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NET-prism 能够实现以核苷酸分辨率专门针对 RNA 聚合酶的转录探测。

NET-prism enables RNA polymerase-dedicated transcriptional interrogation at nucleotide resolution.

机构信息

a Max Planck Research Group 'Chromatin and Ageing', Max Planck Institute for Biology of Ageing , Cologne , Germany.

b Cologne Excellence Cluster on Cellular Stress Responses in Ageing Associated Diseases (CECAD), University of Cologne , Cologne , Germany.

出版信息

RNA Biol. 2019 Sep;16(9):1156-1165. doi: 10.1080/15476286.2019.1621625. Epub 2019 Jun 3.

DOI:10.1080/15476286.2019.1621625
PMID:31156037
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC6693550/
Abstract

The advent of quantitative approaches that enable interrogation of transcription at single nucleotide resolution has allowed a novel understanding of transcriptional regulation previously undefined. However, little is known, at such high resolution, how transcription factors directly influence RNA Pol II pausing and directionality. To map the impact of transcription/elongation factors on transcription dynamics genome-wide at base pair resolution, we developed an adapted NET-seq protocol called NET-prism (Native Elongating Transcription by Polymerase-Regulated Immunoprecipitants in the Mammalian genome). Application of NET-prism on elongation factors (Spt6, Ssrp1), splicing factors (Sf1), and components of the pre-initiation complex (PIC) (TFIID, and Mediator) reveals their inherent command on transcription dynamics, with regards to directionality and pausing over promoters, splice sites, and enhancers/super-enhancers. NET-prism will be broadly applicable as it exposes transcription factor/Pol II dependent topographic specificity and thus, a new degree of regulatory complexity during gene expression.

摘要

定量方法的出现使得能够在单个核苷酸分辨率下研究转录，从而对以前无法定义的转录调控有了新的认识。然而，在如此高的分辨率下，人们对转录因子如何直接影响 RNA Pol II 的暂停和方向性知之甚少。为了在碱基对分辨率上绘制转录/延伸因子对转录动力学的影响图谱，我们开发了一种经过改良的 NET-seq 方案，称为 NET-prism（通过聚合酶调控免疫沉淀在哺乳动物基因组中进行的天然延伸转录）。NET-prism 在延伸因子（Spt6、Ssrp1）、剪接因子（Sf1）和起始前复合物（PIC）组件（TFIID 和 Mediator）上的应用揭示了它们对启动子、剪接位点和增强子/超级增强子上转录方向性和暂停的固有控制。NET-prism 将具有广泛的适用性，因为它揭示了转录因子/Pol II 依赖性的地形特异性，从而在基因表达过程中增加了一个新的调控复杂性程度。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ece3/6693550/7cad29ed4f2e/krnb-16-09-1621625-g005.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ece3/6693550/02c29f55b699/krnb-16-09-1621625-g001.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ece3/6693550/9a0e697c5381/krnb-16-09-1621625-g002.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ece3/6693550/25eb36b8a767/krnb-16-09-1621625-g003.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ece3/6693550/3f295bcbb259/krnb-16-09-1621625-g004.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ece3/6693550/7cad29ed4f2e/krnb-16-09-1621625-g005.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ece3/6693550/02c29f55b699/krnb-16-09-1621625-g001.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ece3/6693550/9a0e697c5381/krnb-16-09-1621625-g002.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ece3/6693550/25eb36b8a767/krnb-16-09-1621625-g003.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ece3/6693550/3f295bcbb259/krnb-16-09-1621625-g004.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ece3/6693550/7cad29ed4f2e/krnb-16-09-1621625-g005.jpg

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