Transcriptional Bursting from the HIV-1 Promoter Is a Significant Source of Stochastic Noise in HIV-1 Gene Expression
TL;DR: In this paper, the authors quantified expression noise from the long terminal repeat (LTR) promoter at different HIV-1 integration sites across the human genome and found that the measured noise levels are inconsistent with constitutive gene expression models, and that each burst generates an average of 2-10 mRNA transcripts before the promoter returned to an inactive state.
About: This article is published in Biophysical Journal.The article was published on 2010-04-21 and is currently open access. It has received 241 citations till now. The article focuses on the topics: HIV Long Terminal Repeat & Regulation of gene expression.
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TL;DR: The identification of replication-competent noninduced proviruses indicates that the size of the latent reservoir-and, hence, the barrier to cure-may be up to 60-fold greater than previously estimated.
1,160 citations
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TL;DR: Stochastic computational modeling successfully accounted for PheB and correctly predicted the dynamics of a Tat mutant that were subsequently confirmed by experiment, illustrating the importance of stochastic fluctuations in gene expression in a mammalian system.
Abstract: Stochastic gene expression has been implicated in a variety of cellular processes, including cell differentiation and disease. In this issue of Cell, Weinberger et al. (2005) take an integrated computational-experimental approach to study the Tat transactivation feedback loop in HIV-1 and show that fluctuations in a key regulator, Tat, can result in a phenotypic bifurcation. This phenomenon is observed in an isogenic population where individual cells display two distinct expression states corresponding to latent and productive infection by HIV-1. These findings demonstrate the importance of stochastic gene expression in molecular "decision-making."
536 citations
TL;DR: Time-lapse fluorescence microscopy is used to analyze 8,000 individual human genomic loci and finds that at virtually all loci, episodic bursting—as opposed to constitutive expression—is the predominant mode of expression.
Abstract: Gene expression occurs either as an episodic process, characterized by pulsatile bursts, or as a constitutive process, characterized by a Poisson-like accumulation of gene products. It is not clear which mode of gene expression (constitutive versus bursty) predominates across a genome or how transcriptional dynamics are influenced by genomic position and promoter sequence. Here, we use time-lapse fluorescence microscopy to analyze 8,000 individual human genomic loci and find that at virtually all loci, episodic bursting—as opposed to constitutive expression—is the predominant mode of expression. Quantitative analysis of the expression dynamics at these 8,000 loci indicates that both the frequency and size of the transcriptional bursts varies equally across the human genome, independent of promoter sequence. Strikingly, weaker expression loci modulate burst frequency to increase activity, whereas stronger expression loci modulate burst size to increase activity. Transcriptional activators such as trichostatin A (TSA) and tumor necrosis factor α (TNF) only modulate burst size and frequency along a constrained trend line governed by the promoter. In summary, transcriptional bursting dominates across the human genome, both burst frequency and burst size vary by chromosomal location, and transcriptional activators alter burst frequency and burst size, depending on the expression level of the locus.
453 citations
Cites background or methods or result from "Transcriptional Bursting from the H..."
...delayed switching to the transcriptional ON state in a two-state transcription model (17) and predicts that noise frequency (not only noise magnitude) (27) is modulated in different genomic or chromatin environments (SI Appendix)....
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...Transcriptional burst dynamics are quantified by deriving analytical expressions for burst size and burst frequency with formulations from previous analyses (13, 14, 17) and low promoter activity assumptions where koff ≫ kon, koff ≫ km, koff ≫ γp and km ≫ ðγm þ γpÞ):...
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...We previously reported that TNF only changes burst frequency of the LTR while conserving burst size in a limited number of isoclones (17), and were interested to see how widespread this phenomena was across the genome....
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...To generate an initial baseline origin for noise-space analysis of the single-cell data, we identified isoclones that were the most Poissonian in their expression fluctuation profiles from a library of isoclonal populations (17) (SI Appendix)....
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...S4), which is consistent with our previous study (17) showing that the LTR promoter displays relatively higher levels of noise than other eukaryotic promoters in yeast (6, 7)....
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TL;DR: This work quantifies copy-number statistics of mRNA from 20 Escherichia coli promoters using single-molecule fluorescence in situ hybridization to characterize the general properties of transcriptional time series and finds that the degree of burstiness is correlated with gene expression level but is largely independent of other parameters of gene regulation.
