The proper activities of enhancers and gene promoters are essential for coordinated transcription within a cell. Although diverse methodologies have been developed to identify enhancers and promoters, most have tacitly assumed that these elements are distinct. However, studies have unexpectedly shown that regulatory elements may have both enhancer and promoter functions. Here we review these results, focusing on the factors that determine the promoter and/or enhancer activity of regulatory elements. We discuss emerging models that define regulatory elements by accessible DNA and their non-mutually-exclusive abilities to drive transcription initiation (promoter activity) and/or to enhance transcription at other such regions (enhancer activity).
This is a preview of subscription content, access via your institution
Open Access articles citing this article.
Intron-mediated enhancement of DIACYLGLYCEROL ACYLTRANSFERASE1 expression in energycane promotes a step change for lipid accumulation in vegetative tissues
Biotechnology for Biofuels and Bioproducts Open Access 14 October 2023
Nature Communications Open Access 21 September 2023
Symmetric inheritance of parental histones governs epigenome maintenance and embryonic stem cell identity
Nature Genetics Open Access 04 September 2023
Access Nature and 54 other Nature Portfolio journals
Get Nature+, our best-value online-access subscription
$29.99 / 30 days
cancel any time
Subscribe to this journal
Receive 12 print issues and online access
$189.00 per year
only $15.75 per issue
Rent or buy this article
Prices vary by article type
Prices may be subject to local taxes which are calculated during checkout
Maurano, M. T. et al. Systematic localization of common disease-associated variation in regulatory DNA. Science 337, 1190–1195 (2012).
Roadmap Epigenomics Consortium et al. Integrative analysis of 111 reference human epigenomes. Nature 518, 317–330 (2015).
Andersson, R. et al. An atlas of active enhancers across human cell types and tissues. Nature 507, 455–461 (2014). This is the largest study to date defining active enhancers from divergent TSSs of eRNAs. It demonstrates exosome-mediated decay of eRNAs and describes transcribed enhancers as being better validated in in vitro reporter assays than enhancers predicted from histone modifications.
Chatterjee, S. & Ahituv, N. Gene regulatory elements, major drivers of human disease. Annu. Rev. Genomics Hum. Genet. 18, 45–63 (2017).
Bradner, J. E., Hnisz, D. & Young, R. A. Transcriptional addiction in cancer. Cell 168, 629–643 (2017).
Miguel-Escalada, I., Pasquali, L. & Ferrer, J. Transcriptional enhancers: functional insights and role in human disease. Curr. Opin. Genet. Dev. 33, 71–76 (2015).
Haberle, V. & Stark, A. Eukaryotic core promoters and the functional basis of transcription initiation. Nat. Rev. Mol. Cell Biol. 19, 621–637 (2018).
Shlyueva, D., Stampfel, G. & Stark, A. Transcriptional enhancers: from properties to genome-wide predictions. Nat. Rev. Genet. 15, 272–286 (2014).
Beagrie, R. A. & Pombo, A. Gene activation by metazoan enhancers: diverse mechanisms stimulate distinct steps of transcription. BioEssays 38, 881–893 (2016).
Lenhard, B., Sandelin, A. & Carninci, P. Metazoan promoters: emerging characteristics and insights into transcriptional regulation. Nat. Rev. Genet. 13, 233–245 (2012).
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). This study demonstrates a unified divergent architecture of candidate enhancers and promoters and observes a correlation between H3K4me3 and transcription levels.
Andersson, R. et al. Nuclear stability and transcriptional directionality separate functionally distinct RNA species. Nat. Commun. 5, 5336 (2014).
Scruggs, B. S. et al. Bidirectional transcription arises from two distinct hubs of transcription factor binding and active chromatin. Mol. Cell 58, 1101–1112 (2015).
Chen, Y. et al. Principles for RNA metabolism and alternative transcription initiation within closely spaced promoters. Nat. Genet. 48, 984–994 (2016).
Koch, F. et al. Transcription initiation platforms and GTF recruitment at tissue-specific enhancers and promoters. Nat. Struct. Mol. Biol. 18, 956–963 (2011).
Andersson, R. et al. Human gene promoters are intrinsically bidirectional. Mol. Cell 60, 346–347 (2015).
