scholarly journals Structural Coupling between RNA Polymerase Composition and DNA Supercoiling in Coordinating Transcription: a Global Role for the Omega Subunit?

mBio ◽  
2011 ◽  
Vol 2 (4) ◽  
Author(s):  
Marcel Geertz ◽  
Andrew Travers ◽  
Sanja Mehandziska ◽  
Patrick Sobetzko ◽  
Sarath Chandra Janga ◽  
...  
Keyword(s):  
Gene Expression ◽  
Rna Polymerase ◽  
Dna Supercoiling ◽  
Dna Topology ◽  
Chromosomal Dna ◽  
Tight Coupling ◽  
Structural Coupling ◽  
Bacterial Physiology ◽  
Transcription Machinery ◽  
The Impact

ABSTRACT In growing bacterial cells, the global reorganization of transcription is associated with alterations of RNA polymerase composition and the superhelical density of the DNA. However, the existence of any regulatory device coordinating these changes remains elusive. Here we show that in an exponentially growing Escherichia coli rpoZ mutant lacking the polymerase ω subunit, the impact of the Eσ38 holoenzyme on transcription is enhanced in parallel with overall DNA relaxation. Conversely, overproduction of σ70 in an rpoZ mutant increases both overall DNA supercoiling and the transcription of genes utilizing high negative superhelicity. We further show that transcription driven by the Eσ38 and Eσ70 holoenzymes from cognate promoters induces distinct superhelical densities of plasmid DNA in vivo. We thus demonstrate a tight coupling between polymerase holoenzyme composition and the supercoiling regimen of genomic transcription. Accordingly, we identify functional clusters of genes with distinct σ factor and supercoiling preferences arranging alternative transcription programs sustaining bacterial exponential growth. We propose that structural coupling between DNA topology and holoenzyme composition provides a basic regulatory device for coordinating genome-wide transcription during bacterial growth and adaptation. IMPORTANCE Understanding the mechanisms of coordinated gene expression is pivotal for developing knowledge-based approaches to manipulating bacterial physiology, which is a problem of central importance for applications of biotechnology and medicine. This study explores the relationships between variations in the composition of the transcription machinery and chromosomal DNA topology and suggests a tight interdependence of these two variables as the major coordinating principle of gene regulation. The proposed structural coupling between the transcription machinery and DNA topology has evolutionary implications and suggests a new methodology for studying concerted alterations of gene expression during normal and pathogenic growth both in bacteria and in higher organisms.

scholarly journals Collective polymerase dynamics emerge from DNA Supercoiling Mediated Transcription

2021 ◽  
Author(s):  
Stuart Sevier ◽  
Sahand Hormoz
Keyword(s):  
Gene Expression ◽  
Rna Polymerase ◽  
Critical Role ◽  
Regulation Of Gene Expression ◽  
Dna Supercoiling ◽  
Toggle Switch ◽  
Endogenous Genes ◽  
Non Local ◽  
Physical Interactions ◽  
The Impact

All biological processes ultimately come from physical interactions. The mechanical properties of DNA play a critical role in transcription. RNA polymerase can over or under twist DNA (referred to as DNA supercoiling) when it moves along a gene resulting in mechanical stresses in DNA that impact its own motion and that of other polymerases. For example, when enough supercoiling accumulates, an isolated polymerase halts and transcription stops. DNA supercoiling can also mediate non-local interactions between polymerases that shape gene expression fluctuations. Here, we construct a comprehensive model of transcription that captures how RNA polymerase motion changes the degree of DNA supercoiling which in turn feeds back into the rate at which polymerases are recruited and move along the DNA. Surprisingly, our model predicts that a group of three or more polymerases move together at a constant velocity and sustain their motion (forming what we call a polymeton) whereas one or two polymerases would have halted. We further show that accounting for the impact of DNA supercoiling on both RNA polymerase recruitment and velocity recapitulates empirical observations of gene expression fluctuations. Finally, we propose a mechanical toggle switch whereby interactions between genes are mediated by DNA twisting as opposed to proteins. Understanding the mechanical regulation of gene expression provides new insights into how endogenous genes can interact and informs the design of new forms of engineered interactions.

