Chromatin Remodeling of the Epidermal Differentiation Complex

表皮分化复合体的染色质重塑

• 批准号: 8887751
• 负责人: Cristina de Guzman Strong
• 金额: $ 33.55万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-03-01 至 2020-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8887751
• 关键词: 1q21; ATAC-seq; Architecture; Binding; Binding Sites; Biology; Cell model; ChIP-seq; Chromatin; Chromatin Remodeling Factor; Chromatin Structure; Clustered Regularly Interspaced Short Palindromic Repeats; Complex; Coupled; Data; Defect; Development; Disease; Elements; Embryo; Enhancers; Epidermis; Epigenetic Process; Future; Gene Activation; Gene Expression; Gene Expression Profile; Gene Expression Regulation; Gene Family; Genes; Genetic Transcription; Genome; Genome engineering; Genomic Segment; Genomic approach; Genomics; Goals; Histocompatibility Testing; Human; Hypersensitivity; In Vitro; Inflammatory; Insulator Elements; Investigation; JUN gene; Knowledge; Link; Mediating; Medicine; Methods; Molecular; Molecular Conformation; Morphogenesis; Mus; Organism; Pathogenicity; Pattern; Phenotype; Proteins; Publishing; Regulatory Element; Research; Resolution; Risk; Role; Skin; Surface; Technology; Testing; Time; Tissues; Transcription Factor AP-1; Transcriptional Activation; Transgenic Mice; base; chromatin remodeling; empowered; filaggrin; functional genomics; genetic approach; histone modification; human tissue; innovation; insight; keratinocyte; keratinocyte differentiation; mouse development; mouse model; next generation sequencing; novel; novel strategies; promoter; public health relevance; skin disorder; tool; transcription factor; transcriptome sequencing

 DESCRIPTION (provided by applicant): The epidermis at the surface of the skin provides the first line of defense for an organism. The expression of the Epidermal Differentiation Complex (EDC) genes is critical to this epidermal barrier function. The EDC encodes 4 gene families (FLG-like, LCE, SPRR, and S100) that are coordinately expressed and provide the essential cellular components that define a functional epidermal keratinocyte. It has been 20 years since the discovery of the EDC and the molecular mechanisms that govern concomitant activation of the EDC in the keratinocyte continues to elude the field. Using an unprecedented and comprehensive genomics approach to skin biology, we directly revisit this important question. Enhancers are important regulatory elements in the genome that are noncoding. Enhancers activate gene expression by proximal looping to the respective gene promoter that is mediated by transcription factor binding. Our recent studies identified a highly conserved enhancer within the EDC, namely 923, whose regulatory activity coincided with and is specific to the spatial and temporal patterning of epidermal differentiation and EDC transcription across the developing mouse epidermis. Using innovative chromosomal conformation capture methods, we further identified dynamic chromatin remodeling of the EDC with respect to the 923 enhancer during keratinocyte differentiation. A role for 923 enhancer activity for EDC activation was further supported by aberrant EDC chromatin remodeling and repressed EDC gene activation upon the loss of AP-1 transcription factor binding to 923. This led us to propose the following hypothesis. The 923 enhancer is a critical regulatory enhancer that imposes a specific configuration of the chromatin architecture for coordinate transcriptional activation of the EDC for epidermal differentiation. We propose the following aims: Aim 1. Identify the conserved epigenetic mechanisms of coordinate EDC gene expression in mice and humans. Here we will test the hypothesis that 923 is released from CTCF-bound insulators thereby allowing accessible AP-1 binding and for 923 to enhance activated EDC gene expression. We will use genomics approaches (4C-seq) to define the chromatin interactions with respect to 923, filaggrin, and a known CTCF binding site and identify the epigenetic mechanisms that govern these interactions (parallel ChIP-seq, RNA-seq, and ATAC-seq studies). Aim 2. Determine the loss of the 923 enhancer in epidermal development. We have successfully generated 5 independent lines of 923 deleted (3) and floxed mice (2) using the state-of-the-art CRISPR/Cas9-mediated genome engineering method. Molecular and cellular studies will be performed to determine the mouse phenotypes. Accomplishments of these aims will have a significant impact on our understanding of the molecular underpinnings linking chromatin architecture to gene expression at an unprecedented resolution in skin biology and relevant to all human tissue types. This study will also provide the necessary tools to develop novel strategies to correct aberrant epidermal differentiation with targeted epigenetic and chromatin remodeling therapies in medicine.

 描述（由适用提供）：皮肤表面的表皮为组织提供了第一道防线。表皮分化复合物（EDC）基因的表达对于这种表皮屏障功能至关重要。 EDC编码4个基因家族（FLG样，LCE，SPRR和S100），它们是协调表达的，并提供了定义功能表皮角质形成细胞的必需细胞成分。自从发现EDC和控制角质形成细胞中EDC激活的分子机制以来，我们已经20年了。使用前所未有且全面的基因组学方法，我们直接重新审视了这个重要问题。增强子是基因组中不编码的重要调节元素。增强子通过代理循环循环到各个基因启动子，该基因启动子由转录因子结合介导。我们最近的研究确定了EDC中高度组成的增强子，即923，其调节活性与表皮分化和EDC转录的空间和临时模式相吻合，并且特定于整个发育中的小鼠表皮上。使用创新的染色体会议捕获方法，我们进一步确定了在角质形成细胞分化过程中相对于923增强剂的EDC的动态染色质重塑。异常的EDC染色质重塑和抑制EDC基因激活，在AP-1转录因子与923结合到923的情况下，进一步支持了923增强子活性的EDC激活的作用。这使我们提出了以下假设。 923增强剂是一个关键的调节增强剂，即无法对染色质结构的特定构型用于坐标EDC的转录激活以进行表皮分化。我们提出以下目的：目标1。确定小鼠和人类坐标EDC基因表达的保守表观遗传机制。在这里，我们将检验以下假设：923是从CTCF结合的绝缘子释放的，从而允许可访问的AP-1结合，并使923增强活化的EDC基因表达。我们将使用基因组学方法（4C-SEQ）来定义相对于923，Filaggrin和已知的CTCF结合位点的染色质相互作用，并确定控制这些相互作用的表观遗传机制（并行芯片seq，rna-seq和atac-seq研究）。目标2。确定表皮发育中923增强子的损失。我们使用最先进的CRISPR/CAS9介导的基因组工程方法成功地生成了5条923条删除（3）和Floxed小鼠（2）的独立线。将进行分子和细胞研究以确定小鼠表型。这些目标的成就将对我们对将染色质结构与基因表达联系起来的分子基础的理解具有重大影响，这在皮肤生物学方面具有前所未有的分辨率，并且与所有人类组织类型有关。这项研究还将提供必要的工具来制定新型策略，以纠正医学中有针对性的表观遗传和染色质重塑疗法的异常表皮分化。