A Comprehensive Atlas of Arabidopsis Regulatory DNA [DAP-seq]

We have developed a high throughput, next-generation DNA sequencing assay for rapid transcription factor binding site (TFBS) discovery in a genomic context. DNA affinity purification sequencing (DAP-seq), which uses affinity-purified transcription factors (TFs) to capture genomic DNA fragments, was applied to all 1,725 Arabidopsis thaliana TFs. High confidence TFBS motifs for 529 TFs and genome-wide enrichment maps for 349 factors were identified. In total,~ 2.7 million TFBS were identified which predict thousands of TF target genes enriched for known and novel functions.. Comparison of TF-binding using cytosine-methylated and -unmethylated genomic DNA revealed a 2-50 fold inhibition at methylated motifs for ~82% (264) of factors tested while 4.6% (15) showed stronger binding to methylated motifs. Finally, we describe how binding of Arabidopsis and maize Auxin Response Factors (ARFs) at phased motif repeats is highly enriched at ARF target gene promoters and how this architecture may allow for stabilization of dimers/multimers. Identification of sequence motifs for 530 transcription factors (529 Arabidopsis thaliana, 1 Zea mays) and genome-wide binding sites for 350 transcription factors (349 Arabidopsis thaliana, 1 Zea mays) by direct sequencing of affinity purified genomic DNA fragments. Idenfication of binding sites for 343 transcription factors (Arabidopsis thaliana) by direct sequencing of affinity purified, PCR amplified DNA libraries. Comparison to ChIP-Seq for ABI5 (AT2G36270). The 530 transcription factors refer to the number of unique proteins assayed on non-amplified DNA libraries (i.e. "characteristics: DNA source" = "col" or "zm"). Additional 5 factors have only data on amplified libraries (i.e. "characteristics: DNA source = colamp"), which do not represent the natural DNA methylation state of the organism (therefore not counted; more details in the "description" field). A subset of 350 of the 530 have both motif and TFBS identification (i.e. "characteristics: subset = TFBS+motif"), and the rest 180 have only motif identification (i.e. "characteristics: subset = motif").

我们开发了一种高通量下一代DNA测序检测方法，用于在基因组背景下快速发现转录因子结合位点（transcription factor binding site, TFBS）。该方法采用DNA亲和纯化测序（DNA affinity purification sequencing, DAP-seq）——即使用亲和纯化的转录因子（transcription factors, TFs）捕获基因组DNA片段——对拟南芥（Arabidopsis thaliana）的全部1725个转录因子进行了检测。最终共鉴定得到529个转录因子的高可信度TFBS基序，以及349个转录因子的全基因组富集图谱。总计鉴定得到约270万个TFBS，这些位点可预测得到数千个具有已知及新功能的转录因子靶基因。通过对比胞嘧啶甲基化与非甲基化基因组DNA的转录因子结合情况，研究发现约82%（264个）的检测转录因子在甲基化基序处的结合能力受到2至50倍的抑制，而4.6%（15个）的转录因子则对甲基化基序表现出更强的结合能力。本研究还阐述了拟南芥与玉米（Zea mays）的生长素响应因子（Auxin Response Factors, ARFs）在相位基序重复序列处的结合，在ARF靶基因启动子区域存在高度富集现象，且这种结构可能有助于二聚体/多聚体的稳定。本研究通过直接测序亲和纯化的基因组DNA片段，共鉴定得到530个转录因子的序列基序（其中529个来自拟南芥，1个来自玉米），以及350个转录因子的全基因组结合位点（其中349个来自拟南芥，1个来自玉米）。另有343个拟南芥转录因子的结合位点，通过对亲和纯化、PCR扩增的DNA文库进行直接测序得到鉴定。将该方法的转录因子结合检测结果与ABI5（AT2G36270）的染色质免疫共沉淀测序（ChIP-Seq）结果进行了对比。上述530个转录因子指的是在非扩增DNA文库（即"特征：DNA来源"为"col"或"zm"）中完成检测的独特蛋白质数量。另有5个转录因子仅在扩增文库（即"特征：DNA来源 = colamp"）中获得数据，由于扩增文库无法反映生物体天然的DNA甲基化状态，因此未被纳入统计（详细说明参见"描述"字段）。在530个转录因子中，有350个同时获得了基序与TFBS鉴定结果（即"特征：子集 = TFBS+motif"），剩余180个仅获得了基序鉴定结果（即"特征：子集 = motif"）。

搜集汇总

背景与挑战

背景概述

该数据集是一个基于DAP-seq技术构建的拟南芥调控DNA全面图谱，涵盖了1725个转录因子的结合位点分析，识别了约270万个转录因子结合位点，并揭示了DNA甲基化对结合的影响。数据集包含936个样本，提供了529个转录因子的高置信度基序和349个因子的全基因组结合位点图谱，为植物基因调控研究提供了重要资源。