Diao, Y. et al. A tiling-deletion-based genetic screen for cis-regulatory element identification in mammalian cells. Nat. Methods 14, 629–635 (2017).
Dao, L. T. M. et al. Genome-wide characterization of mammalian promoters with distal enhancer functions. Nat. Genet. 49, 1073–1081 (2017).
Rajagopal, N. et al. High-throughput mapping of regulatory DNA. Nat. Biotechnol. 34, 167–174 (2016).
Engreitz, J. M. et al. Local regulation of gene expression by lncRNA promoters, transcription and splicing. Nature 539, 452–455 (2016). Diao (2017), Dao (2017) and Engreitz (2016) have observed, using in vivo genome editing and in vitro MPRAs, gene promoters with high enhancer strengths.
De Santa, F. et al. A large fraction of extragenic RNA pol II transcription sites overlap enhancers. PLOS Biol. 8, e1000384 (2010).
Kim, T.-K. et al. Widespread transcription at neuronal activity-regulated enhancers. Nature 465, 182–187 (2010). De Santa (2010) and Kim (2010) are the first studies to observe bidirectional transcription of eRNAs from candidate enhancers at large scale.
Roeder, R. G. The role of general initiation factors in transcription by RNA polymerase II. Trends Biochem. Sci. 21, 327–335 (1996).
Smale, S. T. & Kadonaga, J. T. The RNA polymerase II core promoter. Annu. Rev. Biochem. 72, 449–479 (2003).
Kadonaga, J. T. Perspectives on the RNA polymerase II core promoter. Wiley Interdiscip. Rev. Dev. Biol. 1, 40–51 (2012).
Breathnach, R. & Chambon, P. Organization and expression of eucaryotic split genes coding for proteins. Annu. Rev. Biochem. 50, 349–383 (1981).
Carninci, P. et al. Genome-wide analysis of mammalian promoter architecture and evolution. Nat. Genet. 38, 626–635 (2006).
Orphanides, G., Lagrange, T. & Reinberg, D. The general transcription factors of RNA polymerase II. Genes Dev. 10, 2657–2683 (1996).
Dynlacht, B. D., Hoey, T. & Tjian, R. Isolation of coactivators associated with the TATA-binding protein that mediate transcriptional activation. Cell 66, 563–576 (1991).
Vaquerizas, J. M., Kummerfeld, S. K., Teichmann, S. A. & Luscombe, N. M. A census of human transcription factors: function, expression and evolution. Nat. Rev. Genet. 10, 252–263 (2009).
Lambert, S. A. et al. The human transcription factors. Cell 175, 598–599 (2018).
Vo, N. & Goodman, R. H. CREB-binding protein and p300 in transcriptional regulation. J. Biol. Chem. 276, 13505–13508 (2001).
Malik, S. & Roeder, R. G. Dynamic regulation of pol II transcription by the mammalian mediator complex. Trends Biochem. Sci. 30, 256–263 (2005).
Koutelou, E., Hirsch, C. L. & Dent, S. Y. R. Multiple faces of the SAGA complex. Curr. Opin. Cell Biol. 22, 374–382 (2010).
Spitz, F. & Furlong, E. E. M. Transcription factors: from enhancer binding to developmental control. Nat. Rev. Genet. 13, 613–626 (2012).
Wasserman, W. W. & Sandelin, A. Applied bioinformatics for the identification of regulatory elements. Nat. Rev. Genet. 5, 276–287 (2004).
Lettice, L. A. et al. A long-range Shh enhancer regulates expression in the developing limb and fin and is associated with preaxial polydactyly. Hum. Mol. Genet. 12, 1725–1735 (2003).
Schoenfelder, S. & Fraser, P. Long-range enhancer-promoter contacts in gene expression control. Nat. Rev. Genet. 20, 437–455 (2019).
Robson, M. I., Ringel, A. R. & Mundlos, S. Regulatory landscaping: how enhancer-promoter communication is sculpted in 3D. Mol. Cell 74, 1110–1122 (2019).
Moreau, P. et al. The SV40 72 base repair repeat has a striking effect on gene expression both in SV40 and other chimeric recombinants. Nucleic Acids Res. 9, 6047–6068 (1981).
