دورية أكاديمية
Single-cell nascent RNA sequencing unveils coordinated global transcription.
| العنوان: | Single-cell nascent RNA sequencing unveils coordinated global transcription. |
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| المؤلفون: | Mahat DB; Koch Institute for Integrative Cancer Research and Department of Biology, Massachusetts Institute of Technology, Cambridge, MA, USA., Tippens ND; Koch Institute for Integrative Cancer Research and Department of Biology, Massachusetts Institute of Technology, Cambridge, MA, USA., Martin-Rufino JD; Broad Institute of MIT and Harvard, Cambridge, MA, USA., Waterton SK; Koch Institute for Integrative Cancer Research and Department of Biology, Massachusetts Institute of Technology, Cambridge, MA, USA.; Department of Biology, Stanford University, Stanford, CA, USA., Fu J; Koch Institute for Integrative Cancer Research and Department of Biology, Massachusetts Institute of Technology, Cambridge, MA, USA.; Interdisciplinary Biological Sciences Graduate Program, Northwestern University, Evanston, IL, USA., Blatt SE; Koch Institute for Integrative Cancer Research and Department of Biology, Massachusetts Institute of Technology, Cambridge, MA, USA.; Exact Sciences, Madison, WI, USA., Sharp PA; Koch Institute for Integrative Cancer Research and Department of Biology, Massachusetts Institute of Technology, Cambridge, MA, USA. sharppa@mit.edu. |
| المصدر: | Nature [Nature] 2024 Jul; Vol. 631 (8019), pp. 216-223. Date of Electronic Publication: 2024 Jun 05. |
| نوع المنشور: | Journal Article |
| اللغة: | English |
| بيانات الدورية: | Publisher: Nature Publishing Group Country of Publication: England NLM ID: 0410462 Publication Model: Print-Electronic Cited Medium: Internet ISSN: 1476-4687 (Electronic) Linking ISSN: 00280836 NLM ISO Abbreviation: Nature Subsets: MEDLINE |
| أسماء مطبوعة: | Publication: Basingstoke : Nature Publishing Group Original Publication: London, Macmillan Journals ltd. |
| مواضيع طبية MeSH: | Enhancer Elements, Genetic*/genetics , Gene Expression Regulation*/genetics , Promoter Regions, Genetic*/genetics , RNA*/analysis , RNA*/biosynthesis , RNA*/genetics , Sequence Analysis, RNA*/methods , Single-Cell Gene Expression Analysis*/methods , Transcription, Genetic*, Animals ; Humans ; Mice ; Cell Cycle/genetics ; Click Chemistry/methods ; DNA-Directed RNA Polymerases/analysis ; DNA-Directed RNA Polymerases/metabolism ; Histones/metabolism ; Time Factors |
| مستخلص: | Transcription is the primary regulatory step in gene expression. Divergent transcription initiation from promoters and enhancers produces stable RNAs from genes and unstable RNAs from enhancers 1,2 . Nascent RNA capture and sequencing assays simultaneously measure gene and enhancer activity in cell populations 3 . However, fundamental questions about the temporal regulation of transcription and enhancer-gene coordination remain unanswered, primarily because of the absence of a single-cell perspective on active transcription. In this study, we present scGRO-seq-a new single-cell nascent RNA sequencing assay that uses click chemistry-and unveil coordinated transcription throughout the genome. We demonstrate the episodic nature of transcription and the co-transcription of functionally related genes. scGRO-seq can estimate burst size and frequency by directly quantifying transcribing RNA polymerases in individual cells and can leverage replication-dependent non-polyadenylated histone gene transcription to elucidate cell cycle dynamics. The single-nucleotide spatial and temporal resolution of scGRO-seq enables the identification of networks of enhancers and genes. Our results suggest that the bursting of transcription at super-enhancers precedes bursting from associated genes. By imparting insights into the dynamic nature of global transcription and the origin and propagation of transcription signals, we demonstrate the ability of scGRO-seq to investigate the mechanisms of transcription regulation and the role of enhancers in gene expression. (© 2024. The Author(s).) |
| التعليقات: | Update of: bioRxiv. 2023 Sep 19:2023.09.15.558015. doi: 10.1101/2023.09.15.558015. (PMID: 37745427) |
