Oligomerization‐mediated phase separation in the nucleoid‐associated sensory protein H‐NS is controlled by ambient cues DOI Open Access

Bincy Lukose,

Saloni Goyal,

Athi N. Naganathan

et al.

Protein Science, Journal Year: 2024, Volume and Issue: 34(1)

Published: Dec. 11, 2024

H-NS, a nucleoid-associated protein (NAP) from enterobacteria, regulates gene expression by dynamically transducing environmental cues to conformational assembly and DNA binding. In this work, we show that H-NS Escherichia coli, which can assemble into octameric tetrameric oligomerization states, forms spontaneous micron-sized liquid-like condensates with at sub-physiological concentrations in vitro. The heterotypic are metastable 298 K, partially solubilizing time, while still retaining their properties. display UCST-like phase behavior higher temperatures, but large decrease droplet-assembly propensities 310 K ionic strength. Condensate formation be tuned cyclic manner between the extent of reversibility determined incubation highlighting strong hysteresis. An engineered phospho-mimetic variant (Y61E), is dimeric only weakly binds DNA, unable form condensates. Y61E mutant solubilizes pre-formed few minutes nearly an order magnitude speed-up droplet dissolution relative demonstrating rapid molecular transport dilute condensed phases. Our results establish intrinsically tied not binding also its phase-separation tendencies, showcasing regulatable programmable nature formed archetypal NAP via multiple lifetimes.

Language: Английский

SMC translocation is unaffected by an excess of nucleoid associated proteins in vivo DOI Creative Commons
Zhongqing Ren, Lindsey E. Way, Xindan Wang

et al.

Scientific Reports, Journal Year: 2025, Volume and Issue: 15(1)

Published: Jan. 19, 2025

Genome organization is important for DNA replication, gene expression, and chromosome segregation. In bacteria, two large families of proteins, nucleoid-associated proteins (NAPs) SMC complexes, play roles in organizing the genome. NAPs are highly abundant DNA-binding that can bend, wrap, bridge, compact DNA, while complexes load onto chromosome, translocate on extrude loops. Although capable traversing entire bound by various vivo, it unclear whether translocation influenced NAPs. this study, using Bacillus subtilis as a model system, we expressed collection representative bacterial archaeal introduce distinct structures potentially pose different challenges movement. By fluorescence microscopy chromatin immunoprecipitation, observed these to genome characteristic manners. Using genome-wide conformation capture (Hi-C) assays, found complex traversed without slowing down. Our findings revealed DNA-loop-extruding activity unaffected exogenously which highlights robustness motors busy chromatin.

Language: Английский

Citations

1
Future Directions of the Prokaryotic Chromosome Field DOI
Elio A. Abbondanzieri, Anjana Badrinarayanan, Daniela Barillà

et al.

Molecular Microbiology, Journal Year: 2025, Volume and Issue: 123(2), P. 89 - 100

Published: Feb. 1, 2025

In September 2023, the Biology and Physics of Prokaryotic Chromosomes meeting ran at Lorentz Center in Leiden, The Netherlands. As part workshop, those attendance developed a series discussion points centered around current challenges for field, how these might be addressed, field is likely to develop over next 10 years. staff facilitated discussions via tools aimed optimizing productive interactions. This Perspective article summary reflects state-of-the-art field. It expected help colleagues advancing their own research related prokaryotic chromosomes inspiring novel interdisciplinary collaborations. forward-looking perspective highlights open questions driving builds on impressive recent progress areas as represented by accompanying reviews, perspectives, articles this issue. These underline multi-disciplinary nature multiple length scales which chromatin studied vitro highlight differences similarities bacterial archaeal chromatin-associated processes.

Language: Английский

Citations

1
Antibacterial carbon dots DOI Creative Commons
Shuaishuai Wang, Dapeng Wang, Guoliang Wang

et al.

Materials Today Bio, Journal Year: 2024, Volume and Issue: 30, P. 101383 - 101383

Published: Dec. 6, 2024

Language: Английский

Citations

5
Regulation of Steady State Ribosomal Transcription inMycobacterium tuberculosis: Intersection of Sigma Subunits, Superhelicity, and Transcription Factors. DOI Creative Commons
Ana Ruiz Manzano, Drake Jensen, Eric A. Galburt

et al.

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2025, Volume and Issue: unknown

Published: Feb. 27, 2025

The regulation of ribosomal RNA (rRNA) is closely tied to nutrient availability, growth phase, and global gene expression, serving as a key factor in bacterial adaptability pathogenicity. Mycobacterium tuberculosis (Mtb ) stands out from other species with single operon controlled by two promoters: rrnA P3 P1 high ratio sigma (σ) factors genome size. While the primary σ A known drive transcription, alternative B has been proposed contribute transcription housekeeping genes, including rRNA under range conditions. However, 's precise role remains unclear. Here, we quantify steady-state rates reconstituted reactions establish that -mediated dominates production almost orders magnitude minimal contributions holoenzymes and/or all conditions tested. We measure compare kinetics individual initiation steps for both which, taken together rate measurements, lead us model where exhibit slower DNA unwinding holoenzyme recycling. Our data further demonstrate CarD RbpA reverse or buffer stimulatory effect negative superhelicity on respectively. Lastly, show major determinant increased activity due its N-terminal 205 amino acids. Taken together, our reveal intricate interplay promoter sequence, identity, superhelicity, shaping and, extension, Mtb .

