Structural and Functional Studies of lncRNAs in Gene Activation

lncRNA 在基因激活中的结构和功能研究

基本信息

批准号：
10637407
负责人：
Srinivas Somarowthu
金额：
$ 38.83万
依托单位：
DREXEL UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-04-01 至 2028-02-29
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10637407
关键词：
3-Dimensional Adopted Affinity Binding Sites Biochemical Biological Assay Biological Models CRISPR/Cas technology Cell Proliferation Cell physiology Cells Chemicals Chromatin Complex Cryoelectron Microscopy DNA Development Disease Disease Progression Down-Regulation Elements Epigenetic Process Family Gene Activation Gene Expression Regulation Gene Silencing Genes Genetic Transcription Goals Histones Human Immunoprecipitation Investigation Knowledge Literature Malignant Neoplasms Mammals Maps Micrococcal Nuclease Molecular Mutate Nucleosomes Nucleotides Plants Play Polycomb Process Promoter Regions Proteins RNA Reporting Research Resolution Roentgen Rays Role Site Spectrometry Structure Structure-Activity Relationship Sucrose Technology Testing Transcript Transcription Coactivator Tumor Promotion Tumor Suppressor Proteins Untranslated RNA Visualization WNT Signaling Pathway cell motility chromatin remodeling crosslink deletion analysis experimental study genetic regulatory protein insight knock-down mammalian genome model building novel particle potential biomarker promoter recruit scaffold therapeutic target three dimensional structure transcription factor tumor progression

项目摘要

Project Summary Over the last two decades, breakthroughs in sequencing technologies have revealed that mammalian genomes encode thousands of long non-coding RNAs (lncRNAs). The numbers of human lncRNAs that are known to be dysregulated in diseases are rising at a rapid pace. Emerging evidence suggests that lncRNAs are critical regulators of fundamental cellular processes and disease progression. Despite their significance as potential biomarkers and therapeutic targets, the molecular mechanisms of lncRNAs remain poorly understood. The primary goal of this research is to elucidate in greater detail the mechanisms by which lncRNAs activate transcription. Many lncRNAs are associated with epigenetic machinery; these include histone-modifying complexes, such as the polycomb repressive complex 2, and nucleosome remodeling complexes, such as the SWI/SNF (SWItch/Sucrose Non-Fermentable) complex. While the RNA-PRC2 interactions are under active investigation, the role of lncRNAs in nucleosome remodeling remains uncharacterized. Here, we seek to understand the function of lncRNAs in SWI/SNF-based gene activation. The SWI/SNF complex is a known tumor suppressor, and its subunits are mutated in various cancers. Recently, lncRNAs have been shown to interact with the SWI/SNF complex in mammals and plants. In this proposal, we will utilize lncTCF7 to investigate the role of lncRNAs in SWI/SNF-based gene activation. Previous studies have shown that lncTCF7 activates Wnt signaling by recruiting the SWI/SNF complex. However, the biochemical and structural basis of these interactions is not established. In preliminary studies, we identified SND1 (Staphylococcal Nuclease and Tudor Domain Containing 1) as the top interaction partner of lncTCF7. SND1 is a transcriptional coactivator, and it is known to associate with the SWI/SNF family. Here, we propose a novel mechanism suggesting that lncTCF7 recruits the SWI/SNF complex indirectly via its interaction with SND1. Further, our RNA chemical probing studies identified structural domains and regions of lncTCF7 conserved across mammalian genomes. We hypothesize that these conserved regions and structured elements play critical roles in recruiting protein partners and activating transcription at specific loci. We will test this hypothesis through the following aims: In Aim 1, we will define the functional role of lncTCF7’s domains. Aim 2 will determine the role of lncTCF7-SND1 interaction in recruiting the SWI/SNF complex and activating transcription. In Aim 3, we will determine the 3D structure of lncTCF7 and the lncTCF7-SND1 complex. Together, these studies will provide biochemical and structural knowledge of lncTCF7’s molecular function and expand our understanding of how lncRNAs regulate gene transcription and contribute to disease states.

项目概要在过去的二十年里，测序技术的突破表明，哺乳动物基因组编码数千种长非编码 RNA (lncRNA) 已知的人类 lncRNA 数量。疾病失调现象正在迅速增加，新的证据表明 lncRNA 至关重要。尽管它们具有潜在的重要性，但它们是基本细胞过程和疾病进展的调节因子。由于生物标志物和治疗靶点，lncRNA 的分子机制仍知之甚少。这项研究的主要目标是更详细地阐明 lncRNA 激活的机制许多 lncRNA 与表观遗传机制相关；其中包括组蛋白修饰。复合物，例如多梳抑制复合物 2，和核小体重塑复合物，例如 SWI/SNF（SWIitch/蔗糖不可发酵）复合物，而 RNA-PRC2 相互作用处于活跃状态。调查显示，lncRNA 在核小体重塑中的作用仍未得到表征。了解 lncRNA 在基于 SWI/SNF 的基因激活中的功能 SWI/SNF 复合体是一种已知的肿瘤。最近，lncRNA 已被证明可以相互作用。在本提案中，我们将利用 lncTCF7 来研究哺乳动物和植物中的 SWI/SNF 复合物。 lncRNA在基于SWI/SNF的基因激活中的作用已经表明lncTCF7激活Wnt。然而，这些相互作用的生化和结构基础。在初步研究中，我们鉴定了 SND1（葡萄球菌核酸酶和 Tudor 结构域）。包含 1) 作为 lncTCF7 的顶级相互作用伙伴，SND1 是一种转录共激活因子，已知在此，我们提出了一种新机制，表明 lncTCF7 招募了 SWI/SNF 家族。 SWI/SNF 复合物通过其与 SND1 的相互作用间接实现。此外，我们的 RNA 化学探测研究发现。我们发现，lncTCF7 的结构域和区域在哺乳动物基因组中是保守的。保守区域和结构元件在招募蛋白质伙伴和激活中发挥着关键作用我们将通过以下目标来检验这一假设：在目标 1 中，我们将定义 lncTCF7 结构域的功能作用将决定 lncTCF7-SND1 相互作用在招募中的作用。 SWI/SNF 复合物和激活转录在目标 3 中，我们将确定 lncTCF7 的 3D 结构和 lncTCF7-SND1 复合物，这些研究将提供 lncTCF7 的生化和结构知识。分子功能并扩展我们对 lncRNA 如何调节基因转录并有助于疾病状态。