Predicting Lung Chromatin Access Profiling in an Animal Model

项目总结/摘要 对核小体内特定组蛋白和DNA序列的化学修饰吸引染色质 重塑复合体重塑复合体创造了开放和限制染色质进入的区域， 控制基因表达。环境因素破坏将表观遗传标记分配给组蛋白和/或DNA， 改变染色质通路，改变基因表达，增加疾病风险。 本文综述了转座酶可及染色质检测技术的最新进展， ATAC测序（ATAC-seq）使得通过核苷酸分辨率鉴定染色质通路成为可能。 ~5000个细胞。ATAC-seq确定开放染色质、DNA结合蛋白、个体 核小体和染色质致密化。定义染色质通路提供了关于功能性 早产和机械通气（PTMV）可能会改变DNA调控区的状态。我们 将使用ATAC-seq来确定PTMV对我们独特的早产羔羊模型中染色质重塑的影响 支气管肺发育不良（BPD）。我们修订后的R21的新数据表明：1）染色质景观模式 PTMV羔羊与健康对照羔羊在肺和脑中存在差异，2）这些差异是 反映在来自PTMV羔羊与健康对照羔羊的CD 4 + T细胞内，和3）管家基因是 在羔羊CD 4 + T细胞、肺、脑和人B细胞中是保守的。我们假设PTMV对 影响指导染色质重塑和建立适当的 整个基因组的染色质通路。我们将通过以下两个具体目标来测试我们的假设。 具体目标1。确定PTMV诱导的染色质景观的改变。组织特异 将通过比较肺和脑之间的结果来识别染色质景观的差异。 具体目标2。比较CD 4 + T细胞中染色质景观的改变与 在目标1中获得的肺和脑。将确定染色质景观的组织特异性差异 通过比较同一羔羊的肺和脑中的CD 4 + T细胞的结果。 我们修改后的R21应用程序的结果将是重要的，因为它们将识别CD 4 + T细胞， 肺和脑中染色质景观表观遗传变化的可获得生物标志物， 在BPD进展的早产儿中受到不利影响，但不容易进行分子分析。 创新方法将联合收割机ATAC-seq与我们的早产羔羊BPD模型相结合。我们的合作，多重- PI成员带来了表观遗传学、BPD早产羔羊模型和白细胞生物学方面的专业知识。平移 在CD 4+淋巴细胞中定义与改变的基因相关的表观遗传生物标志物， 在肺和脑组织中表达。我们的R21项目的影响将是一种可能适用的新方法 对于有BPD风险的人类早产儿，部分原因是ATAC-seq分析现在可用于单细胞。

PROJECT SUMMARY/ABSTRACT Chemical modifications to specific histones and DNA sequences within nucleosomes attract chromatin remodeling complexes. Remodeling complexes create regions of open and restricted chromatin access to control gene expression. Environmental factors disrupt assigning epigenetic marks to histones and/or DNA that alter chromatin access, altering gene expression and increasing disease risk. The recent development of the cutting-edge technology assay for transposase-accessible chromatin, using ATAC sequencing (ATAC-seq), makes identification possible of chromatin access with nucleotide resolution on ~5000 cells. ATAC-seq determines the genomic location of open chromatin, DNA binding proteins, individual nucleosomes, and chromatin compaction. Defining chromatin access provides information about the functional state of DNA regulatory regions that may be altered by preterm birth and mechanical ventilation (PTMV). We will use ATAC-seq to identify the impact of PTMV on chromatin remodeling in our unique preterm lamb model of bronchopulmonary dysplasia (BPD). Our revised R21’s new data show that 1) chromatin landscape patterns are different in lung and brain from PTMV lambs versus healthy control lambs, 2) these differences are reflected within CD4+ T cells from PTMV lambs versus healthy control lambs, and 3) housekeeping genes are conserved among lamb CD4+ T cells, lung, brain, and human B cells. We hypothesize that PTMV negatively impact assignment of epigenetic marks necessary to direct chromatin remodeling and establish proper chromatin access throughout the genome. We will test our hypothesis by doing the following 2 specific aims. Specific Aim 1. Determine alterations to chromatin landscape induced by PTMV. Tissue-specific differences in chromatin landscapes will be identified by comparing results between lung and brain. Specific Aim 2. Compare alterations in chromatin landscape in CD4+ T cells with those identified in the lung and brain obtained in Aim 1. Tissue-specific differences in chromatin landscapes will be identified by comparing results between CD4+ T cells and lung and brain from the same lamb. Results of our revised R21 application will be significant because they will identify CD4+ T cells as accessible biomarkers of epigenetic changes in chromatin landscape in the lung and brain, which become adversely affected in preterm infants with evolving BPD but are not readily accessible for molecular analysis. Innovative approaches combine ATAC-seq with our preterm lamb model of BPD. Our collaborative, multiple- PI members bring expertise in epigenetics, preterm lamb model of BPD, and leukocyte biology. Translational potential arises from defining epigenetic biomarkers in CD4+ lymphocytes that correlate with altered gene expression in lung and brain tissue. Impact of our R21 project will be a novel approach that may be applicable to human preterm infants at risk for BPD, in part because ATAC-seq analysis now is available for single cells.