期刊
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
卷 117, 期 31, 页码 18540-18549出版社
NATL ACAD SCIENCES
DOI: 10.1073/pnas.2005019117
关键词
RNA polymerase; liquid-liquid phase separation; biomolecular condensate; spatial organization; single-molecule tracking
资金
- NSF [DBI-0742708]
- Natural Sciences and Engineering Research Council of Canada [RGPIN-2017-04435]
- Fonds de Recherche du Quebec-Nature et Technologies [B3X-209443]
Once described as mere bags of enzymes, bacterial cells are in fact highly organized, with many macromolecules exhibiting non-uniform 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 demonstrates that this process can serve as a mechanism for intracellular organization in prokaryotes and eukaryotes alike.
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