Abstract: Gene activity is described by the time series of discrete, stochastic mRNA production events. This transcriptional time series shows intermittent, bursty behavior. One consequence of this temporal intricacy is that gene expression can be tuned by varying different features of the time series. Here we quantify copy-number statistics of mRNA from 20 Escherichia coli promoters using single-molecule fluorescence in situ hybridization in order to characterize the general properties of these transcriptional time series. We find that the degree of burstiness is correlated with gene expression level but is largely independent of other parameters of gene regulation. The observed behavior can be explained by the underlying variation in the duration of bursting events. Using Shannon's mutual information function, we estimate the mutual information transmitted between an outside stimulus, such as the extracellular concentration of inducer molecules, and intracellular levels of mRNA. This suggests that the outside stimulus transmits information reflected in the properties of transcriptional time series.
396 citations
TL;DR: It is shown that positive supercoiling buildup on a DNA segment by transcription slows down transcription elongation and eventually stops transcription initiation, and proves that transcriptional bursting of highly expressed genes in bacteria is primarily caused by reversible gyrase dissociation from and rebinding to aDNA segment, changing the supercoiled level of the segment.
357 citations
Cites background from "Transcriptional Bursting from the H..."
..., 2012), and fluorescent protein (Singh et al., 2010) or luciferase (Suter et al....
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...…single mRNA production in real time (Chubb et al., 2006; Golding et al., 2005; Hocine et al., 2013; Larson et al., 2011; Lionnet et al., 2011; Muramoto et al., 2012), and fluorescent protein (Singh et al., 2010) or luciferase (Suter et al., 2011) as gene expression reporter in live cells....
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References
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TL;DR: In this paper, an exact method is presented for numerically calculating, within the framework of the stochastic formulation of chemical kinetics, the time evolution of any spatially homogeneous mixture of molecular species which interreact through a specified set of coupled chemical reaction channels.
5,875 citations
TL;DR: It is demonstrated that the requirement for the tat gene can be offset by placing constitutive promoters upstream of the vector transcript, and the improved design presented here should facilitate testing of lentivirus vectors.
Abstract: Vectors derived from human immunodeficiency virus (HIV) are highly efficient vehicles for in vivo gene delivery. However, their biosafety is of major concern. Here we exploit the complexity of the HIV genome to provide lentivirus vectors with novel biosafety features. In addition to the structural genes, HIV contains two regulatory genes, tat and rev, that are essential for HIV replication, and four accessory genes that encode critical virulence factors. We previously reported that the HIV type 1 accessory open reading frames are dispensable for efficient gene transduction by a lentivirus vector. We now demonstrate that the requirement for the tat gene can be offset by placing constitutive promoters upstream of the vector transcript. Vectors generated from constructs containing such a chimeric long terminal repeat (LTR) transduced neurons in vivo at very high efficiency, whether or not they were produced in the presence of Tat. When the rev gene was also deleted from the packaging construct, expression of gag and pol was strictly dependent on Rev complementation in trans. By the combined use of a separate nonoverlapping Rev expression plasmid and a 5' LTR chimeric transfer construct, we achieved optimal yields of vector of high transducing efficiency (up to 10(7) transducing units [TU]/ml and 10(4) TU/ng of p24). This third-generation lentivirus vector uses only a fractional set of HIV genes: gag, pol, and rev. Moreover, the HIV-derived constructs, and any recombinant between them, are contingent on upstream elements and trans complementation for expression and thus are nonfunctional outside of the vector producer cells. This split-genome, conditional packaging system is based on existing viral sequences and acts as a built-in device against the generation of productive recombinants. While the actual biosafety of the vector will ultimately be proven in vivo, the improved design presented here should facilitate testing of lentivirus vectors.
3,063 citations
TL;DR: Stochastic gene expression has important consequences for cellular function, being beneficial in some contexts and harmful in others, including the stress response, metabolism, development, the cell cycle, circadian rhythms, and aging.
2,471 citations
TL;DR: The results of a genome-wide analysis of human cells suggest that most protein-coding genes, including most genes thought to be transcriptionally inactive, experience transcription initiation, and that transcription initiation at most genes is a general phenomenon in human cells.
1,927 citations
TL;DR: Global analysis of cellular transcription indicated that active genes were preferential integration targets, particularly genes that were activated in cells after infection by HIV-1, and this data suggests how selective targeting promotes aggressive HIV replication.
1,808 citations