scholarly journals Composition of Transcription Machinery and Its Crosstalk with Nucleoid-Associated Proteins and Global Transcription Factors

Biomolecules ◽  
2021 ◽  
Vol 11 (7) ◽  
pp. 924
Author(s):  
Georgi Muskhelishvili ◽  
Patrick Sobetzko ◽  
Sanja Mehandziska ◽  
Andrew Travers
Keyword(s):  
Rna Polymerase ◽  
Spatial Organization ◽  
Physiological State ◽  
Central Element ◽  
Sigma Factors ◽  
Spatiotemporal Patterns ◽  
Temporal Expression ◽  
Dna Topoisomerases ◽  
Transcription Machinery ◽  
Associated Proteins

The coordination of bacterial genomic transcription involves an intricate network of interdependent genes encoding nucleoid-associated proteins (NAPs), DNA topoisomerases, RNA polymerase subunits and modulators of transcription machinery. The central element of this homeostatic regulatory system, integrating the information on cellular physiological state and producing a corresponding transcriptional response, is the multi-subunit RNA polymerase (RNAP) holoenzyme. In this review article, we argue that recent observations revealing DNA topoisomerases and metabolic enzymes associated with RNAP supramolecular complex support the notion of structural coupling between transcription machinery, DNA topology and cellular metabolism as a fundamental device coordinating the spatiotemporal genomic transcription. We analyse the impacts of various combinations of RNAP holoenzymes and global transcriptional regulators such as abundant NAPs, on genomic transcription from this viewpoint, monitoring the spatiotemporal patterns of couplons—overlapping subsets of the regulons of NAPs and RNAP sigma factors. We show that the temporal expression of regulons is by and large, correlated with that of cognate regulatory genes, whereas both the spatial organization and temporal expression of couplons is distinctly impacted by the regulons of NAPs and sigma factors. We propose that the coordination of the growth phase-dependent concentration gradients of global regulators with chromosome configurational dynamics determines the spatiotemporal patterns of genomic expression.

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 Mycobacterial HelD is a nucleic acids-clearing factor for RNA polymerase

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Tomáš Kouba ◽  
Tomáš Koval’ ◽  
Petra Sudzinová ◽  
Jiří Pospíšil ◽  
Barbora Brezovská ◽  
...  
Keyword(s):  
Nucleic Acids ◽  
Rna Polymerase ◽  
Active Site ◽  
Mycobacterium Smegmatis ◽  
Rna Synthesis ◽  
Environmental Changes ◽  
Transcription Elongation ◽  
Specific Interactions ◽  
Inactive State ◽  
Transcription Machinery

AbstractRNA synthesis is central to life, and RNA polymerase (RNAP) depends on accessory factors for recovery from stalled states and adaptation to environmental changes. Here, we investigated the mechanism by which a helicase-like factor HelD recycles RNAP. We report a cryo-EM structure of a complex between the Mycobacterium smegmatis RNAP and HelD. The crescent-shaped HelD simultaneously penetrates deep into two RNAP channels that are responsible for nucleic acids binding and substrate delivery to the active site, thereby locking RNAP in an inactive state. We show that HelD prevents non-specific interactions between RNAP and DNA and dissociates stalled transcription elongation complexes. The liberated RNAP can either stay dormant, sequestered by HelD, or upon HelD release, restart transcription. Our results provide insights into the architecture and regulation of the highly medically-relevant mycobacterial transcription machinery and define HelD as a clearing factor that releases RNAP from nonfunctional complexes with nucleic acids.

scholarly journals Live-cell imaging reveals the spatiotemporal organization of endogenous RNA polymerase II phosphorylation at a single gene

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Linda S. Forero-Quintero ◽  
William Raymond ◽  
Tetsuya Handa ◽  
Matthew N. Saxton ◽  
Tatsuya Morisaki ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Single Molecule ◽  
Computational Models ◽  
Spatial Organization ◽  
Fluorescent Antibody ◽  
Single Gene ◽  
Single Copy ◽  
Living Cells ◽  
Live Cell