Language: Английский

Citations

0
Biophysical modeling reveals the transcriptional regulatory mechanism of Spo0A, the master regulator in starving Bacillus subtilis DOI Creative Commons
Y.-H Zhang, Cristina S.D. Palma, Zhuo Chen

et al.

mSystems, Journal Year: 2025, Volume and Issue: unknown

Published: April 29, 2025

ABSTRACT In starving Bacillus subtilis bacteria , the initiation of two survival programs—biofilm formation and sporulation—is controlled by same phosphorylated master regulator, Spo0A~P. Its gene, spo0A, is transcribed from promoters, P v s, that are, respectively, regulated RNA polymerase (RNAP) holoenzymes bearing σ A H . Notably, transcription directly autoregulated Spo0A ~ binding sites known as 0A1, 0A2, 0A3 box, located in between promoters. It remains unclear whether, at onset starvation, these boxes activate or repress spo0A expression, whether transcriptional feedback plays a role increase expression. Based on experimental data promoter activities under systematic perturbation architecture, we developed biophysical model regulation Spo0A~P to each 0A boxes. The predicts its does not affect RNAP recruitment promoters but instead affects rate. Moreover, effects are mainly repressive saturated early starvation. Therefore, expression driven holoenzyme levels. Additionally, reveal affinity strongest weakest 0A2 there attractive forces occupied Our findings, addition clarifying how sporulation regulator controlled, offer framework predict regulatory outcomes complex gene-regulatory mechanisms. IMPORTANCE Cell differentiation often critical for survival. bacteria, decisions regulators control. understanding transcriptionally required understand differentiation. However, many cases, underlying complex, with multiple factor making it challenging dissect exact Here, address this problem Spo0A. Using model, quantitatively characterize effect individual promoter. Furthermore, allows us identify specific step affected binding. Such promising quantitative study wide range involved feedback.

Language: Английский

Citations

0
The pheS5-fit95 interface is essential for eliciting hallmark phenotype of fitA76 in Escherichia coli DOI

Praveen Belagal

Molecular Biology Reports, Journal Year: 2025, Volume and Issue: 52(1)

Published: April 30, 2025

Language: Английский

Citations

0
More than chromosome organizers: Unique mycobacterial nucleoid-associated proteins DOI

Kornel Milcarz,

Joanna Hołówka, Agnieszka Strzałka

et al.

Microbiological Research, Journal Year: 2025, Volume and Issue: unknown, P. 128239 - 128239

Published: May 1, 2025

Language: Английский

Citations

0
Identification of a novel alternate promoter element in the pheST operon of Escherichia coli DOI

Praveen Belagal

Molecular Biology Reports, Journal Year: 2024, Volume and Issue: 51(1)

Published: Oct. 17, 2024

Language: Английский

Citations

3
Towards a Better Control of Engineered Circuit Transcription in Bacterial Genomes DOI Creative Commons
Nazim Sarica, Laurent Jannière,

Brian C. Jester

et al.

Applied Microbiology, Journal Year: 2025, Volume and Issue: 5(1), P. 17 - 17

Published: Feb. 7, 2025

The transcription of genes and engineered circuits can deeply vary when inserted into different genomic loci. This unpredictable performance, termed context sensitivity, complicates strain development. Although the causes mechanisms sensitivity are emerging, it is poorly known how to engineer synthetic pathways isolated from it. Using tools biology for designing inserting various reporter cassettes in Escherichia coli genome RT-qPCR directly measuring gene transcription, we first surveyed landscape at 214 positions cells grown glucose or glycerol. results show deep variations cassette with respect position (up 160-fold) growth condition a 30-fold). We then demonstrated that this position-dependent variability strongly reduced insulated an artificial protein-bound DNA loop. Finally, measured two loop-insulated positions. depends on relative orientation genes, promoter strength, positive supercoiling. present model suggesting looping important cause be used better controlling circuits.

Language: Английский

Citations

0
Clues to transcription/replication collision‐induced DNA damage: it was RNAP, in the chromosome, with the fork DOI
Matthew Cooke, Christophe Herman, Priya Sivaramakrishnan

et al.

FEBS Letters, Journal Year: 2024, Volume and Issue: unknown

Published: Nov. 24, 2024

DNA replication and RNA transcription processes compete for the same template and, thus, frequently collide. These transcription–replication collisions are thought to lead genomic instability, which places a selective pressure on organisms avoid them. Here, we review predisposing causes, molecular mechanisms, downstream consequences of (TRCs) with strong emphasis prokaryotic model systems, before contrasting findings cases in eukaryotic systems. Current research points structure as primary determinant steady‐state TRC levels polymerase regulation inducer excess TRCs. We proposed mechanisms TRC‐induced damage, attempting clarify their mechanistic requirements. Finally, discuss what drives genomes select against

Language: Английский

Citations

2