| References: | Birney, E. et al. Identification and analysis of functional elements in 1% of the human genome by the ENCODE pilot project. Nature 447, 799–816 (2007). (PMID: 17571346) Core, L. J., Waterfall, J. J. & Lis, J. T. Nascent RNA sequencing reveals widespread pausing and divergent initiation at human promoters. Science 322, 1845–1848 (2008). (PMID: 190569412833333) Core, L. J. et al. Analysis of nascent RNA identifies a unified architecture of initiation regions at mammalian promoters and enhancers. Nat. Genet. 46, 1311–1320 (2014). (PMID: 253839684254663) Chubb, J. R., Trcek, T., Shenoy, S. M. & Singer, R. H. Transcriptional pulsing of a developmental gene. Curr. Biol. 16, 1018–1025 (2006). (PMID: 167139604764056) Raj, A., Peskin, C. S., Tranchina, D., Vargas, D. Y. & Tyagi, S. Stochastic mRNA synthesis in mammalian cells. PLoS Biol. 4, e309 (2006). (PMID: 170489831563489) Fukaya, T., Lim, B. & Levine, M. Enhancer control of transcriptional bursting. Cell 166, 358–368 (2016). (PMID: 272931914970759) Larsson, A. J. M. et al. Genomic encoding of transcriptional burst kinetics. Nature 424, 147 (2019). Hah, N. et al. A rapid, extensive, and transient transcriptional response to estrogen signaling in breast cancer cells. Cell 145, 622–634 (2011). (PMID: 215494153099127) Long, H. K., Prescott, S. L. & Wysocka, J. Ever-changing landscapes: transcriptional enhancers in development and evolution. Cell 167, 1170–1187 (2016). (PMID: 278632395123704) Levo, M. et al. Transcriptional coupling of distant regulatory genes in living embryos. Nature 605, 754–760 (2022). (PMID: 355086629886134) Hindorff, L. A. et al. Potential etiologic and functional implications of genome-wide association loci for human diseases and traits. Proc. Natl Acad. Sci. USA 106, 9362–9367 (2009). (PMID: 194742942687147) Shlyueva, D., Stampfel, G. & Stark, A. Transcriptional enhancers: from properties to genome-wide predictions. Nat. Rev. Genet. 15, 272–286 (2014). (PMID: 24614317) Tippens, N. D. et al. Transcription imparts architecture, function and logic to enhancer units. Nat. Genet. 52, 1067–1075 (2020). (PMID: 329589507541647) Kwak, H., Fuda, N. J., Core, L. J. & Lis, J. T. Precise maps of RNA polymerase reveal how promoters direct initiation and pausing. Science 339, 950–953 (2013). (PMID: 234306543974810) Kolb, H. C., Finn, M. G. & Sharpless, K. B. Click chemistry: diverse chemical function from a few good reactions. Angew. Chem. Int. Ed. 40, 2004–2021 (2001). Core, L. J. et al. Defining the status of RNA polymerase at promoters. Cell Rep. 2, 1025–1035 (2012). (PMID: 230627133483431) Shah, S. et al. Dynamics and spatial genomics of the nascent transcriptome by intron seqFISH. Cell 174, 363–376.e16 (2018). (PMID: 298873816046268) Levsky, J. M., Shenoy, S. M., Pezo, R. C. & Singer, R. H. Single-cell gene expression profiling. Science 297, 836–840 (2002). (PMID: 12161654) Femino, A. M., Fay, F. S., Fogarty, K. & Singer, R. H. Visualization of single RNA transcripts in situ. Science 280, 585–590 (1998). (PMID: 9554849) Erhard, F. et al. scSLAM-seq reveals core features of transcription dynamics in single cells. Nature 571, 419–423 (2019). (PMID: 31292545) Muse, G. W. et al. RNA polymerase is poised for activation across the genome. Nat. Genet. 39, 1507–1511 (2007). (PMID: 179940212365887) Deng, Q., Ramsköld, D., Reinius, B. & Sandberg, R. Single-cell RNA-seq reveals dynamic, random monoallelic gene expression in mammalian cells. Science 343, 193–196 (2014). (PMID: 24408435) Jonkers, I., Kwak, H. & Lis, J. T. Genome-wide dynamics of Pol II elongation and its interplay with promoter proximal pausing, chromatin, and exons. eLife 3, e02407 (2014). (PMID: 248430274001325) Suter, D. M. et al. Mammalian genes are transcribed with widely different bursting kinetics. Science 332, 472–474 (2011). (PMID: 21415320) Ramalingam, V., Natarajan, M., Johnston, J. & Zeitlinger, J. TATA and paused promoters active in differentiated tissues have distinct expression characteristics. Mol. Syst. Biol. 17, e9866 (2021). (PMID: 335438297863008) Pimmett, V. L. et al. Quantitative imaging of transcription in living Drosophila embryos reveals the impact of core promoter motifs on promoter state dynamics. Nat. Commun. 