Banerji, J., Rusconi, S. & Schaffner, W. Expression of a beta-globin gene is enhanced by remote SV40 DNA sequences. Cell 27, 299–308 (1981). Moreau (1981) and Banerji (1981) are the first studies to discover enhancers through the identification of a 72-bp repeat sequence that could enhance gene transcription in SV40. They further show that such enhancers can act both upstream and downstream of, and at various distances from, a reporter gene.
Benoist, C. & Chambon, P. In vivo sequence requirements of the SV40 early promotor region. Nature 290, 304–310 (1981).
Banerji, J., Olson, L. & Schaffner, W. A lymphocyte-specific cellular enhancer is located downstream of the joining region in immunoglobulin heavy chain genes. Cell 33, 729–740 (1983).
Neuberger, M. S. Expression and regulation of immunoglobulin heavy chain gene transfected into lymphoid cells. EMBO J. 2, 1373–1378 (1983).
Schaffner, W. Enhancers, enhancers — from their discovery to today’s universe of transcription enhancers. Biol. Chem. 396, 311–327 (2015).
Furlong, E. E. M. & Levine, M. Developmental enhancers and chromosome topology. Science 361, 1341–1345 (2018).
Catarino, R. R. & Stark, A. Assessing sufficiency and necessity of enhancer activities for gene expression and the mechanisms of transcription activation. Genes Dev. 32, 202–223 (2018).
Forrest, A. R. R. et al. A promoter-level mammalian expression atlas. Nature 507, 462–470 (2014).
Consortium, R. E. et al. Integrative analysis of 111 reference human epigenomes. Nature 518, 317–330 (2015).
Bernstein, B. E. et al. An integrated encyclopedia of DNA elements in the human genome. Nature 489, 57–74 (2012).
Arner, E. et al. Transcribed enhancers lead waves of coordinated transcription in transitioning mammalian cells. Science 347, 1010–1014 (2015).
Zavolan, M., van Nimwegen, E. & Gaasterland, T. Splice variation in mouse full-length cDNAs identified by mapping to the mouse genome. Genome Res. 12, 1377–1385 (2002).
Landry, J.-R., Mager, D. L. & Wilhelm, B. T. Complex controls: the role of alternative promoters in mammalian genomes. Trends Genet. 19, 640–648 (2003).
Valen, E. et al. Genome-wide detection and analysis of hippocampus core promoters using DeepCAGE. Genome Res. 19, 255–265 (2009).
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).
Core, L. & Adelman, K. Promoter-proximal pausing of RNA polymerase II: a nexus of gene regulation. Genes Dev. 33, 960–982 (2019).
Seila, A. C. et al. Divergent transcription from active promoters. Science 322, 1849–1851 (2008).
Preker, P. et al. RNA exosome depletion reveals transcription upstream of active human promoters. Science 322, 1851–1854 (2008). Core (2008), Seila (2008) and Preker (2008) are the first studies to independently observe divergent transcription from mammalian gene promoters.
Jensen, T. H., Jacquier, A. & Libri, D. Dealing with pervasive transcription. Mol. Cell 52, 473–484 (2013).
Seila, A. C., Core, L. J., Lis, J. T. & Sharp, P. A. Divergent transcription: a new feature of active promoters. Cell Cycle 8, 2557–2564 (2009).
Ntini, E. et al. Polyadenylation site-induced decay of upstream transcripts enforces promoter directionality. Nat. Struct. Mol. Biol. 20, 923–928 (2013).
Almada, A. E., Wu, X., Kriz, A. J., Burge, C. B. & Sharp, P. A. Promoter directionality is controlled by U1 snRNP and polyadenylation signals. Nature 499, 360–363 (2013).
Furey, T. S. ChIP-seq and beyond: new and improved methodologies to detect and characterize protein–DNA interactions. Nat. Rev. Genet. 13, 840–852 (2012).
Bernstein, B. E. et al. Methylation of histone H3 Lys 4 in coding regions of active genes. Proc. Natl Acad. Sci. USA 99, 8695–8700 (2002).
Lee, C. K., Shibata, Y., Rao, B., Strahl, B. D. & Lieb, J. D. Evidence for nucleosome depletion at active regulatory regions genome-wide. Nat. Genet. 36, 900–905 (2004).
