Regulatory Elements, Transcriptional

监管要素 ,转录
  • 文章类型: Journal Article
    顺式调节元件(CREs)在协调整个生物系统中的基因表达方面至关重要。功能CRE的准确识别和深入表征对于在细胞过程中解码基因调控网络至关重要。在这项研究中,我们开发了用于转座酶可访问染色质的Kethoxal辅助单链DNA分析测序(KAS-ATAC-seq),以定量分析CREs的转录活性。KAS-ATAC-seq的主要优势在于其在近端和远端ATAC-seq峰内的ssDNA水平的精确测量,能够识别转录调控序列。该特征特别擅长定义单链转录增强子(SSTE)。SSTE高度富含新生RNA和定义细胞同一性的特异性转录因子(TF)结合位点。此外,KAS-ATAC-seq提供了各种SSTE亚型的详细表征和功能含义。我们对小鼠神经分化过程中的CRE的分析表明,KAS-ATAC-seq可以有效地识别响应于视黄酸(RA)治疗的立即早期激活的CRE。我们的发现表明KAS-ATAC-seq提供了转录中功能CRE的更精确注释。KAS-ATAC-seq的未来应用将有助于阐明不同生物过程中基因调控的复杂动力学。
    Cis-regulatory elements (CREs) are pivotal in orchestrating gene expression throughout diverse biological systems. Accurate identification and in-depth characterization of functional CREs are crucial for decoding gene regulation networks during cellular processes. In this study, we develop Kethoxal-Assisted Single-stranded DNA Assay for Transposase-Accessible Chromatin with Sequencing (KAS-ATAC-seq) to quantitatively analyze the transcriptional activity of CREs. A main advantage of KAS-ATAC-seq lies in its precise measurement of ssDNA levels within both proximal and distal ATAC-seq peaks, enabling the identification of transcriptional regulatory sequences. This feature is particularly adept at defining Single-Stranded Transcribing Enhancers (SSTEs). SSTEs are highly enriched with nascent RNAs and specific transcription factors (TFs) binding sites that define cellular identity. Moreover, KAS-ATAC-seq provides a detailed characterization and functional implications of various SSTE subtypes. Our analysis of CREs during mouse neural differentiation demonstrates that KAS-ATAC-seq can effectively identify immediate-early activated CREs in response to retinoic acid (RA) treatment. Our findings indicate that KAS-ATAC-seq provides more precise annotation of functional CREs in transcription. Future applications of KAS-ATAC-seq would help elucidate the intricate dynamics of gene regulation in diverse biological processes.
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  • 文章类型: Journal Article
    转座元件(TE)和其他重复区已被证明含有基因调控元件,包括转录因子结合位点。然而,已证明由重复所携带的调节元件难以使用短读取测序测定(例如ChIP-seq或ATAC-seq)来表征。大多数调控基因组学分析流水线丢弃与多个基因组位置一致的“多重映射”读取。因为多联读取主要来自重复,当前的分析管道无法检测到重复区域中发生的大部分监管事件。为了解决这个缺点,我们开发了Allo,一种以高效的方式分配多映射读取的新方法,准确,和用户友好的方式。Allo将多映射读段的概率映射与识别潜在峰的读段分布特征的卷积神经网络相结合,在多任务读取分配中提供增强的准确性。Allo还以校正的对齐文件的形式提供读取级别输出,使其与现有的调控基因组学分析管道和下游测峰仪兼容。在CTCFChIP-seq数据的演示应用程序中,我们表明,Allo导致发现了数千个新的CTCF峰。这些峰中的许多含有预期的同源基序和/或用作TAD边界。我们还将Allo应用于不同的ENCODEChIP-seq数据集集合,导致转录因子和重复元件家族之间的多个先前未识别的相互作用。最后,我们表明,Allo可能特别有利于识别着丝粒的ChIP-seq峰,接近分段重复的基因,在年轻的TEs中,在这些地区进行新的监管分析。
    Transposable elements (TEs) and other repetitive regions have been shown to contain gene regulatory elements, including transcription factor binding sites. However, regulatory elements harbored by repeats have proven difficult to characterize using short-read sequencing assays such as ChIP-seq or ATAC-seq. Most regulatory genomics analysis pipelines discard \"multimapped\" reads that align equally well to multiple genomic locations. Because multimapped reads arise predominantly from repeats, current analysis pipelines fail to detect a substantial portion of regulatory events that occur in repetitive regions. To address this shortcoming, we developed Allo, a new approach to allocate multimapped reads in an efficient, accurate, and user-friendly manner. Allo combines probabilistic mapping of multimapped reads with a convolutional neural network that recognizes the read distribution features of potential peaks, offering enhanced accuracy in multimapping read assignment. Allo also provides read-level output in the form of a corrected alignment file, making it compatible with existing regulatory genomics analysis pipelines and downstream peak-finders. In a demonstration application on CTCF ChIP-seq data, we show that Allo results in the discovery of thousands of new CTCF peaks. Many of these peaks contain the expected cognate motif and/or serve as TAD boundaries. We additionally apply Allo to a diverse collection of ENCODE ChIP-seq data sets, resulting in multiple previously unidentified interactions between transcription factors and repetitive element families. Finally, we show that Allo may be particularly beneficial in identifying ChIP-seq peaks at centromeres, near segmentally duplicated genes, and in younger TEs, enabling new regulatory analyses in these regions.
