Organization and replicon interactions within the highly segmented genome of Borrelia burgdorferi

Borrelia burgdorferi, a causative agent of Lyme disease, contains the most segmented bacterial genome known to date, with one linear chromosome and over twenty plasmids. How this unusually complex genome is organized, and whether and how the different replicons interact are unclear. We recently demonstrated that B. burgdorferi is polyploid and that the copies of the chromosome and plasmids are regularly spaced in each cell, which is critical for faithful segregation of the genome to daughter cells. Regular spacing of the chromosome is controlled by two separate partitioning systems that involve the protein pairs ParA/ParZ and ParB/Smc. Here, using chromosome conformation capture (Hi-C), we characterized the organization of the B. burgdorferi genome and the interactions between the replicons. We uncovered that although the linear chromosome lacks contacts between the two replication arms, the two telomeres are in frequent contact. Moreover, several plasmids specifically interact with the chromosome oriC region, and a subset of plasmids interact with each other more than with others. We found that Smc and the Smc-like MksB protein mediate long-range interactions on the chromosome, but they minimally affect plasmid-chromosome or plasmid-plasmid interactions. Finally, we found that disruption of the two partition systems leads to chromosome restructuring, correlating with the mis-positioning of chromosome oriC. Altogether, this study revealed the conformation of a complex genome and analyzed the contribution of the partition systems and SMC family proteins to this organization. This work expands the understanding of the organization and maintenance of multipartite bacterial genomes.Author summary: Genomes are highly organized in cells to facilitate biological processes. Borrelia burgdorferi, an agent of Lyme disease, carries one linear chromosome and more than twenty plasmids in what is known as one of the most segmented bacterial genomes. How the different replicons interact with each other is unclear. Here we investigate the organization of this highly segmented genome and the protein factors that contribute to this organization. Using chromosome conformation capture assays, we determined the interactions within the chromosome, between the chromosome and plasmids, and between the plasmids. We found that the two telomeres of the linear chromosome interact with each other; a subset of plasmids interact with the chromosomal replication origin region; and a subset of plasmids preferentially interact with one another. Finally, we revealed that two structural maintenance of chromosomes (SMC) family proteins, Smc and MksB, promote long-range DNA interactions on the chromosome, and the two partition systems, ParA/ParZ and ParB/Smc, contribute to chromosome structure. Altogether, we characterized the conformation of a complex genome and investigated the functions of different genome organizers. Our study advances the understanding of the organization of highly segmented bacterial genomes.