Schübeler, D. et al. The histone modification pattern of active genes revealed through genome-wide chromatin analysis of a higher eukaryote. Genes Dev. 18, 1263–1271 (2004).
Roh, T., Ngau, W. C., Cui, K., Landsman, D. & Zhao, K. High-resolution genome-wide mapping of histone modifications. Nat. Biotechnol. 22, 1013–1016 (2004).
Song, L. et al. Open chromatin defined by DNaseI and FAIRE identifies regulatory elements that shape cell-type identity. Genome Res. 21, 1757–1767 (2011).
Kim, T. H. et al. A high-resolution map of active promoters in the human genome. Nature 436, 876–880 (2005).
Zeitlinger, J. et al. RNA polymerase stalling at developmental control genes in the Drosophila melanogaster embryo. Nat. Genet. 39, 1512–1516 (2007).
Muse, G. W. et al. RNA polymerase is poised for activation across the genome. Nat. Genet. 39, 1507–1511 (2007).
Klemm, S. L., Shipony, Z. & Greenleaf, W. J. Chromatin accessibility and the regulatory epigenome. Nat. Rev. Genet. 20, 207–220 (2019).
Creyghton, M. P. et al. Histone H3K27ac separates active from poised enhancers and predicts developmental state. Proc. Natl Acad. Sci. USA 107, 21931–21936 (2010).
Heintzman, N. D. et al. Distinct and predictive chromatin signatures of transcriptional promoters and enhancers in the human genome. Nat. Genet. 39, 311–318 (2007).
Robertson, A. G. et al. Genome-wide relationship between histone H3 lysine 4 mono- and tri-methylation and transcription factor binding. Genome Res. 18, 1906–1917 (2008). Heintzman (2007) and Robertson (2008) identify a signature of high H3K4me1 and low H3K4me3 at candidate enhancers that could be used to discern enhancers from promoters and predict their locations genome-wide.
Ernst, J. et al. Mapping and analysis of chromatin state dynamics in nine human cell types. Nature 473, 43–49 (2011).
Hoffman, M. M. et al. Unsupervised pattern discovery in human chromatin structure through genomic segmentation. Nat. Methods 9, 473–476 (2012).
Ernst, J. & Kellis, M. ChromHMM: automating chromatin-state discovery and characterization. Nat. Methods 9, 215–216 (2012).
Melgar, M. F., Collins, F. S. & Sethupathy, P. Discovery of active enhancers through bidirectional expression of short transcripts. Genome Biol. 12, R113 (2011).
Wu, H. et al. Tissue-specific RNA expression marks distant-acting developmental enhancers. PLOS Genet. 10, e1004610 (2014).
Andersson, R., Sandelin, A. & Danko, C. G. A unified architecture of transcriptional regulatory elements. Trends Genet. 31, 426–433 (2015).
van Arensbergen, J. et al. Genome-wide mapping of autonomous promoter activity in human cells. Nat. Biotechnol. 35, 145–153 (2017).
Landolin, J. M. et al. Sequence features that drive human promoter function and tissue specificity. Genome Res. 20, 890–898 (2010).
Barski, A. et al. High-resolution profiling of histone methylations in the human genome. Cell 129, 823–837 (2007).
Soares, L. M. et al. Determinants of histone H3K4 methylation patterns. Mol. Cell 68, 773–785.e6 (2017).
Pekowska, A. et al. H3K4 tri-methylation provides an epigenetic signature of active enhancers. EMBO J. 30, 4198–4210 (2011).
Henriques, T. et al. Widespread transcriptional pausing and elongation control at enhancers. Genes Dev. 32, 26–41 (2018).
Rennie, S. et al. Transcription start site analysis reveals widespread divergent transcription in D. melanogaster and core promoter-encoded enhancer activities. Nucleic Acids Res. 46, 5455–5469 (2018).
Mifsud, B. et al. Mapping long-range promoter contacts in human cells with high-resolution capture Hi-C. Nat. Genet. 47, 598–606 (2015).
Schoenfelder, S. et al. The pluripotent regulatory circuitry connecting promoters to their long-range interacting elements. Genome Res. 25, 582–597 (2015).
Javierre, B. M. et al. Lineage-specific genome architecture links enhancers and non-coding disease variants to target gene promoters. Cell 167, 1369–1384.e19 (2016).
