Bacterial cell division must occur at a specific site in the cell to ensure transfer of equal copies of the genome to each newborn cell. The first step in cell division is the polymerization of the widely conserved tubulin-like protein, FtsZ, at midcell to form a ring-shaped structure, called the Z-ring. The Z-ring recruits all other division proteins to this site to allow division. So mechanisms responsible for spatial control of division involve controlling the precise positioning of the Z-ring at midcell. Previously we have shown, using the Gram-positive bacterium, B. subtilis, that the ability to form Z-rings at midcell is correlated with the progression of the initiation phase of DNA replication. This suggests that the accuracy of Z-ring positioning is coupled with replication initiation, and has led to the hypothesis that a specific replication protein(s) has a role in Z-ring positioning. To test this, we correlated the stability of each DNA replication protein with the frequency of midcell Z-rings in a DNA replication initiation mutant. Interestingly, our results showed that one of these proteins, the DNA replication initiator, DnaA, was significantly unstable when a low frequency of midcell Z-rings was observed. This indicates DnaA as a regulator of Z-ring positioning. Consistent with this, we showed that a low frequency of midcell Z-rings occurs when DnaA is inactivated by (i) expression of a DnaA inhibitor protein, SirA; and (ii) deleting the dnaA gene by replacing it with a plasmid replicon. Results strikingly showed that midcell Z-ring frequency was decreased significantly under both conditions. This raises the intriguing possibility that DnaA has a role in the intracellular Z-ring positioning process. We are currently examining whether DnaA achieves this via a separate unidentified (non-replicative) function or through replication itself. Our data currently favour that a non-replicative mechanism involving changes to chromosome structure and/or organization around oriC.