乳腺癌细胞基因组内G-四链体的形成特征及调节因素研究

G-四链体控制癌变功能基因的表达，但乳腺癌细胞基因组G-四链体形成特征及调节因素的研究还很匮乏。我们基于以往研究提出假想：乳腺癌细胞中癌变调节基因启动子的G-四链体形成不同于正常细胞；微环境和生理条件及某些天然化合物可以影响G-四链体形成来调节基因表达。本项目有三个研究目标：1. 用染色质免疫共沉淀法鉴定和比较正常乳腺细胞及乳腺癌细胞基因组中G-四链体的分布，并对应转录组数据以发现G-四链体形成与基因表达相关性。2. 研究微环境及生理因素（缺氧和pH值改变）对乳腺癌细胞基因组G-四链体形成及原癌基因表达影响，分析二者对应关系。3. 利用体外实验筛选出能够稳定G-四链体结构或抑制其解旋酶活性的天然化合物，并研究对细胞内G-四链体形成影响及抑癌活性。本项目将系统性地阐明乳腺癌细胞基因组G-四链体的形成特征，研究调节G-四链体形成的微环境及生理因素和天然化合物，为乳腺癌病人临床治疗提供理论依据。

G-quadruplexes regulate the expression of genes involved in oncogenesis. Currently, gaps still exist regarding our knowledge for the features of G-quadruplex formation in the genome of breast cancer cells and its regulatory factors. Based on previous studies, we hypothesized that the landscapes of G-quadruplex formation in the promoters of cancer-regulatory genes in breast cancer cells are different from these in normal mammary epithelial cells; physiological conditions, microenvironmental factors and certain natural compounds may affect G-quadruplex formation and consequently regulate gene expression. This project has three specific aims. In Aim 1, we will use chromatin immunoprecipitation (ChIP) to map the regions of G-quadruplex formation in the genomes of normal mammary epithelial cells and breast cancer cells, and determine the G-quadruplex formation difference between these two cell lines. In addition, we will use the RNA-seq technique to determine the transcriptomes of these two cell lines, and individually correlate the G-quadruplex formation sections and transcription regions in each cell line. In Aim 2, we will investigate the effects of microenvironmental and physiological factors, including hypoxia and pH changes, on G-quadruplex formation and oncogene expression in breast cancer cells. In Aim 3, we will carry out in vitro assays to screen for natural compounds that can either stabilize G-quadruplex structures or inhibit the activities of helicases with that can resolve G-quadruplexes. Overall, this project will systematically characterize G-quadruplex formation in the genome of breast cancer cells, and determine the effects of physiological conditions, microenvironmental factors and natural compounds on G-quadruplex formation. The achievements of this project will provide theoretical insights to the clinical treatment of breast cancer patients.