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  • 文章类型: Journal Article
    基因调控元件中的核苷酸变化是神经元发育和疾病的重要决定因素。在来自妊娠中期皮质和大脑类器官的原代人细胞中使用大规模平行报告子测定法,我们询问了102,767个开放染色质区域的顺式调节活性,包括数千个具有细胞类型特异性可及性的序列和与脑基因调控相关的变异。在原代细胞中,我们鉴定了46,802个活性增强子序列和164个改变增强子活性的变体。类器官和原代细胞的活性相当,这表明类器官为发育中的皮质提供了一个适当的模型。使用深度学习,我们解码了增强子活性的序列基础和上游调节因子。这项工作建立了人类神经元发育中功能基因调控元件和变体的综合目录。
    Nucleotide changes in gene regulatory elements are important determinants of neuronal development and diseases. Using massively parallel reporter assays in primary human cells from mid-gestation cortex and cerebral organoids, we interrogated the cis-regulatory activity of 102,767 open chromatin regions, including thousands of sequences with cell type-specific accessibility and variants associated with brain gene regulation. In primary cells, we identified 46,802 active enhancer sequences and 164 variants that alter enhancer activity. Activity was comparable in organoids and primary cells, suggesting that organoids provide an adequate model for the developing cortex. Using deep learning we decoded the sequence basis and upstream regulators of enhancer activity. This work establishes a comprehensive catalog of functional gene regulatory elements and variants in human neuronal development.
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  • 文章类型: Journal Article
    人脑中细胞类型特异性基因调控元件中的核苷酸变体是人类疾病的危险因素。我们测量了来自616个人类死后大脑的分选神经元和非神经元的1932等分试样的染色质可及性,并确定了34,539个开放染色质区域,具有染色质可及性定量性状基因座(caQTL)。只有10.4%的caQTL在神经元和非神经元之间共享,它支持大脑调节组的细胞类型特异性遗传调控。结合等位基因特异性染色质可及性可改善统计精细作图并完善疾病风险基础的分子机制。在诱导的兴奋性神经元中使用大规模平行报告子测定法,我们筛选了19,893个脑QTL,并鉴定了476个调控变体的功能影响.合并,这一综合资源捕获了人脑调节组的变化,并提供了对疾病病因的见解。
    Nucleotide variants in cell type-specific gene regulatory elements in the human brain are risk factors for human disease. We measured chromatin accessibility in 1932 aliquots of sorted neurons and non-neurons from 616 human postmortem brains and identified 34,539 open chromatin regions with chromatin accessibility quantitative trait loci (caQTLs). Only 10.4% of caQTLs are shared between neurons and non-neurons, which supports cell type-specific genetic regulation of the brain regulome. Incorporating allele-specific chromatin accessibility improves statistical fine-mapping and refines molecular mechanisms that underlie disease risk. Using massively parallel reporter assays in induced excitatory neurons, we screened 19,893 brain QTLs and identified the functional impact of 476 regulatory variants. Combined, this comprehensive resource captures variation in the human brain regulome and provides insights into disease etiology.
