Function of GATA3-PARP1 complex in breast cancer during GATA3-mediated cellular reprogramming

GATA3-PARP1复合物在GATA3介导的细胞重编程过程中在乳腺癌中的功能

• 批准号: 10679740
• 负责人: Mikhala Cooper
• 金额: $ 4.77万
• 依托单位: UNIVERSITY OF NORTH DAKOTA
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-05-23 至 2025-11-22
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10679740
• 关键词: Address; Architecture; Binding; Binding Sites; Biology; Breast Cancer Cell; Breast Cancer Patient; Breast Cancer cell line; Cancer Biology; Cells; ChIP-seq; Chromatin; Chromatin Structure; Complex; DNA; DNA Repair; Data; Development; Disease; Doxycycline; Enhancers; Enzymes; Epigenetic Process; Epithelium; Essential Genes; Estrogen Receptor alpha; Eukaryotic Cell; GATA3 gene; Gene Activation; Gene Expression; Gene Expression Regulation; Genes; Genome; Genomic DNA; Goals; Histones; Homeostasis; Human Genome; Immune; Immunoprecipitation; Interruption; Knowledge; Learning; MDA MB 231; Maintenance; Mammary Neoplasms; Mass Spectrum Analysis; Measures; Mediating; Methods; Molecular; Mutate; Mutation; Nucleosomes; Pathway interactions; Phenotype; Poly(ADP-ribose) Polymerases; Property; Proteins; Research; Role; Site; Specificity; System; Testing; Tissues; Transcriptional Activation; Work; chromatin modification; cofactor; epithelial to mesenchymal transition; experimental study; innovation; insight; malignant breast neoplasm; mammary epithelium; next generation sequencing; novel; prevent; transcription factor

Project Summary GATA3 is a transcription factor that acts as a pioneer factor in breast cancer cells. However, the function of pioneer factors isn’t well understood. Specifically, how they activate silent chromatin is largely unknown. The long-term goal is to provide novel insights into the molecular mechanisms of pioneer factor- mediated cellular reprogramming at a chromatin level. More than 7 million GATA3 motifs exist in the human genome, but the experimental data from GATA3 ChIP- seq analysis indicates less than 1% of the motifs are occupied by the pioneer factor, GATA3. In addition to binding selectivity, pioneer factor action, including GATA3, is known to be site-specific (context-dependent), and they can only induce chromatin opening and enhancer formation at a subset of binding sites. The overall objective of this proposal is to identify the binding specificity of the pioneer factor, GATA3, in the genome and elucidate the impact of the interaction between GATA3 and chromatin-modification enzyme, Poly ADP-ribose polymerase 1 (PARP1), on gene activation. Our preliminary data strongly suggest that PARP1 is involved in such context-dependent action of GATA3. The central hypothesis is that GATA3-mediated cellular reprogramming is dependent on the PARP1 activity. The rationale for this study is that the identification of the roles of GATA3 and PARP1 interaction will provide novel molecular mechanisms underlying the site- specific expression of pioneer factor activities during cellular reprogramming. The central hypothesis will be tested by (1) investigating the function of PARP1 in the binding selectivity of GATA3 and (2) identifying the role(s) of GATA3-PARP1 interaction in gene activation. To test this hypothesis, I will use our unique DOX- inducible system in MDA-MB-231 basal breast cancer cells. In this system, exogenous GATA3 expression initiates mesenchymal-to-epithelial transition (MET). This system recapitulates GATA3 function in patient breast tumors since GATA3 expression is known to be higher in the luminal/epithelial breast tumors, and is essential for the maintenance of epithelial phenotypes. Under the first aim, we will investigate changes in GATA3 binding across the genome in the presence and absence of PARP1. The second aim will utilize next- generation sequencing to evaluate the effects that GATA3 and PARP1 have on the steps in pioneer factor- mediated cellular reprogramming. The proposed research is innovative because we are addressing the novel concept of context-dependent enhancer formation by a pioneer factor during cellular reprogramming. This research is significant because it will fill a critical gap in our knowledge about how pioneer factors mediate cell fate transition.

项目摘要 GATA 3是一种转录因子，在乳腺癌细胞中充当先锋因子。然而， 先锋因素并没有被很好地理解。具体来说，它们如何激活沉默的染色质在很大程度上取决于 未知长期目标是提供新的见解的分子机制的先锋因子- 在染色质水平介导细胞重编程。 人类基因组中存在超过700万个GATA 3基序，但来自GATA 3 ChIP的实验数据- seq分析表明，不到1%的基序被先锋因子GATA 3占据。除了 结合选择性，先锋因子作用，包括GATA 3，已知是位点特异性的（依赖于环境）， 并且它们只能在结合位点的子集处诱导染色质开放和增强子形成。整体 该提案的目的是鉴定基因组中先锋因子GATA 3的结合特异性， 阐明GATA 3与染色质修饰酶聚ADP-核糖之间相互作用的影响 聚合酶1（PARP 1）对基因激活的影响。我们的初步数据强烈表明PARP 1参与了 GATA 3这种依赖于上下文的作用。核心假设是GATA 3介导的细胞凋亡可能是一个重要的机制。 重编程依赖于PARP 1活性。这项研究的基本原理是， GATA 3和PARP 1相互作用的作用将提供新的分子机制， 在细胞重编程期间先锋因子活性的特异性表达。核心假设是 通过（1）研究PARP 1在GATA 3的结合选择性中的功能和（2）鉴定 GATA 3-PARP 1相互作用在基因激活中的作用。为了验证这个假设，我将使用我们独特的DOX- MDA-MB-231基底乳腺癌细胞中的诱导系统。在该系统中，外源GATA 3表达 启动间充质至上皮转变（MET）。该系统概括了患者中的GATA 3功能 乳腺肿瘤，因为已知GATA 3表达在管腔/上皮乳腺肿瘤中更高， 对维持上皮表型至关重要。在第一个目标下，我们将调查 在存在和不存在PARP 1的情况下跨基因组的GATA 3结合。第二个目标将利用下一个- 代测序，以评估GATA 3和PARP 1对先锋因子- 介导的细胞重编程。这项研究是创新性的，因为我们正在解决小说 在细胞重编程过程中由先锋因子形成的背景依赖性增强子的概念。 这项研究意义重大，因为它将填补我们关于先驱因素如何介导的知识的关键空白 细胞命运转变