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  • Review Article
  • Published:

Eukaryotic transcriptional dynamics: from single molecules to cell populations

Nature Reviews Genetics volume 14, pages 572–584 (2013)Cite this article

Subjects

Key Points

  • Single-molecule and genome-wide studies of transcription reveal the importance of dynamics for understanding the mechanisms of gene regulation.

  • A single gene can be regulated by dozens of factors interacting in a combinatorial manner. Observing all such interactions experimentally is still a daunting task, but computational models of transcription dynamics can provide insight into the underlying mechanisms.

  • Molecular models of transcription often emphasize sequential, ordered recruitment, for example in the formation of a pre-initiation complex at a promoter. These sequential processes are best-described by non-equilibrium thermodynamics, in which kinetics and energy dependence are treated explicitly.

  • Models based on non-equilibrium thermodynamics provide insight into a range of transcription phenomena, including nucleosome positioning, transcriptional bursting, and refractory periods during transcription.

Abstract

Transcriptional regulation is achieved through combinatorial interactions between regulatory elements in the human genome and a vast range of factors that modulate the recruitment and activity of RNA polymerase. Experimental approaches for studying transcription in vivo now extend from single-molecule techniques to genome-wide measurements. Parallel to these developments is the need for testable quantitative and predictive models for understanding gene regulation. These conceptual models must also provide insight into the dynamics of transcription and the variability that is observed at the single-cell level. In this Review, we discuss recent results on transcriptional regulation and also the models those results engender. We show how a non-equilibrium description informs our view of transcription by explicitly considering time- and energy-dependence at the molecular level.

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Figure 1: Points of view on transcriptional regulation.
Figure 2: Experimental techniques to study transcriptional kinetics.

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Acknowledgements

We would like to thank the members of the Transcription Imaging Consortium at the Janelia Farm Research Campus for their important contributions to the theories proposed here. We also thank B. Lewis for critical reading of the manuscript. This work is supported in part by funding from the US National Institutes of Health (NIH) grants GM57071, 84364 and 86217 to R.H.S. D.R.L. is supported by the Intramural Research Program of the NIH, National Cancer Institute, Center for Cancer Research.

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Authors and Affiliations

  1. Laboratory of Biological Modeling, National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK), National Institutes of Health, Bethesda, 20892, Maryland, USA

    Antoine Coulon & Carson C. Chow

  2. Department of Anatomy and Structural Biology, Albert Einstein College of Medicine, Bronx, 10461, New York, USA

    Robert H. Singer

  3. Janelia Farm Research Campus of the Howard Hughes Medical Institute (HHMI), Ashburn, 20147, Virginia, USA

    Robert H. Singer

  4. Laboratory of Receptor Biology and Gene Expression, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, 20892, Maryland, USA

    Daniel R. Larson

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  1. Antoine Coulon
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Correspondence to Daniel R. Larson.

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PowerPoint slides

Glossary

Chromatin remodellers

Along with chromatin modifiers, these are complexes and enzymes that affect the status of chromatin, through conformational changes (such as nucleosome displacement or eviction, or histone replacement) or by depositing or removing covalent marks on histone tails. These processes are accompanied by the hydrolysis of coenzymes, releasing chemical energy.

Chromatin immunoprecipitation

(ChIP). A method to assess the occupancy at a given genomic locus by a particular factor. It is carried out by amplifying DNA fragments that have been crosslinked to the factor of interest and pulled down using an antibody.

Re-ChIP

A modification of the chromatin immunoprecipitation (ChIP) method. By successively using several antibodies, it allows an assessment of the co-occupancy of a locus by multiple factors.

Equilibrium

For a chemical system to be at equilibrium, every reaction must occur in both directions with equal probability (or rate). Hence, a system can reach a steady state without ever being at equilibrium.

Steady state

Refers to a system that does not evolve over time. This concept may apply to various descriptions such as a set of concentrations of molecular species or the probability distribution of a set of features among a population of cells (for example, nucleosome positions on a specific promoter).

Fluorescence recovery after photobleaching

(FRAP). An experimental microscopy method to assess the mobility of molecules in living cells. In FRAP, the rate at which fluorescently labelled molecules repopulate a region of the cell that has been photobleached reflects both their diffusion and binding to chromatin.

Fluorescence correlation spectroscopy

(FCS). An experimental microscopy method to assess the mobility of molecules in living cells. In FCS, those properties are derived from the temporal fluctuations of fluorescence due to molecules entering and leaving a small optically defined volume of the cell.

MS2 and/or PP7 RNA labelling

A microscopy technique for labelling, in live cells, the transcripts from an artificial gene construct. Many molecules of fluorescent proteins (MS2 or PP7) bind each RNA on a specific cassette, thus allowing the monitoring of the number of nascent RNAs being transcribed at the gene locus over time.

Genomic nuclear run-on followed by high-throughput sequencing

(GRO-seq). This approach uses nuclear run-on methodology to map transcriptionally engaged polymerases on a genome-wide level. This approach constitutes a direct measure of transcriptional activity.

Flux

The net flux of a reaction is the difference between the rates at which it is observed to occur in one direction versus the other. When a system satisfies detailed balance, the net fluxes of all reactions are null.

Equilibrium constant

The ratio of association and dissociation rates. In an equilibrium context, this describes the affinity of a molecule for a binding site and directly relates to interaction energy. In a non-equilibrium context, such a ratio does not reflect the interaction energy and should not be called an equilibrium constant

Refractoriness

A time distribution function that displays an increasing phase for short delays is said to be refractory because — as opposed to an exponential distribution — the probability for the random event to occur is not constant but increases over time, thus shaping the distribution and reflecting the underlying biomolecular mechanics.

Memoryless

A process is memoryless if the time it takes to complete is exponentially distributed, indicating that its probability of completion does not change over time and is hence independent of the past. A succession of memoryless events (for example, sequential recruitment of factors) can lead to memory.

Single-molecule fluorescence in situ hybridization

(smFISH). A microscopy technique for labelling and visualizing RNA in fixed cells using many probes that are hybridized to a single transcript. For each cell, this technique allows the counting of the number of RNAs at the transcription site (that is, nascent RNAs) and also the number of RNAs in the cell.

Hill coefficient

A value describing the steepness of a dose–response curve at the level of transition between a low value and a high response. It reflects the level of cooperativity in the binding of the regulator and equals 1 in the case of uncooperative binding.

Metastable

A biochemical or conformational feature can be qualified as metastable if it is very unlikely to disappear spontaneously without the intervention of an energy- dependent enzymatic reaction. For example, post-transcriptional modifications of histone tails are metastable, as are certain conformational or remodelled states of chromatin.

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Coulon, A., Chow, C., Singer, R. et al. Eukaryotic transcriptional dynamics: from single molecules to cell populations. Nat Rev Genet 14, 572–584 (2013). https://doi.org/10.1038/nrg3484

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