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  • 文章类型: Journal Article
    与精神疾病相关的大多数遗传变异位于基因组的非编码区。为了调查它们的功能含义,我们整合了来自PsychENCODE联盟和其他已发表资料的表观遗传学数据,构建了一个综合的候选脑顺式调节元件图谱.使用深度学习,我们对这些元件的序列语法进行建模,并预测谱系特异性转录因子的结合位点如何在各种类型的神经胶质和神经元中促进细胞类型特异性基因调控。元素进化史表明,大脑中的新调控信息主要是通过保守哺乳动物元素中的较小序列突变而出现的,而不是全新的人类或灵长类动物特异性序列。然而,灵长类动物特异性候选元素,特别是那些在胎儿大脑发育和兴奋性神经元和星形胶质细胞中活跃的人,与大脑相关的人类特征的遗传力有关。此外,我们介绍心理屏幕,一个基于网络的平台,提供来自精神疾病患者和健康对照者不同脑细胞类型的PsychENCODE生成的遗传和表观遗传数据的交互式可视化。
    Most genetic variants associated with psychiatric disorders are located in noncoding regions of the genome. To investigate their functional implications, we integrate epigenetic data from the PsychENCODE Consortium and other published sources to construct a comprehensive atlas of candidate brain cis-regulatory elements. Using deep learning, we model these elements\' sequence syntax and predict how binding sites for lineage-specific transcription factors contribute to cell type-specific gene regulation in various types of glia and neurons. The elements\' evolutionary history suggests that new regulatory information in the brain emerges primarily via smaller sequence mutations within conserved mammalian elements rather than entirely new human- or primate-specific sequences. However, primate-specific candidate elements, particularly those active during fetal brain development and in excitatory neurons and astrocytes, are implicated in the heritability of brain-related human traits. Additionally, we introduce PsychSCREEN, a web-based platform offering interactive visualization of PsychENCODE-generated genetic and epigenetic data from diverse brain cell types in individuals with psychiatric disorders and healthy controls.
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  • 文章类型: Journal Article
    ChIP-Atlas(https://chip-atlas.org/)提供了一套用于分析表观基因组景观的数据挖掘工具,由超过376.000个公共ChIP-seq的全面集成提供动力,ATAC-seq,来自六种代表性模型生物的DNase-seq和亚硫酸氢盐-seq实验。为了解开染色质结构的复杂性,它介导了细胞内转录和表型多样性的调节组引发的产生,我们报告了ChIP-Atlas3.0通过在新合并的“注释轨道”部分中纳入基因组和表观基因组特征的其他轨道,从而增强了清晰度。轨迹包括染色体构象(Hi-C和eQTL数据集),转录调控元件(ChromHMM和FANTOM5增强子),以及与疾病和表型相关的基因组变异(GWASSNP和ClinVar变异)。这些注释轨道与其他实验轨道一起很容易访问,促进更好地阐明转录和表型性状多样化的染色质结构。此外,\'差异分析,\'一个新的在线工具,比较查询表观基因组数据以识别差异结合,可访问,和使用ChIP-seq的甲基化区域,ATAC-seq和DNase-seq,和亚硫酸氢盐-SEQ数据集,分别。注释轨道和差异分析工具的集成,加上持续的数据扩展,使ChIP-Atlas3.0成为挖掘转录调控机制景观的强大资源,从而提供有价值的观点,特别是遗传疾病研究和药物发现。
    ChIP-Atlas (https://chip-atlas.org/) presents a suite of data-mining tools for analyzing epigenomic landscapes, powered by the comprehensive integration of over 376 000 public ChIP-seq, ATAC-seq, DNase-seq and Bisulfite-seq experiments from six representative model organisms. To unravel the intricacies of chromatin architecture that mediates the regulome-initiated generation of transcriptional and phenotypic diversity within cells, we report ChIP-Atlas 3.0 that enhances clarity by incorporating additional tracks for genomic and epigenomic features within a newly consolidated \'annotation track\' section. The tracks include chromosomal conformation (Hi-C and eQTL datasets), transcriptional regulatory elements (ChromHMM and FANTOM5 enhancers), and genomic variants associated with diseases and phenotypes (GWAS SNPs and ClinVar variants). These annotation tracks are easily accessible alongside other experimental tracks, facilitating better elucidation of chromatin architecture underlying the diversification of transcriptional and phenotypic traits. Furthermore, \'Diff Analysis,\' a new online tool, compares the query epigenome data to identify differentially bound, accessible, and methylated regions using ChIP-seq, ATAC-seq and DNase-seq, and Bisulfite-seq datasets, respectively. The integration of annotation tracks and the Diff Analysis tool, coupled with continuous data expansion, renders ChIP-Atlas 3.0 a robust resource for mining the landscape of transcriptional regulatory mechanisms, thereby offering valuable perspectives, particularly for genetic disease research and drug discovery.
