Clusters of bacterial RNA polymerase are biomolecular condensates that assemble through liquid–liquid phase separation
Anne‐Marie Ladouceur, Baljyot Singh Parmar, Stefan Biedzinski, James Wall, S. Graydon Tope, David E. Cohn, Albright Kim, Nicolas Soubry, Rodrigo Reyes‐Lamothe, Stephanie C. Weber
Abstract
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.