Xu, Z., Wei, G., Chepelev, I., Zhao, K. & Felsenfeld, G. Mapping of INS promoter interactions reveals its role in long-range regulation of SYT8 transcription. Nat. Struct. Mol. Biol. 18, 372–378 (2011).
Andersson, R. Promoter or enhancer, what’s the difference? Deconstruction of established distinctions and presentation of a unifying model. BioEssays 37, 314–323 (2015).
Mikhaylichenko, O. et al. The degree of enhancer or promoter activity is reflected by the levels and directionality of eRNA transcription. Genes Dev. 32, 42–57 (2018).
Canver, M. C. et al. BCL11A enhancer dissection by Cas9-mediated in situ saturating mutagenesis. Nature 527, 192–197 (2015).
Korkmaz, G. et al. Functional genetic screens for enhancer elements in the human genome using CRISPR–Cas9. Nat. Biotechnol. 34, 192–198 (2016).
Sanjana, N. E. et al. High-resolution interrogation of functional elements in the noncoding genome. Science 353, 1545–1549 (2016).
Klann, T. S. et al. CRISPR–Cas9 epigenome editing enables high-throughput screening for functional regulatory elements in the human genome. Nat. Biotechnol. 35, 561–568 (2017).
Carleton, J. B., Berrett, K. C. & Gertz, J. Multiplex enhancer interference reveals collaborative control of gene regulation by estrogen receptor α-bound enhancers. Cell Syst. 5, 333–344.e5 (2017).
Simeonov, D. R. et al. Discovery of stimulation-responsive immune enhancers with CRISPR activation. Nature 549, 111–115 (2017).
Nguyen, T. A. et al. High-throughput functional comparison of promoter and enhancer activities. Genome Res. 26, 1023–1033 (2016). This is the first study to systematically compare the promoter and enhancer potentials of a large number of candidate sequences using MPRAs.
Weingarten-Gabbay, S. et al. Systematic interrogation of human promoters. Genome Res. 29, 171–183 (2019).
Mattioli, K. et al. High-throughput functional analysis of lncRNA core promoters elucidates rules governing tissue specificity. Genome Res. 29, 344–355 (2019).
Patwardhan, R. P. et al. Massively parallel functional dissection of mammalian enhancers in vivo. Nat. Biotechnol. 30, 265–270 (2012).
Melnikov, A. et al. Systematic dissection and optimization of inducible enhancers in human cells using a massively parallel reporter assay. Nat. Biotechnol. 30, 271–277 (2012).
Kheradpour, P. et al. Systematic dissection of regulatory motifs in 2000 predicted human enhancers using a massively parallel reporter assay. Genome Res. 23, 800–811 (2013).
Kwasnieski, J. C., Mogno, I., Myers, C. A., Corbo, J. C. & Cohen, B. A. Complex effects of nucleotide variants in a mammalian cis-regulatory element. Proc. Natl Acad. Sci. USA 109, 19498–19503 (2012).
Smith, R. P. et al. Massively parallel decoding of mammalian regulatory sequences supports a flexible organizational model. Nat. Genet. 45, 1021–1028 (2013).
Grossman, S. R. et al. Systematic dissection of genomic features determining transcription factor binding and enhancer function. Proc. Natl Acad. Sci. USA 114, E1291–E1300 (2017).
Shen, S. Q. et al. Massively parallel cis-regulatory analysis in the mammalian central nervous system. Genome Res. 26, 238–255 (2016).
Haberle, V. et al. Transcriptional cofactors display specificity for distinct types of core promoters. Nature 570, 122–126 (2019).
Kwasnieski, J. C., Fiore, C., Chaudhari, H. G. & Cohen, B. A. High-throughput functional testing of ENCODE segmentation predictions. Genome Res. 24, 1595–1602 (2014).
Chaudhari, H. G. & Cohen, B. A. Local sequence features that influence AP-1-regulatory activity. Genome Res. 28, 171–181 (2018).
Muerdter, F. et al. Resolving systematic errors in widely used enhancer activity assays in human cells. Nat. Methods 15, 141–149 (2018).
Fenouil, R. et al. CpG islands and GC content dictate nucleosome depletion in a transcription-independent manner at mammalian promoters. Genome Res. 22, 2399–2408 (2012).
