Department of Bionanoscience, Kavli Institute of Nanoscience Delft, Delft University of Technology, Van der Massweg 9, 2629HZ Delft, Netherlands.
BITZ Transformation Lab, Deggendorf Institute of Technology, 94363 Oberschneiding, Germany.
Nucleic Acids Res. 2024 Nov 27;52(21):13255-13268. doi: 10.1093/nar/gkae936.
The ParABS system plays a critical role in bacterial chromosome segregation. The key component of this system, ParB, loads and spreads along DNA to form a local protein-DNA condensate known as a partition complex. As bacterial chromosomes are heavily supercoiled due to the continuous action of RNA polymerases, topoisomerases and nucleoid-associated proteins, it is important to study the impact of DNA supercoiling on the ParB-DNA partition complex formation. Here, we use an in-vitro single-molecule assay to visualize ParB on supercoiled DNA. Unlike most DNA-binding proteins, individual ParB proteins are found to not pin plectonemes on supercoiled DNA, but freely diffuse along supercoiled DNA. We find that DNA supercoiling enhances ParB-DNA condensation, which initiates at lower ParB concentrations than on DNA that is torsionally relaxed. ParB proteins induce a DNA-protein condensate that strikingly absorbs all supercoiling writhe. Our findings provide mechanistic insights that have important implications for our understanding of bacterial chromosome organization and segregation.
ParABS 系统在细菌染色体分离中起着关键作用。该系统的关键组成部分 ParB 加载并沿 DNA 扩散，形成局部的蛋白质-DNA 凝聚物，称为分区复合物。由于 RNA 聚合酶、拓扑异构酶和类核相关蛋白的持续作用，细菌染色体严重超螺旋化，因此研究 DNA 超螺旋对 ParB-DNA 分区复合物形成的影响很重要。在这里，我们使用体外单分子测定法在超螺旋 DNA 上可视化 ParB。与大多数 DNA 结合蛋白不同，单个 ParB 蛋白不会在超螺旋 DNA 上固定扭结，而是自由扩散沿超螺旋 DNA。我们发现 DNA 超螺旋增强了 ParB-DNA 的凝聚，这比在扭转松弛的 DNA 上起始的 ParB 浓度更低。ParB 蛋白诱导的 DNA-蛋白质凝聚物显著吸收所有超螺旋扭曲。我们的发现提供了对细菌染色体组织和分离的理解具有重要意义的机制见解。