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  • 文章类型: Journal Article
    无法在多细胞系统中可缩放和精确地测量发育顺式调节元件(CREs)的活性是基因组学的瓶颈。在这里,我们开发了一种双RNA盒,它将多重单细胞报告分子测定固有的检测和定量任务解耦。报告表达的结果测量在多个数量级上是准确的,精度接近泊松计数噪声设定的极限。与通过环化的RNA条形码稳定一起,这些可扩展的单细胞定量表达报告基因提供了高对比度的读数,类似于经典的原位测定,但完全来自测序。在早期哺乳动物发育的多细胞体外模型中筛选>200个可接近染色质区域,我们确定了13个(8个以前未表征的)自主和细胞类型特异性发育CRE。我们进一步证明了嵌合CRE对产生同源的两细胞型活性谱,并评估了来自具有扰动的转录因子结合位点的CRE变体的功能增益和功能丧失的多细胞表达表型。单细胞定量表达报告基因可应用于发育和多细胞系统,以定量表征天然,在规模上扰动和合成的CRE,具有高灵敏度和单细胞分辨率。
    The inability to scalably and precisely measure the activity of developmental cis-regulatory elements (CREs) in multicellular systems is a bottleneck in genomics. Here we develop a dual RNA cassette that decouples the detection and quantification tasks inherent to multiplex single-cell reporter assays. The resulting measurement of reporter expression is accurate over multiple orders of magnitude, with a precision approaching the limit set by Poisson counting noise. Together with RNA barcode stabilization via circularization, these scalable single-cell quantitative expression reporters provide high-contrast readouts, analogous to classic in situ assays but entirely from sequencing. Screening >200 regions of accessible chromatin in a multicellular in vitro model of early mammalian development, we identify 13 (8 previously uncharacterized) autonomous and cell-type-specific developmental CREs. We further demonstrate that chimeric CRE pairs generate cognate two-cell-type activity profiles and assess gain- and loss-of-function multicellular expression phenotypes from CRE variants with perturbed transcription factor binding sites. Single-cell quantitative expression reporters can be applied in developmental and multicellular systems to quantitatively characterize native, perturbed and synthetic CREs at scale, with high sensitivity and at single-cell resolution.
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  • 文章类型: Journal Article
    鉴于转录因子(TF)结合变化带来的复杂性,了解顺式调节元件对基因调节的影响提出了许多挑战。染色质可及性,结构限制,和细胞类型的差异。这篇综述讨论了基因调控网络在增强对转录调控的理解中的作用,并涵盖了从基于表达的方法到有监督的机器学习的构建方法。此外,关键的实验方法,包括MPRA和基于CRISPR-Cas9的筛查,这显著有助于理解TF结合偏好和顺式调节元件功能,正在探索。最后,分析了机器学习和人工智能解开顺式监管逻辑的潜力。这些计算上的进步对精准医学有着深远的影响,治疗靶点发现,以及健康和疾病遗传变异的研究。
    Understanding the influence of cis-regulatory elements on gene regulation poses numerous challenges given complexities stemming from variations in transcription factor (TF) binding, chromatin accessibility, structural constraints, and cell-type differences. This review discusses the role of gene regulatory networks in enhancing understanding of transcriptional regulation and covers construction methods ranging from expression-based approaches to supervised machine learning. Additionally, key experimental methods, including MPRAs and CRISPR-Cas9-based screening, which have significantly contributed to understanding TF binding preferences and cis-regulatory element functions, are explored. Lastly, the potential of machine learning and artificial intelligence to unravel cis-regulatory logic is analyzed. These computational advances have far-reaching implications for precision medicine, therapeutic target discovery, and the study of genetic variations in health and disease.