AbstractThe carboxyl-terminal domain of RNA polymerase II (RNAP2) is phosphorylated during transcription in eukaryotic cells. While residue-specific phosphorylation has been mapped with exquisite spatial resolution along the 1D genome in a population of fixed cells using immunoprecipitation-based assays, the timing, kinetics, and spatial organization of phosphorylation along a single-copy gene have not yet been measured in living cells. Here, we achieve this by combining multi-color, single-molecule microscopy with fluorescent antibody-based probes that specifically bind to different phosphorylated forms of endogenous RNAP2 in living cells. Applying this methodology to a single-copy HIV-1 reporter gene provides live-cell evidence for heterogeneity in the distribution of RNAP2 along the length of the gene as well as Serine 5 phosphorylated RNAP2 clusters that remain separated in both space and time from nascent mRNA synthesis. Computational models determine that 5 to 40 RNAP2 cluster around the promoter during a typical transcriptional burst, with most phosphorylated at Serine 5 within 6 seconds of arrival and roughly half escaping the promoter in ~1.5 minutes. Taken together, our data provide live-cell support for the notion of efficient transcription clusters that transiently form around promoters and contain high concentrations of RNAP2 phosphorylated at Serine 5.

scholarly journals Initiation and elongation of polyribonucleotide chains on rat ventral-prostate chromatin transcribed by homologous ribonucleic acid polymerase B.

1977 ◽  
Vol 166 (2) ◽  
pp. 189-198 ◽  
Author(s):  
P Thomas ◽  
P Davies ◽  
K Griffiths
Keyword(s):  
Rna Polymerase ◽  
Rna Synthesis ◽  
Prostate Gland ◽  
Ventral Prostate ◽  
Control Of Gene Expression ◽  
Rat Ventral Prostate ◽  
Associated Proteins ◽  
Ribonucleoside Triphosphate ◽  
Androgenic Steroids ◽  
Transcriptional Process

The characteristics of initiation of RNA synthesis and the elongation of RNA chains on rat ventral-prostate chromatin by RNA polymerase B were investigated by two methods. 1. Initiation was carried out under low-salt conditions with three ribonucleoside triphosphates, and elongation was begun in the absence of reinitiation by the addition of the fourth ribonucleoside triphosphate and increasing the salt concentration. 2. Stable initiation complexes were formed by preincubation of enzyme with template at 37 degrees C, elongation was started by the addition of all four ribonucleoside triphosphates and reinitiation or spurious RNA synthesis was prevented by rifamycin AF/013. The latter method gave more reliable results. The dependence of those parameters on the androgenic status of the animal was studied. During the first 24h after castration, elongation was mainly affected, whereas after 72h a smaller number of initiation sites for RNA polymerase B on chromatin was evident. Considerable diurnal variations in the various parameters were observed. Changes in the relative concentrations of the chromatin-associated proteins were also observed after castration. In the rat ventral-prostate gland androgenic steroids may not only influence one stage of the transcriptional process, but may affect many factors involved in the control of gene expression.

scholarly journals Spatiotemporal patterns of synchronous heavy rainfall events in East Asia during the Baiu season

2021 ◽  
Author(s):  
Frederik Wolf ◽  
Ugur Ozturk ◽  
Kevin Cheung ◽  
Reik V. Donner
Keyword(s):  
System Dynamics ◽  
East Asia ◽  
Heavy Rainfall ◽  
Spatial Organization ◽  
Monsoon Season ◽  
Spatiotemporal Patterns ◽  
Extreme Weather Events ◽  
Earth System ◽  
Rainfall Events ◽  
Heavy Rainfall Events

<p>Investigating the synchrony and interdependency of heavy rainfall occurrences is crucial to understand the underlying physical mechanisms and reduce physical and economic damages by improved forecasting strategies. In this context, studies utilizing functional network representations have recently contributed to significant advances in the understanding and prediction of extreme weather events.</p><p>To thoroughly expand on previous works employing the latter framework to the East Asian Summer Monsoon (EASM) system, we focus here on changes in the spatial organization of synchronous heavy precipitation events across the monsoon season (April to August) by studying the temporal evolution of corresponding network characteristics in terms of a sliding window approach. Specifically, we utilize functional climate networks together with event coincidence analysis for identifying and characterizing synchronous activity from daily rainfall estimates with <span>a spatial resolution of 0.25° </span>between 1998 and 2018. Our results demonstrate that the formation of the Baiu front as a main feature of the EASM is reflected by a double-band structure of synchronous heavy rainfall with two centers north and south of the front. Although the two separated bands are strongly related to either low- or high-level winds which are commonly assumed to be independent, we provide evidence that it is rather their mutual interconnectivity that changes during the different phases of the EASM season in a characteristic way.</p><p>Our findings shed some new light on the interplay between tropical and extratropical factors controlling the EASM intraseasonal evolution, which could potentially help improving future forecasts of the Baiu onset in different regions of East Asia.</p><p> </p><p>Further details: F. Wolf, U. Ozturk, K. Cheung, R.V. Donner: Spatiotemporal patterns of synchronous heavy rainfall events in East Asia during the Baiu season. Earth System Dynamics (in review). Discussion Paper: Earth System Dynamics Discussions, (2020)</p>

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