAa (RNA activation) is a method of gene activation that uses promoter targeting small RNAs. The RNA used in this method is known as saRNA (small activating RNA). RNAa has potential applications in treatment of various diseases and construction of high expression vectors. However, the underlying mechanism of its action is still not fully understood. In previous studies, we explored the role of open chromatin in the functional mechanism of RNAa, where saRNA was used to target promoters of enhancer-containing or CpG island-containing genes, which are regulated by chromatin regulatory elements, such as UCOE (Ubiquitous Chromatin-opening Element) and MAR (Matrix Attachment Region), respectively. The results showed modulated expression of target genes compared with RNAa treatment alone. As both, UCOE and MAR, can interact with the host chromatin, the results indicated that the 3D chromatin structure played an important role in RNAa binding and effect. The aim of this project is to study the interaction between the UCOE or MAR and host chromatin, as well as the interaction between promoters targeted by RNAa and other promoters. The techniques of ChIA-PET (Chromatin Interaction Analysis using Paired-End Tag sequencing) and 4C (circular chromatin conformation capture) will be used to elucidate the relationship between the 3D chromatin structure and RNAa efficiency. The implementation of this project will help to integrate the 3D chromatin and open chromatin mechanisms of RNAa, the latter was explored in our previous project, and lay the foundation for the construction of efficient expression vectors.