scholarly journals Properties of Gene Expression and Chromatin Structure with Mechanically Regulated Transcription

2018 ◽  
Author(s):  
Stuart A. Sevier ◽  
Herbert Levine
Keyword(s):  
Gene Expression ◽  
Mechanical Properties ◽  
Dna Structure ◽  
Genetic System ◽  
Dna Supercoiling ◽  
Simple Description ◽  
Gene Orientation ◽  
Physical Elements ◽  
Transcriptional Bursting ◽  
The Impact

The mechanical properties of transcription have emerged as central elements in our understanding of gene expression. Recent work has been done introducing a simple description of the basic physical elements of transcription where RNA elongation, RNA polymerase (RNAP) rotation and DNA supercoiling are coupled [1]. Here we generalize this framework to accommodate the behavior of many RNAPs operating on multiple genes on a shared piece of DNA. The resulting framework is combined with well-established stochastic processes of transcription resulting in a model which characterizes the impact of the mechanical properties of transcription on gene expression and DNA structure. Transcriptional bursting readily emerges as a common phenomenon with origins in the geometric nature of the genetic system and results in the bounding of gene expression statistics. Properties of a multiple gene system are examined with special attention paid to role that genome composition (gene orientation, size, and intergenic distance) plays in the ability of genes to transcribe. The role of transcription in shaping DNA structure is examined and the possibility of transcription driven domain formation is discussed.PACS numbers:

scholarly journals Quantitative modelling predicts the impact of DNA methylation on RNA polymerase II traffic

2019 ◽  
Vol 116 (30) ◽  
pp. 14995-15000 ◽  
Author(s):  
Justyna Cholewa-Waclaw ◽  
Ruth Shah ◽  
Shaun Webb ◽  
Kashyap Chhatbar ◽  
Bernard Ramsahoye ◽  
...  
Keyword(s):  
Gene Expression ◽  
Dna Methylation ◽  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Rett Syndrome ◽  
Transcription Initiation ◽  
Expression Patterns ◽  
Global Gene Expression ◽  
A Genome ◽  
The Impact

Patterns of gene expression are primarily determined by proteins that locally enhance or repress transcription. While many transcription factors target a restricted number of genes, others appear to modulate transcription levels globally. An example is MeCP2, an abundant methylated-DNA binding protein that is mutated in the neurological disorder Rett syndrome. Despite much research, the molecular mechanism by which MeCP2 regulates gene expression is not fully resolved. Here, we integrate quantitative, multidimensional experimental analysis and mathematical modeling to indicate that MeCP2 is a global transcriptional regulator whose binding to DNA creates “slow sites” in gene bodies. We hypothesize that waves of slowed-down RNA polymerase II formed behind these sites travel backward and indirectly affect initiation, reminiscent of defect-induced shockwaves in nonequilibrium physics transport models. This mechanism differs from conventional gene-regulation mechanisms, which often involve direct modulation of transcription initiation. Our findings point to a genome-wide function of DNA methylation that may account for the reversibility of Rett syndrome in mice. Moreover, our combined theoretical and experimental approach provides a general method for understanding how global gene-expression patterns are choreographed.

scholarly journals Regulation of Chlamydia Gene Expression by Tandem Promoters with Different Temporal Patterns

2015 ◽  
Vol 198 (2) ◽  
pp. 363-369 ◽  
Author(s):  
Christopher J. Rosario ◽  
Ming Tan
Keyword(s):  
Gene Expression ◽  
Transcriptional Regulation ◽  
Transcription Factors ◽  
Rna Polymerase ◽  
Expression Patterns ◽  
Dna Supercoiling ◽  
Early Gene ◽  
Developmental Cycle ◽  
Content Type ◽  
Intracellular Infection

