Integrative and quantitative view of the CtrA regulatory network in a stalked budding bacterium

The Alphaproteobacteria show a remarkable diversity of cell cycle-dependent developmental patterns, which are governed by the conserved CtrA pathway. Its central component CtrA is a DNA-binding response regulator that is controlled by a complex two-component signaling network, mediating distinct transcriptional programs in the two offspring. The CtrA pathway has been studied intensively and was shown to consist of an upstream part that reads out the developmental state of the cell and a downstream part that integrates the upstream signals and mediates CtrA phosphorylation. However, the role of this circuitry in bacterial diversification remains incompletely understood. We have therefore investigated CtrA regulation in the morphologically complex stalked budding alphaproteobacterium Hyphomonas neptunium. Compared to relatives dividing by binary fission, H. neptunium shows distinct changes in the role and regulation of various pathway components. Most notably, the response regulator DivK, which normally links the upstream and downstream parts of the CtrA pathway, is dispensable, while downstream components such as the pseudokinase DivL, the histidine kinase CckA, the phosphotransferase ChpT and CtrA are essential. Moreover, CckA is compartmentalized to the nascent bud without forming distinct polar complexes and CtrA is not regulated at the level of protein abundance. We show that the downstream pathway controls critical functions such as replication initiation, cell division and motility. Quantification of the signal flow through different nodes of the regulatory cascade revealed that the CtrA pathway is a leaky pipeline and must involve thus-far unidentified factors. Collectively, the quantitative system-level analysis of CtrA regulation in H. neptunium points to a considerable evolutionary plasticity of cell cycle regulation in alphaproteobacteria and leads to hypotheses that may also hold in well-established model organisms such as Caulobacter crescentus.