The aim of our group is to understand the molecular basis of cellular differentiation, morphogenesis, and cell division in bacteria and to elucidate how one-dimensional genetic information is translated into defined spatial and temporal regulatory patterns. To address these questions, we are using a combination of cell biological, biochemical, structural, and bioinformatic approaches. Our studies focus primarily on the model organism Caulobacter crescentus, a Gram-negative bacterium that is characterized by its unique developmental cycle. In addition, we have extended our work to alternative model systems to facilitate comparative studies.
Intracellular protein gradients play a critical role in the spatial organization of both prokaryotic and eukaryotic cells, but in most cases the precise mechanisms underlying their formation are still unclear. We are investigating the function of a gradient-forming system, based on the conserved ATPase MipZ, that determines division site placement in the differentiating bacterium Caulobacter crescentus. MipZ interacts with a kinetochore-like nucleoprotein complex formed by the DNA partitioning protein ParB in proximity of the chromosomal origin of replication. Upon entry into S-phase, the two newly duplicated origin regions are segregated and tethered to opposite cell poles, giving rise to a bipolar distribution of MipZ, with a defined concentration minimum at the cell center. Since MipZ acts as an inhibitor of divisome formation, its gradient-like pattern effectively confines cytokinesis to the midcell region.
In collaboration with the group of Peter Lenz, we are using a combination of biochemistry, cell biology and computational modelling to investigate the mechanism driving formation of the MipZ gradient. We have solved the crystal structures of MipZ in the apo and ATP-bound state and started to dissect the determinants that ensures the dynamic recruitment of MipZ to the polar regions of the cell. Our results indicate that MipZ alternates between distinct conformational states that display marked differences in their interaction networks and diffusion rates. As a consequence, the protein undergoes an elaborate localization cycle, involving its oscillation between the polar ParB complexes and pole-distal regions of the nucleoid. The MipZ gradient thus represents the steady-state distribution of molecules in a highly dynamic system. Our work enlightens the principles that govern the establishment of a regulatory protein gradient within the confined space of the bacterial cytoplasm and provide the basis for the future implementation of gradient-forming systems in synthetic cells.