ABSTRACTChlamydiais a genus of pathogenic bacteria with an unusual intracellular developmental cycle marked by temporal waves of gene expression. The three main temporal groups of chlamydial genes are proposed to be controlled by separate mechanisms of transcriptional regulation. However, we have noted genes with discrepancies, such as the early genednaKand the midcycle genesbioYandpgk, which have promoters controlled by the late transcriptional regulators EUO and σ28. To resolve this issue, we analyzed the promoters of these three genesin vitroand inChlamydia trachomatisbacteria grown in cell culture. Transcripts from the σ28-dependent promoter of each gene were detected only at late times in the intracellular infection, bolstering the role of σ28RNA polymerase in late gene expression. In each case, however, expression prior to late times was due to a second promoter that was transcribed by σ66RNA polymerase, which is the major form of chlamydial polymerase. These results demonstrate that chlamydial genes can be transcribed from tandem promoters with different temporal profiles, leading to a composite expression pattern that differs from the expression profile of a single promoter. In addition, tandem promoters allow a gene to be regulated by multiple mechanisms of transcriptional regulation, such as DNA supercoiling or late regulation by EUO and σ28. We discuss how tandem promoters broaden the repertoire of temporal gene expression patterns in the chlamydial developmental cycle and can be used to fine-tune the expression of specific genes.IMPORTANCEChlamydiais a pathogenic bacterium that is responsible for the majority of infectious disease cases reported to the CDC each year. It causes an intracellular infection that is characterized by coordinated expression of chlamydial genes in temporal waves. Chlamydial transcription has been shown to be regulated by DNA supercoiling, alternative forms of RNA polymerase, and transcription factors, but the number of transcription factors found inChlamydiais far fewer than the number found in most bacteria. This report describes the use of tandem promoters that allow the temporal expression of a gene or operon to be controlled by more than one regulatory mechanism. This combinatorial strategy expands the range of expression patterns that are available to regulate chlamydial genes.

scholarly journals New Insights into Structural and Functional Roles of Indole-3-acetic acid (IAA): Changes in DNA Topology and Gene Expression in Bacteria

Biomolecules ◽  
2019 ◽  
Vol 9 (10) ◽  
pp. 522 ◽  
Author(s):  
Defez ◽  
Valenti ◽  
Andreozzi ◽  
Romano ◽  
Ciaramella ◽  
...  
Keyword(s):  
Gene Expression ◽  
Acetic Acid ◽  
Conformational Changes ◽  
Regulation Of Gene Expression ◽  
Dna Supercoiling ◽  
Dna Topology ◽  
In Vitro Assays ◽  
Bacterial Cells ◽  
Indole 3 Acetic Acid

: Indole-3-acetic acid (IAA) is a major plant hormone that affects many cellular processes in plants, bacteria, yeast, and human cells through still unknown mechanisms. In this study, we demonstrated that the IAA-treatment of two unrelated bacteria, the Ensifer meliloti 1021 and Escherichia coli, harboring two different host range plasmids, influences the supercoiled state of the two plasmid DNAs in vivo. Results obtained from in vitro assays show that IAA interacts with DNA, leading to DNA conformational changes commonly induced by intercalating agents. We provide evidence that IAA inhibits the activity of the type IA topoisomerase, which regulates the DNA topological state in bacteria, through the relaxation of the negative supercoiled DNA. In addition, we demonstrate that the treatment of E. meliloti cells with IAA induces the expression of some genes, including the ones related to nitrogen fixation. In contrast, these genes were significantly repressed by the treatment with novobiocin, which reduces the DNA supercoiling in bacterial cells. Taking into account the overall results reported, we hypothesize that the IAA action and the DNA structure/function might be correlated and involved in the regulation of gene expression. This work points out that checking whether IAA influences the DNA topology under physiological conditions could be a useful strategy to clarify the mechanism of action of this hormone, not only in plants but also in other unrelated organisms.

scholarly journals How Single-Molecule Localization Microscopy Expanded Our Mechanistic Understanding of RNA Polymerase II Transcription

2021 ◽  
Vol 22 (13) ◽  
pp. 6694
Author(s):  
Peter Hoboth ◽  
Ondřej Šebesta ◽  
Pavel Hozák
Keyword(s):  
Gene Expression ◽  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Single Molecule ◽  
Spatial Organization ◽  
Super Resolution ◽  
Temporal Organization ◽  
Transcription Machinery ◽  
Mechanistic Understanding ◽  
Rnapii Transcription

