Clusters of bacterial RNA polymerase are biomolecular condensates that assemble through liquid-liquid phase separation

Proceedings of the National Academy of Sciences of the United States of America
Anne-Marie LadouceurStephanie C Weber

Abstract

Once described as mere "bags of enzymes," bacterial cells are in fact highly organized, with many macromolecules exhibiting nonuniform localization patterns. Yet the physical and biochemical mechanisms that govern this spatial heterogeneity remain largely unknown. Here, we identify liquid-liquid phase separation (LLPS) as a mechanism for organizing clusters of RNA polymerase (RNAP) in Escherichia coli Using fluorescence imaging, we show that RNAP quickly transitions from a dispersed to clustered localization pattern as cells enter log phase in nutrient-rich media. RNAP clusters are sensitive to hexanediol, a chemical that dissolves liquid-like compartments in eukaryotic cells. In addition, we find that the transcription antitermination factor NusA forms droplets in vitro and in vivo, suggesting that it may nucleate RNAP clusters. Finally, we use single-molecule tracking to characterize the dynamics of cluster components. Our results indicate that RNAP and NusA molecules move inside clusters, with mobilities faster than a DNA locus but slower than bulk diffusion through the nucleoid. We conclude that RNAP clusters are biomolecular condensates that assemble through LLPS. This work provides direct evidence for LLPS in bacteria and...Continue Reading

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Citations

Aug 23, 2020·International Journal of Molecular Sciences·Alain A M André, Evan Spruijt
Dec 10, 2020·Proceedings of the National Academy of Sciences of the United States of America·Grace A RosenJeff Gelles
Dec 11, 2020·Biochemical Society Transactions·Pinaki Swain, Stephanie C Weber
Jan 3, 2021·Journal of Molecular Biology·Bede Portz, James Shorter
Nov 14, 2020·Biophysical Journal·Christopher A AzaldeguiJulie S Biteen
Jan 17, 2021·Biophysical Journal·Ibraheem AlshareedahPriya R Banerjee
Feb 14, 2021·Biochimica Et Biophysica Acta. Molecular Cell Research·Miguel Ángel Robles-RamosBegoña Monterroso
Mar 20, 2021·Current Protocols·Rachel A VictorJacob C Schwartz
Mar 28, 2021·Trends in Cell Biology·Marina Feric, Tom Misteli
Apr 9, 2021·Frontiers in Microbiology·Vic Norris, Camille Ripoll
Apr 21, 2021·Current Opinion in Microbiology·Vidhyadhar Nandana, Jared M Schrader
Jun 2, 2021·EcoSal Plus·Maxime LerouxRodrigo Reyes-Lamothe
Jul 1, 2021·Cell Reports·Michael H Nichols, Victor G Corces
Jul 23, 2021·Biochimica Et Biophysica Acta. Molecular Cell Research·Semen V NesterovVladimir N Uversky
Jul 6, 2021·Molecular Microbiology·Josh McQuailSivaramesh Wigneshweraraj
Dec 22, 2020·ACS Nano·Anthony Birnie, Cees Dekker

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Methods Mentioned

BETA
Fluorescence
fluorescence microscopy
DNA array

Software Mentioned

SMAUG
IUPred

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