Genome segregation in bacteria: investigating protein dynamics and mechanisms in the cell at single molecule level

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Bacterial multidrug resistance is a global burden on human health worldwide.Large, low copy number plasmids responsible for antibiotic resistance haveevolved strategies to ensure their faithful distribution at celldivision. Multidrug resistance plasmids harbour their own survival system, apartition cassette, which ensures an accurate segregation of the plasmids atcell division. When this system malfunctions, the plasmid is not stablyinherited and is ultimately lost causing bacteria to become sensitive toantimicrobials. The multidrug resistance plasmid TP228 replicates at lowcopy number in Escherichia coli. Its partition cassette encodes two proteins:ParF, an ATPase, and ParG, a DNA-binding protein that associates to aspecific site on the plasmid. By using super-resolution microscopy on livecells, we have shown that ParG-plasmid complexes are entrapped within athree-dimensional ParF meshwork that assembles through the volume of thebacterial chromosome. When the ParG protein is defective, the ParG-plasmidcomplex is excluded from the chromosome volume and lost at the followingcell division. We have proposed a Venus flytrap model as a mechanism forplasmid segregation (1).This project will investigate the localization of ParFG-plasmid complexes inthe cell and the dynamics of complex formation at single molecule level toshed light on the mechanistic details underpinning plasmid segregation.