Molecular Mechanism of Chromosome Segregation by the E. coli Min system

Cell division is one of the most essential processes underlying life. Of importance is certainly the equal partitioning of the genetic material into the two daughter cells termed chromosome segregation. It is still debated if entropic forces alone are sufficient to fulfill this task, or whether additional dedicated protein machineries play an active role in this process. The E. coli Min system, consisting of the proteins MinC, MinD and MinE, is well known for its function in defining mid-cell and directing there the FtsZ-ring, which marks the position of the future division site. The Min system is characterized by a pole-to-pole oscillation, corresponding to a time-averaged bipolar protein distribution. It has been discovered in our lab that MinD is able to directly bind DNA in vitro and in vivo. This led to the proposal of a Brownian Ratchet-like model for chromosome segregation, in which membrane-bound MinD provides DNA tethering sites and biases the diffusion of the duplicated chromosomes in the direction of the poles. However, the molecular details of this mechanism were still lacking. Here I used several in vitro and in vivo assays to understand better how MinD binds to the DNA and to clarify what the role of other proteins, such as MinC, MinE and FtsZ, is. Specifically, I performed ChIP-Seq to study the genome-wide binding of MinD in cells with and without the endogenous Min system and found that MinD does not associate to specific chromosomal macrodomains. This supports the notion that the Min system assists the segregation of the chromosomal bulk. Furthermore, analysis of the transcriptome revealed that the Min system does not function as a global transcriptional regulator. Using electrophoretic mobility shift assays (EMSAs) with purified proteins, I established that the MinC-MinD complex has a much higher affinity for DNA than the individual proteins. Lipid vesicles coated with MinC-MinD complexes could tether plasmid DNA to the lipid-associated pellet in co-sedimentation assays. These findings advocate ...