Biddie, S. C. et al. Transcription factor AP1 potentiates chromatin accessibility and glucocorticoid receptor binding. Mol. Cell 43, 145–155 (2011).
Iwafuchi-Doi, M. & Zaret, K. S. Pioneer transcription factors in cell reprogramming. Genes Dev. 28, 2679–2692 (2014).
Nakayama, R. T. et al. SMARCB1 is required for widespread BAF complex-mediated activation of enhancers and bivalent promoters. Nat. Genet. 49, 1613–1623 (2017).
Colbran, L. L., Chen, L. & Capra, J. A. Sequence characteristics distinguish transcribed enhancers from promoters and predict their breadth of activity. Genetics 211, 1205–1217 (2019).
Farley, E. K. et al. Suboptimization of developmental enhancers. Science 350, 325–328 (2015).
Farley, E. K., Olson, K. M., Zhang, W., Rokhsar, D. S. & Levine, M. S. Syntax compensates for poor binding sites to encode tissue specificity of developmental enhancers. Proc. Natl Acad. Sci. USA 113, 6508–6513 (2016).
Cheng, Y. et al. Principles of regulatory information conservation between mouse and human. Nature 515, 371–375 (2014).
Kim, T.-K. & Shiekhattar, R. Architectural and functional commonalities between enhancers and promoters. Cell 162, 948–959 (2015).
Struhl, K. Transcriptional noise and the fidelity of initiation by RNA polymerase II. Nat. Struct. Mol. Biol. 14, 103–105 (2007).
Zabidi, M. A. et al. Enhancer–core–promoter specificity separates developmental and housekeeping gene regulation. Nature 518, 556–559 (2015).
Dickel, D. E. et al. Ultraconserved enhancers are required for normal development. Cell 172, 491–499.e15 (2018).
Pennacchio, L. A. et al. In vivo enhancer analysis of human conserved non-coding sequences. Nature 444, 499–502 (2006).
Kidwell, M. & Lisch, D. Transposable elements and host genome evolution. Trends Ecol. Evol. 15, 95–99 (2000).
Wu, X. & Sharp, P. A. Divergent transcription: a driving force for new gene origination? Cell 155, 990–996 (2013).
Carelli, F. N., Liechti, A., Halbert, J., Warnefors, M. & Kaessmann, H. Repurposing of promoters and enhancers during mammalian evolution. Nat. Commun. 9, 4066 (2018).
Halfon, M. S. Studying transcriptional enhancers: the founder fallacy, validation creep, and other biases. Trends Genet. 35, 93–103 (2019).
Ramirez-Carrozzi, V. R. et al. A unifying model for the selective regulation of inducible transcription by CpG islands and nucleosome remodeling. Cell 138, 114–128 (2009).
Kolovos, P., Knoch, T. A., Grosveld, F. G., Cook, P. R. & Papantonis, A. Enhancers and silencers: an integrated and simple model for their function. Epigenetics Chromatin 5, 1 (2012).
Sima, J. et al. Identifying cis elements for spatiotemporal control of mammalian DNA replication. Cell 176, 816–830.e18 (2019).
Kawai, J. et al. Functional annotation of a full-length mouse cDNA collection. Nature 409, 685–690 (2001).
Takahashi, H., Lassmann, T., Murata, M. & Carninci, P. 5′ end-centered expression profiling using cap-analysis gene expression and next-generation sequencing. Nat. Protoc. 7, 542–561 (2012).
Yamashita, R. et al. Genome-wide characterization of transcriptional start sites in humans by integrative transcriptome analysis. Genome Res. 21, 775–789 (2011).
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).
Kruesi, W. S., Core, L. J., Waters, C. T., Lis, J. T. & Meyer, B. J. Condensin controls recruitment of RNA polymerase II to achieve nematode X-chromosome dosage compensation. eLife 2, e00808 (2013).
Lam, M. T. Y. et al. Rev-Erbs repress macrophage gene expression by inhibiting enhancer-directed transcription. Nature 498, 511–515 (2013).
Mayer, A. et al. Native elongating transcript sequencing reveals human transcriptional activity at nucleotide resolution. Cell 161, 541–554 (2015).