Classical models of gene expression were built using genetics and biochemistry. Although these approaches are powerful, they have very limited consideration of the spatial and temporal organization of gene expression. Although the spatial organization and dynamics of RNA polymerase II (RNAPII) transcription machinery have fundamental functional consequences for gene expression, its detailed studies have been abrogated by the limits of classical light microscopy for a long time. The advent of super-resolution microscopy (SRM) techniques allowed for the visualization of the RNAPII transcription machinery with nanometer resolution and millisecond precision. In this review, we summarize the recent methodological advances in SRM, focus on its application for studies of the nanoscale organization in space and time of RNAPII transcription, and discuss its consequences for the mechanistic understanding of gene expression.

scholarly journals How Single-Molecule Localization Microscopy Expanded Our Mechanistic Understanding of RNA Polymerase II Transcription

2021 ◽  
Author(s):  
Peter Hoboth ◽  
Ondřej Šebesta ◽  
Pavel Hozak
Keyword(s):  
Gene Expression ◽  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Single Molecule ◽  
Spatial Organization ◽  
Super Resolution ◽  
Temporal Organization ◽  
Transcription Machinery ◽  
Mechanistic Understanding ◽  
Rnapii Transcription

Classical models of gene expression were built using genetics and biochemistry. Although these approaches are powerful, they have very limited consideration of the spatial and temporal organization of gene expression. Although the spatial organization and dynamics of RNA polymerase II (RNAPII) transcription machinery has fundamental functional consequences for gene expression, its detailed studies have been for long time abrogated by the limits of classical light microscopy. The advent of super-resolution microscopy (SRM) techniques allowed for the visualization of the RNAPII transcription machinery with nanometer resolution and millisecond precision. In this review, we summarize the recent methodological advances in SRM, focus on its application for studies of the nanoscale organization in space and time of RNAPII transcription, and discuss its consequences for the mechanistic understanding of gene expression.

scholarly journals Quantitative modelling predicts the impact of DNA methylation on RNA polymerase II traffic

2018 ◽  
Author(s):  
Justyna Cholewa-Waclaw ◽  
Ruth Shah ◽  
Shaun Webb ◽  
Kashyap Chhatbar ◽  
Bernard Ramsahoye ◽  
...  
Keyword(s):  
Gene Expression ◽  
Dna Methylation ◽  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Rett Syndrome ◽  
Transcription Initiation ◽  
Expression Patterns ◽  
Global Gene Expression ◽  
A Genome ◽  
The Impact

Patterns of gene expression are primarily determined by proteins that locally enhance or repress transcription. While many transcription factors target a restricted number of genes, others appear to modulate transcription levels globally. An example is MeCP2, an abundant methylated-DNA binding protein that is mutated in the neurological disorder Rett Syndrome. Despite much research, the molecular mechanism by which MeCP2 regulates gene expression is not fully resolved. Here we integrate quantitative, multi-dimensional experimental analysis and mathematical modelling to show that MeCP2 is a novel type of global transcriptional regulator whose binding to DNA creates "slow sites" in gene bodies. Waves of slowed-down RNA polymerase II formed behind these sites travel backward and indirectly affect initiation, reminiscent of defect-induced shock waves in non-equilibrium physics transport models. This mechanism differs from conventional gene regulation mechanisms, which often involve direct modulation of transcription initiation. Our findings uncover a genome-wide function of DNA methylation that may account for the reversibility of Rett syndrome in mice. Moreover, our combined theoretical and experimental approach provides a general method for understanding how global gene expression patterns are choreographed.

scholarly journals The dormancy-specific regulator, SutA, is intrinsically disordered and modulates transcription initiation in Pseudomonas aeruginosa

2018 ◽  
Author(s):  
Megan Bergkessel ◽  
Brett M. Babin ◽  
David G. VanderVelde ◽  
Michael J. Sweredoski ◽  
Annie Moradian ◽  
...  
Keyword(s):  
Gene Expression ◽  
Pseudomonas Aeruginosa ◽  
Rna Polymerase ◽  
Sigma Factor ◽  
Growth Arrest ◽  
Rrna Genes ◽  
Bacterial Physiology ◽  
Intrinsically Disordered ◽  
Intrinsically Disordered Regions ◽  
Structural Methods

