High resolution characterization of epigenetic dynamics in gastrointestinal cell plasticity

胃肠道细胞可塑性表观遗传动力学的高分辨率表征

基本信息

批准号：
10576232
负责人：
Unmesh Jadhav
金额：
$ 12.38万
依托单位：
UNIVERSITY OF SOUTHERN CALIFORNIA
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：
2023-02-01 至 2025-01-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10576232
关键词：
Ablation Abnormal Cell Address Binding Biological Assay Biological Process Boranes Cancer Etiology Cell Nucleus Cells Cellular Assay Cessation of life Charge Chemicals Chromatin Chromatin Structure Cluster Analysis Colorectal Colorectal Cancer Complementary DNA Computing Methodologies DNA DNA Damage DNA Methylation DNA Sequence Alteration DNA amplification DNA analysis Data Development Developmental Gene Disease Disease Progression Enhancers Epigenetic Process Epithelial Cells Epithelium Event Evolution Future Gene Expression Gene Expression Profile Gene Mutation Genes Genetic Genetic Transcription Genome Genomic approach Intestines Island KRAS2 gene Knowledge Malignant Neoplasms Maps Measures Mesenchymal Messenger RNA Methods Methylation Modality Modification Molecular Mutation Natural regeneration Process Proteins Protocols documentation RNA Regulator Genes Research Resolution Signal Transduction Sorting Stains TP53 gene Technology Testing Therapeutic Intervention Time Tissues base bisulfite cell behavior cell type disease prognosis drug development early onset colorectal cancer epigenomics gastrointestinal histone modification innovation intestinal epithelium magnetic beads multiple omics new therapeutic target novel preservation promoter stem cells therapy design tissue regeneration tissue repair transcription factor transcriptomic profiling tumor tumorigenesis

项目摘要

Project Summary Continuous differentiation of intestinal stem cells (ISCs) maintains the integrity of intestinal epithelium while producing diverse functional cells at rapid pace. This epithelium also displays remarkable cell plasticity, including the ability of mature cells to dedifferentiate into stem cells upon loss of ISCs. As many genetic and signaling drivers of this cell fate reversion are being discovered, cell intrinsic epigenetic controls that must be rewired for cell plasticity, remain largely unknown. Using state-of-the-art single-cell sc-Multiome assays that reveal chromatin accessibility (scATAC-seq) and gene expression (scRNA-seq) in parallel, we have recently produced high-quality data comparing native epithelial cells and cells undergoing regeneration upon ISC ablation. Using these data and innovative computational methods, we will chart the chromatin and gene expression dynamic underlying cell fate changes in intestinal regeneration, and identify key epigenetic modulators and transcription factor controllers of this process. In conjunction with these analyses, we propose to delineate promoter and enhancer relevant epigenetic controls (histone modifications) through the time course of regeneration. Aberrant chromatin structure can cause gene dysregulation and contribute to tumorigenesis, including colorectal cancer. DNA methylation is the most frequently modulated epigenetic modality in colorectal as well as many other cancers. However, determinants of the reciprocal interactions between DNA methylation and mutational changes or other epigenetic layers have not been studied critically. Also missing is the knowledge of how early in tumorigenesis does the DNA methylation change occur and how uniform are the changes in DNA methylation across cells. Technological limitations have been the most significant barrier for such analyses, as current methods to characterize DNA methylation at single-cell level suffer from low CpG coverage limited to gene promoters and GpG islands, which precludes understanding of the true breadth of DNA methylation change, particularly at gene regulatory enhancers. We propose to override these limitations through development of a novel assay for characterizing DNA methylation as well as RNA expression at single-cell level. We combine chemical methods geared to preserve DNA integrity and gentler nuclei handling based on magnetic bead capture in our method called scTaMT-seq, which will provide a robust protocol for analyzing DNA methylation in unprecedented detail at highest possible resolution. This technology will propel the understanding of epigenomic impact on various biological processes, particularly cancer.

项目概要肠干细胞（ISC）的持续分化维持肠上皮的完整性，同时快速产生多种功能细胞。这种上皮还表现出显着的细胞可塑性，包括成熟细胞在 ISC 丢失后去分化为干细胞的能力。由于许多遗传和这种细胞命运逆转的信号驱动因素正在被发现，细胞内在的表观遗传控制必须是细胞可塑性的重新连接，仍然很大程度上未知。使用最先进的单细胞 sc-Multiome 检测并行揭示染色质可及性 (scATAC-seq) 和基因表达 (scRNA-seq)，我们最近产生了比较天然上皮细胞和 ISC 再生细胞的高质量数据消融。利用这些数据和创新的计算方法，我们将绘制染色质和基因的图表肠道再生中潜在细胞命运变化的表达动态变化，并确定关键的表观遗传该过程的调节剂和转录因子控制器。结合这些分析，我们建议描述随着时间的推移启动子和增强子相关的表观遗传控制（组蛋白修饰）再生过程。异常的染色质结构可导致基因失调并导致肿瘤发生，包括结直肠癌。 DNA 甲基化也是结直肠中最常调节的表观遗传模式和许多其他癌症一样。然而，DNA 甲基化和 DNA 甲基化之间相互作用的决定因素突变变化或其他表观遗传层尚未得到严格研究。还缺少知识 DNA 甲基化变化在肿瘤发生的早期发生以及变化的均匀性如何跨细胞的 DNA 甲基化。技术限制是此类活动的最大障碍分析，因为目前在单细胞水平上表征 DNA 甲基化的方法存在 CpG 较低的问题覆盖范围仅限于基因启动子和 GpG 岛，这妨碍了对基因启动子和 GpG 岛的真正广度的理解 DNA 甲基化变化，尤其是基因调控增强子的甲基化变化。我们建议克服这些限制通过开发一种新的测定方法来表征 DNA 甲基化以及 RNA 表达单细胞水平。我们结合了旨在保持 DNA 完整性的化学方法和更温和的细胞核处理基于我们称为 scTaMT-seq 的方法中的磁珠捕获，该方法将为以尽可能高的分辨率以前所未有的细节分析 DNA 甲基化。这项技术将推动了解表观基因组对各种生物过程，特别是癌症的影响。