Nojima, T. et al. Mammalian NET-seq reveals genome-wide nascent transcription coupled to RNA processing. Cell 161, 526–540 (2015).
Weber, C. M., Ramachandran, S. & Henikoff, S. Nucleosomes are context-specific, H2A.Z-modulated barriers to RNA polymerase. Mol. Cell 53, 819–830 (2014).
Nechaev, S. et al. Global analysis of short RNAs reveals widespread promoter-proximal stalling and arrest of Pol II in Drosophila. Science 327, 335–338 (2010).
Berman, B. P. et al. Exploiting transcription factor binding site clustering to identify cis-regulatory modules involved in pattern formation in the Drosophila genome. Proc. Natl Acad. Sci. USA 99, 757–762 (2002).
May, D. et al. Large-scale discovery of enhancers from human heart tissue. Nat. Genet. 44, 89–93 (2011).
Visel, A. et al. ChIP-seq accurately predicts tissue-specific activity of enhancers. Nature 457, 854–858 (2009).
Wang, Z. et al. Combinatorial patterns of histone acetylations and methylations in the human genome. Nat. Genet. 40, 897–903 (2008).
Thurman, R. E. et al. The accessible chromatin landscape of the human genome. Nature 489, 75–82 (2012).
Cusanovich, D. A. et al. A single-cell atlas of in vivo mammalian chromatin accessibility. Cell 174, 1309–1324.e18 (2018).
Pradeepa, M. M. et al. Histone H3 globular domain acetylation identifies a new class of enhancers. Nat. Genet. 48, 681–686 (2016).
Rajagopal, N. et al. Distinct and predictive histone lysine acetylation patterns at promoters, enhancers, and gene bodies. G3 4, 2051–2063 (2014).
Jinek, M. et al. A programmable dual-RNA-guided DNA endonuclease in adaptive bacterial immunity. Science 337, 816–821 (2012).
Qi, L. S. et al. Repurposing CRISPR as an RNA-guided platform for sequence-specific control of gene expression. Cell 152, 1173–1183 (2013).
Fulco, C. P. et al. Systematic mapping of functional enhancer-promoter connections with CRISPR interference. Science 354, 769–773 (2016).
Matharu, N. et al. CRISPR-mediated activation of a promoter or enhancer rescues obesity caused by haploinsufficiency. Science 363, eaau0629 (2019).
Lopes, R., Korkmaz, G. & Agami, R. Applying CRISPR–Cas9 tools to identify and characterize transcriptional enhancers. Nat. Rev. Mol. Cell Biol. 17, 597–604 (2016).
Patwardhan, R. P. et al. High-resolution analysis of DNA regulatory elements by synthetic saturation mutagenesis. Nat. Biotechnol. 27, 1173–1175 (2009).
Inoue, F. et al. A systematic comparison reveals substantial differences in chromosomal versus episomal encoding of enhancer activity. Genome Res. 27, 38–52 (2017).
Arnold, C. D. et al. Genome-wide quantitative enhancer activity maps identified by STARR-seq. Science 339, 1074–1077 (2013).
Arnold, C. D. et al. Genome-wide assessment of sequence-intrinsic enhancer responsiveness at single-base-pair resolution. Nat. Biotechnol. 35, 136–144 (2017).
R.A. was supported by the Independent Research Fund Denmark (6108-00038B), the European Research Council (ERC) under the European Union’s Horizon 2020 research and innovation programme (StG no. 638173), and the Novo Nordisk Foundation (NNF18OC0052570). A.S. was supported by the ERC under the European Union’s Horizon 2020 research and innovation programme (MSCA ITN pHioniC), the Lundbeck Foundation, the Danish Cancer Society, the Danish Council for Independent Research and Innovation Fund Denmark. We thank L. van Duin, M. Wu and A. Thieffry for critical comments on the manuscript.
The authors declare no competing interests.
Peer review information
Nature Reviews Genetics thanks K. Adelman and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
- RNA polymerase II
(RNAPII). An enzyme that catalyses the transcription of DNA to RNA, including mRNAs and many long non-coding RNAs.
- Transcription start site
(TSS). The first transcribed genomic nucleotide of a transcript.