ABSTRACTThough bacteria in nature are often nutritionally limited and growing slowly, most of our understanding of core cellular processes such as transcription comes from studies in a handful of model organisms doubling rapidly under nutrient-replete conditions. We previously identified a small protein of unknown function, called SutA, in a global screen of proteins synthesized in Pseudomonas aeruginosa under growth arrest (Babin BM, et al. (2016) SutA is a bacterial transcription factor expressed during slow growth in Pseudomonas aeruginosa. PNAS 113(5):E597-605). SutA binds RNA polymerase (RNAP), causing widespread changes in gene expression, including upregulation of the ribosomal RNA (rRNA) genes. Here, using biochemical and structural methods, we examine how SutA interacts with RNAP and the functional consequences of these interactions. We show that SutA consists of a central α-helix with unstructured N- and C-terminal tails, and binds to the β1 domain of RNAP. It activates transcription from the P. aeruginosa rrn promoter by both the housekeeping sigma factor holoenzyme (Eσ70) and the general stress response sigma factor holoenzyme (EσS) in vitro, and its N-terminal tail is required for activation in both holoenzyme contexts. However, we find that the interaction between SutA and each holoenzyme is distinct, with the SutA C-terminal tail and an acidic loop unique to σ70 playing the determining roles in these differences. Our results add SutA to a growing list of transcription regulators that use their intrinsically disordered regions to remodel transcription complexes.SIGNIFICANCELittle is known about how bacteria regulate their activities during periods of dormancy, yet growth arrest dominates bacterial existence in most environments and is directly relevant to the problem of physiological antibiotic tolerance. Though much is known about transcription in the model organism, Escherichia coli, even there, our understanding of gene expression during dormancy is incomplete. Here we explore how transcription under growth arrest is modulated in Pseudomonas aeruginosa by the small acidic protein, SutA. We show that SutA binds to RNA polymerase and controls transcription by a mechanism that is distinct from other known regulators. Our work underscores the potential for fundamental, mechanistic discovery in this important and understudied realm of bacterial physiology.

scholarly journals The Yersinia enterocolitica pYV Virulence Plasmid Contains Multiple Intrinsic DNA Bends Which Melt at 37°C

1999 ◽  
Vol 181 (14) ◽  
pp. 4198-4204 ◽  
Author(s):  
John R. Rohde ◽  
Xing-she Luan ◽  
Harold Rohde ◽  
James M. Fox ◽  
S. A. Minnich
Keyword(s):  
Gene Expression ◽  
Yersinia Enterocolitica ◽  
Conformational Transition ◽  
Dna Supercoiling ◽  
Dna Topology ◽  
Phenotypic Switching ◽  
Virulence Plasmid ◽  
Secretion Systems ◽  
Temperature Dependent ◽  
Virulence Gene Expression

ABSTRACT Temperature has a pleiotropic effect on Yersinia enterocolitica gene expression. Temperature-dependent phenotypes include the switching between two type III protein secretion systems, flagellum biosynthesis (≤30°C) and virulence plasmid-encoded Yop secretion (37°C). The mechanism by which temperature exerts this change in genetic programming is unclear; however, altered gene expression by temperature-dependent changes in DNA topology has been implicated. Here, we present evidence that the Y. enterocolitica virulence plasmid, pYV, undergoes a conformational transition between 30 and 37°C. Using a simplified two-dimensional, single-gel assay, we show that pYV contains multiple regions of intrinsic curvature, including virF, the positive activator of virulence genes. These bends are detectable at 30°C but melt at 37°C, the temperature at which the cells undergo phenotypic switching. We also show that pACYC184, a plasmid used as a reporter of temperature-induced changes in DNA supercoiling, has a single region of intrinsic bending detected by our assay. Topoisomers of pACYC184, with and without this bend, isolated from Y. enterocolitica were resolved by using chloroquine gels. The single bend has a dramatic influence on temperature-dependent DNA supercoiling. These data suggest that the Y. enterocolitica pYV plasmid may undergo a conformational change at the host temperature due to melting of DNA bends followed by compensatory adjustments in superhelical density. Hence, changes in DNA topology may be the temperature-sensing mechanism for virulence gene expression in Y. enterocolitica and other enteric pathogens.

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