12, 4504 (2021). (PMID: 343019368302612) Subramanian, A. et al. Gene set enrichment analysis: a knowledge-based approach for interpreting genome-wide expression profiles. Proc. Natl Acad. Sci. USA 102, 15545–15550 (2005). (PMID: 161995171239896) Xie, X. et al. Systematic discovery of regulatory motifs in human promoters and 3′ UTRs by comparison of several mammals. Nature 434, 338–345 (2005). (PMID: 157356392923337) Patange, S. et al. MYC amplifies gene expression through global changes in transcription factor dynamics. Cell Rep. 38, 110292 (2022). (PMID: 350813488849550) Ochiai, H. et al. Genome-wide kinetic properties of transcriptional bursting in mouse embryonic stem cells. Sci. Adv. 6, eaaz6699 (2020). (PMID: 325964487299619) Whitfield, M. L. et al. Identification of genes periodically expressed in the human cell cycle and their expression in tumors. Mol. Biol. Cell 13, 1977–2000 (2002). (PMID: 12058064117619) Salmen, F. et al. High-throughput total RNA sequencing in single cells using VASA-seq. Nat. Biotechnol. 40, 1780–1793 (2022). (PMID: 357609149750877) McKellar, D. W. et al. Spatial mapping of the total transcriptome by in situ polyadenylation. Nat. Biotechnol. 41, 513–520 (2023). (PMID: 36329320) Robbins, E. & Borun, T. W. The cytoplasmic synthesis of histones in HELA cells and its temporal relationship to DNA replication. Proc. Natl Acad. Sci. USA 57, 409–416 (1967). (PMID: 16591485335521) Marzluff, W. F., Wagner, E. J. & Duronio, R. J. Metabolism and regulation of canonical histone mRNAs: life without a poly(A) tail. Nat. Rev. Genet. 9, 843–854 (2008). (PMID: 189275792715827) Riba, A. et al. Cell cycle gene regulation dynamics revealed by RNA velocity and deep-learning. Nat. Commun. 13, 2865 (2022). (PMID: 356063839126911) Waisman, A. et al. Cell cycle dynamics of mouse embryonic stem cells in the ground state and during transition to formative pluripotency. Sci Rep. 9, 8051 (2019). (PMID: 311427856541595) Beyrouthy, M. J. et al. Identification of G1-regulated genes in normally cycling human cells. PLoS ONE 3, e3943 (2008). (PMID: 190797742600614) van Dam, S., Võsa, U., van der Graaf, A., Franke, L. & de Magalhães, J. P. Gene co-expression analysis for functional classification and gene–disease predictions. Brief. Bioinform. 19, 575–592 (2017). (PMID: 6054162) Zhang, Y. et al. Regulation of cell cycle progression by forkhead transcription factor FOXO3 through its binding partner DNA replication factor Cdt1. Proc. Natl Acad. Sci. USA 109, 5717–5722 (2012). (PMID: 224519353326503) Whyte, W. A. et al. Master transcription factors and mediator establish super-enhancers at key cell identity genes. Cell 153, 307–319 (2013). (PMID: 235823223653129) Ohtsuka, M., Inoko, H., Kulski, J. K. & Yoshimura, S. Major histocompatibility complex (Mhc) class Ib gene duplications, organization and expression patterns in mouse strain C57BL/6. BMC Genomics 9, 178 (2008). (PMID: 184168562375909) Agrawal, P. et al. Genome editing demonstrates that the −5 kb Nanog enhancer regulates Nanog expression by modulating RNAPII initiation and/or recruitment. J. Biol. Chem. 296, 100189 (2021). (PMID: 33334884) Tesar, P. J. et al. New cell lines from mouse epiblast share defining features with human embryonic stem cells. Nature 448, 196–199 (2007). (PMID: 17597760) Li, Y. et al. CRISPR reveals a distal super-enhancer required for Sox2 expression in mouse embryonic stem cells. PLoS ONE 9, e114485 (2014). (PMID: 254862554259346) Xie, L. et al. A dynamic interplay of enhancer elements regulates Klf4 expression in naive pluripotency. Genes Dev. 31, 1795–1808 (2017). (PMID: 289827625666677) Moorthy, S. D. et al. Enhancers and super-enhancers have an equivalent regulatory role in embryonic stem cells through regulation of single or multiple genes. Genome Res. 27, 246–258 (2017). (PMID: 278951095287230) Mahat, D. B. et al. Base-pair-resolution genome-wide mapping of active RNA polymerases using precision nuclear run-on (PRO-seq). Nat. Protoc. 