Mammalian Genes Are Transcribed with Widely Different Bursting Kinetics

Real-time monitoring of gene expression reveals transcription kinetics of mammalian genes. In prokaryotes and eukaryotes, most genes appear to be transcribed during short periods called transcriptional bursts, interspersed by silent intervals. We describe how such bursts generate gene-specific temporal patterns of messenger RNA (mRNA) synthesis in mammalian cells. To monitor transcription at high temporal resolution, we established various gene trap cell lines and transgenic cell lines expressing a short-lived luciferase protein from an unstable mRNA, and recorded bioluminescence in real time in single cells. Mathematical modeling identified gene-specific on- and off-switching rates in transcriptional activity and mean numbers of mRNAs produced during the bursts. Transcriptional kinetics were markedly altered by cis-regulatory DNA elements. Our analysis demonstrated that bursting kinetics are highly gene-specific, reflecting refractory periods during which genes stay inactive for a certain time before switching on again.

[1]  Alexander van Oudenaarden,et al.  Single molecule fluorescent in situ hybridization (smFISH) of C. elegans worms and embryos. , 2012, WormBook : the online review of C. elegans biology.

[2]  Liat Rosenfeld,et al.  Single-allele analysis of transcription kinetics in living mammalian cells , 2010, Nature Methods.

[3]  Elizabeth A. Calle,et al.  Quantifying E. coli Proteome and Transcriptome with Single-Molecule Sensitivity in Single Cells , 2010, Science.

[4]  Alexander van Oudenaarden,et al.  Transcript counting in single cells reveals dynamics of rDNA transcription , 2010, Molecular systems biology.

[5]  Alexander van Oudenaarden,et al.  Variability in gene expression underlies incomplete penetrance , 2009, Nature.

[6]  Yoav Freund,et al.  Visualization of Individual Scr mRNAs during Drosophila Embryogenesis Yields Evidence for Transcriptional Bursting , 2009, Current Biology.

[7]  D. Larson,et al.  A single molecule view of gene expression. , 2009, Trends in cell biology.

[8]  Leah M. Octavio,et al.  Epigenetic and Conventional Regulation Is Distributed among Activators of FLO11 Allowing Tuning of Population-Level Heterogeneity in Its Expression , 2009, PLoS genetics.

[9]  John T. Lis,et al.  Defining mechanisms that regulate RNA polymerase II transcription in vivo , 2009, Nature.

[10]  M. Levine,et al.  Synchronous and Stochastic Patterns of Gene Activation in the Drosophila Embryo , 2009, Science.

[11]  Robert Tjian,et al.  Imaging transcription in living cells. , 2009, Annual review of biophysics.

[12]  D. Larson,et al.  Single-RNA counting reveals alternative modes of gene expression in yeast , 2008, Nature Structural &Molecular Biology.

[13]  M. Elowitz,et al.  Frequency-modulated nuclear localization bursts coordinate gene regulation , 2008, Nature.

[14]  J. Paulsson,et al.  Effects of Molecular Memory and Bursting on Fluctuations in Gene Expression , 2008, Science.

[15]  R. Singer,et al.  Transcriptional Pulsing of a Developmental Gene , 2006, Current Biology.

[16]  N. Friedman,et al.  Stochastic protein expression in individual cells at the single molecule level , 2006, Nature.

[17]  Esteban O. Mazzoni,et al.  Stochastic spineless expression creates the retinal mosaic for colour vision , 2006, Nature.

[18]  E. Cox,et al.  Real-Time Kinetics of Gene Activity in Individual Bacteria , 2005, Cell.

[19]  Felix Naef,et al.  Circadian Gene Expression in Individual Fibroblasts Cell-Autonomous and Self-Sustained Oscillators Pass Time to Daughter Cells , 2004, Cell.

[20]  R. Mantovani,et al.  The molecular biology of the CCAAT-binding factor NF-Y. , 1999, Gene.

[21]  J. Peccoud,et al.  Markovian Modeling of Gene-Product Synthesis , 1995 .

[22]  M. Ryan,et al.  Foot‐and‐mouth disease virus 2A oligopeptide mediated cleavage of an artificial polyprotein. , 1994, The EMBO journal.

[23]  D. Tranchina,et al.  Mechanistic model of bursts in mRNA synthesis , 2006 .

[24]  Seung Hyun Yoo.  INAUGURAL ARTICLE: PERIOD2::LUCIFERASE real-time reporting of circadian dynamics reveals persistent circadian oscillations in mouse peripheral tissues , 2004 .