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  • 文章类型: Journal Article
    响应于扰动的差异基因表达至少部分地由转录因子(TF)和其他蛋白质在特定基因组区域的结合的变化介导。这些顺式调节元件(CREs)与其靶基因的关联是一项具有挑战性的任务,对于解决许多生物学和机理问题至关重要。许多目前的方法依赖于染色质构象捕获技术或单细胞相关方法来建立CRE与基因的关联。这些方法可能是有效的,但有局限性,包括决议,可检测的关联距离的间隙,和成本。作为替代,我们开发了DegCre,一种非参数方法,用于评估扰动诱导的差异基因表达的测量值与CREs的差异调节信号之间的相关性,以对可能的CRE-基因关联进行评分。它有几个独特的特点,包括以下能力:使用任何类型的CRE活性测量;产生CRE到基因对的概率评分;并在宽范围的序列距离上评估CRE到基因对。我们将DegCre应用于六个数据集,每个都采用不同的扰动,并包含各种调节信号测量,包括染色质开放性,组蛋白修饰,TF入住率。为了测试它们的功效,我们将DegCre关联与Hi-C循环调用和CRISPR验证的CRE到基因关联进行比较,通过DegCre建立与竞争方法相当或优于竞争方法的良好性能。DegCre是一种从扰动-差异角度将CREs与基因关联的新方法,具有与现有方法互补的优势,并允许对基因调控有新的见解。
    Differential gene expression in response to perturbations is mediated at least in part by changes in binding of transcription factors (TFs) and other proteins at specific genomic regions. Association of these cis-regulatory elements (CREs) with their target genes is a challenging task that is essential to address many biological and mechanistic questions. Many current approaches rely on chromatin conformation capture techniques or single-cell correlational methods to establish CRE-to-gene associations. These methods can be effective but have limitations, including resolution, gaps in detectable association distances, and cost. As an alternative, we have developed DegCre, a nonparametric method that evaluates correlations between measurements of perturbation-induced differential gene expression and differential regulatory signal at CREs to score possible CRE-to-gene associations. It has several unique features, including the ability to use any type of CRE activity measurement, yield probabilistic scores for CRE-to-gene pairs, and assess CRE-to-gene pairings across a wide range of sequence distances. We apply DegCre to six data sets, each using different perturbations and containing a variety of regulatory signal measurements, including chromatin openness, histone modifications, and TF occupancy. To test their efficacy, we compare DegCre associations to Hi-C loop calls and CRISPR-validated CRE-to-gene associations, establishing good performance by DegCre that is comparable or superior to competing methods. DegCre is a novel approach to the association of CREs to genes from a perturbation-differential perspective, with strengths that are complementary to existing approaches and allow for new insights into gene regulation.
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  • 文章类型: Journal Article
    心律失常的遗传易感性一直是研究的领域。研究最初集中在罕见的遗传性心律失常,但是在过去的二十年里,遗传变异(单核苷酸多态性)在心率中的作用,节奏,心律失常也被考虑在内。特别是,全基因组关联研究已经确定了数百个与定量心电图性状相关的基因组基因座,心房颤动,和不太常见的心律失常,如Brugada综合征。已发现大量相关变体系统地定位在控制编码转录因子的基因的组织特异性和时间转录的非编码调控元件中。离子通道,和其他蛋白质。然而,由于复杂的组织特异性,因果变异的鉴定及其对表型影响的机制已被证明是困难的,时间解决,依赖于条件,和调节元件的组合功能,以及它们在不同模型物种中的适度保护。在这次审查中,我们讨论了旨在鉴定和表征性状相关变异调节元件及其影响心率或心律的分子机制的研究工作.
    Genetic predisposition to cardiac arrhythmias has been a field of intense investigation. Research initially focused on rare hereditary arrhythmias, but over the last two decades, the role of genetic variation (single nucleotide polymorphisms) in heart rate, rhythm, and arrhythmias has been taken into consideration as well. In particular, genome-wide association studies have identified hundreds of genomic loci associated with quantitative electrocardiographic traits, atrial fibrillation, and less common arrhythmias such as Brugada syndrome. A significant number of associated variants have been found to systematically localize in non-coding regulatory elements that control the tissue-specific and temporal transcription of genes encoding transcription factors, ion channels, and other proteins. However, the identification of causal variants and the mechanism underlying their impact on phenotype has proven difficult due to the complex tissue-specific, time-resolved, condition-dependent, and combinatorial function of regulatory elements, as well as their modest conservation across different model species. In this review, we discuss research efforts aimed at identifying and characterizing-trait-associated variant regulatory elements and the molecular mechanisms underlying their impact on heart rate or rhythm.
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