11, 1455–1476 (2016). (PMID: 274428635502525) Habib, N. et al. Div-Seq: single-nucleus RNA-seq reveals dynamics of rare adult newborn neurons. Science 353, 925–928 (2016). (PMID: 274712525480621) Wu, J., McKeague, M. & Sturla, S. J. Nucleotide-resolution genome-wide mapping of oxidative DNA damage by Click-Code-Seq. J. Am. Chem. Soc. 140, 9783–9787 (2018). (PMID: 29944356) Miller, G. P. & Kool, E. T. Versatile 5′-functionalization of oligonucleotides on solid support: amines, azides, thiols, and thioethers via phosphorus chemistry. J. Org. Chem. 69, 2404–2410 (2004). (PMID: 15049637) Zhou, L., Myers, A. N., Vandersteen, J. G., Wang, L. & Wittwer, C. T. Closed-tube genotyping with unlabeled oligonucleotide probes and a saturating DNA dye. Clin. Chem. 50, 1328–1335 (2004). (PMID: 15166111) Gu, W. et al. Depletion of abundant sequences by hybridization (DASH): using Cas9 to remove unwanted high-abundance species in sequencing libraries and molecular counting applications. Genome Biol. 17, 2408 (2016). Marcel, M. Cutadapt removes adapter sequences from high-throughput sequencing reads. EMBnet J. 17, 10–12 (2011). Dodt, M., Roehr, J. T., Ahmed, R. & Dieterich, C. FLEXBAR—flexible barcode and adapter processing for next-generation sequencing platforms. Biology 1, 895–905 (2012). (PMID: 248325234009805) Langmead, B. & Salzberg, S. L. Fast gapped-read alignment with Bowtie 2. Nat. Methods 9, 357–359 (2012). (PMID: 223882863322381) Smith, T., Heger, A. & Sudbery, I. UMI-tools: modeling sequencing errors in unique molecular identifiers to improve quantification accuracy. Genome Res. 27, 491–499 (2017). (PMID: 281005845340976) Audibert, A., Weil, D. & Dautry, F. In vivo kinetics of mRNA splicing and transport in mammalian cells. Mol. Cell. Biol. 22, 6706–6718 (2002). (PMID: 12215528134034) Clement, J. Q., Qian, L., Kaplinsky, N. & Wilkinson, M. F. The stability and fate of a spliced intron from vertebrate cells. RNA 5, 206–220 (1999). (PMID: 100241731369753) Coulon, A. et al. Kinetic competition during the transcription cycle results in stochastic RNA processing. eLife 3, e03939 (2014). (PMID: 252713744210818) Neugebauer, K. M. Nascent RNA and the coordination of splicing with transcription. Cold Spring Harb. Perspect. Biol. 11, a032227 (2019). (PMID: 313713516671939) Rabani, M. et al. Metabolic labeling of RNA uncovers principles of RNA production and degradation dynamics in mammalian cells. Nat. Biotechnol. 29, 436–442 (2011). (PMID: 215160853114636) Rabani, M. et al. High-resolution sequencing and modeling identifies distinct dynamic RNA regulatory strategies. Cell 159, 1698–1710 (2014). (PMID: 254975484272607) Singh, J. & Padgett, R. A. Rates of in situ transcription and splicing in large human genes. Nat. Struct. Mol. Biol. 16, 1128–1133 (2009). (PMID: 198207122783620) Danko, C. G. et al. Identification of active transcriptional regulatory elements from GRO-seq data. Nat. Methods 12, 433–438 (2015). (PMID: 257994414507281) Dreos, R., Ambrosini, G., Groux, R., Cavin Périer, R. & Bucher, P. The eukaryotic promoter database in its 30th year: focus on non-vertebrate organisms. Nucleic Acids Res. 45, D51–D55 (2017). (PMID: 27899657) Tome, J. M., Tippens, N. D. & Lis, J. T. Single-molecule nascent RNA sequencing identifies regulatory domain architecture at promoters and enhancers. Nat. Genet. 322, 1845 (2018). Satija, R., Farrell, J. A., Gennert, D., Schier, A. F. & Regev, A. Spatial reconstruction of single-cell gene expression data. Nat. Biotechnol. 33, 495–502 (2015). (PMID: 258679234430369) Chae, M., Danko, C. G. & Kraus, W. L. groHMM: a computational tool for identifying unannotated and cell type-specific transcription units from global run-on sequencing data. BMC Bioinformatics 16, 222 (2015). (PMID: 261734924502638) Hu, S. et al. Transcription factor antagonism regulates heterogeneity in embryonic stem cell states. Mol. Cell 82, 4410–4427.e12 (2022). (PMID: 363565839722640) |
| معلومات مُعتمدة: | P01 CA042063 United States CA NCI NIH HHS; P30 CA014051 United States CA NCI NIH HHS; R01 GM034277 United States GM NIGMS NIH HHS |
| المشرفين على المادة: | EC 2.7.7.6 (DNA-Directed RNA Polymerases) 0 (Histones) 63231-63-0 (RNA) |
| تواريخ الأحداث: | Date Created: 20240605 Date Completed: 20240703 Latest Revision: 20240708 |
| رمز التحديث: | 20240709 |
| مُعرف محوري في PubMed: | PMC11222150 |
| DOI: | 10.1038/s41586-024-07517-7 |
| PMID: | 38839954 |
| قاعدة البيانات: | MEDLINE |
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Full Text Finder | تدمد: | 1476-4687 |
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| DOI: | 10.1038/s41586-024-07517-7 |