- General transcription factors
(GTFs). Proteins that, together with RNA polymerase II, make up the pre-initiation complex.
- Pre-initiation complex
A polypeptide complex consisting of RNA polymerase II and general transcription factors. This forms around the transcription start site and primes RNA polymerase II for transcription.
- TATA box
A T/A-rich sequence that lies upstream (typically 24–30 bp) of transcription start sites, with a role in positioning the pre-initiation complex.
The initiator sequence; a sequence pattern often overlapping transcription start sites.
- Transcription factors
(TFs). Sequence-specific DNA-binding proteins with a role in regulating transcription.
- Nucleosome-depleted region
(NDR). A region depleted of nucleosomes, often identified by DNaseI hypersensitivity, and often carrying regulatory potential.
- Nascent-RNA techniques
A wide range of methods aimed at capturing RNA as it is being transcribed. See Box 2.
A method for identifying accessible regions of the genome, based on DNaseI hypersensitivity. See Box 3.
- Promoter-upstream transcript
(PROMPT). A short RNA (also known as upstream antisense RNA (uaRNA)) that is transcribed upstream and on the opposite strand from an mRNA transcription start site and is typically degraded by the nuclear exosome. It has many similarities to an enhancer RNA.
- Nuclear exosome
A multi-protein complex responsible for the degradation of RNAs from the 3′ end.
- Poly(A) sites
Sequence patterns (AT/ATAA) associated with the 3′ ends of genes, but that also occur with high frequency in intergenic DNA.
Chromatin immunoprecipitation coupled to sequencing; a method for finding DNA–protein interactions by combining immunoprecipitation and high-throughput DNA sequencing. See Box 3.
- Enhancer RNAs
(eRNAs). Short RNAs (<500 bp) that are transcribed from enhancers, with many similarities to promoter-upstream transcripts (PROMPTs).
Assay for transposase-accessible chromatin; a method for identifying accessible regions of the genome, based on transposase activity.
- CpG islands
Genomic sequences that are not depleted of cytosine–phosphate–guanine (CpG) dinucleotides, which would occur by 5-methylcytosine deamination. They often overlap or are near transcription start sites. Most definitions set a minimum length (for example, 200 or 500 bp) and a minimum observed/expected CpG ratio.
Cap analysis of gene expression; a method to identify transcription start sites by sequencing the 5′ ends of capped, steady-state RNAs. See Box 2.
- Expression quantitative trait loci
(eQTLs). Regions of DNA in which genetic variation is associated with variability in the expression of one or more genes.
- Massively parallel reporter assays
(MPRAs). Methods that can measure the promoter or enhancer activity of many candidate DNA sequences in parallel. See Box 5.
- DNase hypersensitive sites
(DHSs). Highly accessible genomic regions identified by DNase-seq.
- Pioneer transcription factors
Transcription factors that can directly bind nucleosomal DNA, possibly in compacted chromatin.
- Homotypic attraction
A force driving chromatin with similar characteristics or associated proteins to self-associate. The physical association may be formed by protein bridges or liquid–liquid phase separation, the latter of which involves molecules separating into liquid condensates with specific compositions.
- Ascertainment bias
Drawing general conclusions based on biased sampling of non-representative examples.
- Validation creep
Treating predictions as validated entities, in either the same study or subsequent studies. This type of bias is related to ascertainment bias and overfitting.
About this article
Cite this article
Andersson, R., Sandelin, A. Determinants of enhancer and promoter activities of regulatory elements. Nat Rev Genet 21, 71–87 (2020). https://doi.org/10.1038/s41576-019-0173-8
This article is cited by
Genome Biology (2023)
Study of the regulatory elements of the Ovalbumin gene promoter using CRISPR technology in chicken cells
Journal of Biological Engineering (2023)
Understanding the roles and regulation patterns of circRNA on its host gene in tumorigenesis and tumor progression
Journal of Experimental & Clinical Cancer Research (2023)
Multiple Fra-1-bound enhancers showing different molecular and functional features can cooperate to repress gene transcription
Cell & Bioscience (2023)
Intron-mediated enhancement of DIACYLGLYCEROL ACYLTRANSFERASE1 expression in energycane promotes a step change for lipid accumulation in vegetative tissues
Biotechnology for Biofuels and Bioproducts (2023)