Cell differentiation and division in Caulobacter crescentus are regulated by a signal transduction pathway mediated by the histidine kinase DivJ and the essential response regulator DivK. Here we report genetic and biochemical evidence that the DivJ and DivK proteins function to control the activity of CtrA, a response regulator required for multiple cell cycle events, including f lagellum biosynthesis, DNA replication, and cell division. Temperature-sensitive sokA (suppressor of divK) alleles were isolated as extragenic suppressors of a cold-sensitive divK mutation and mapped to the C terminus of the CtrA protein. The sokA alleles also suppress the lethal phenotype of a divK gene disruption and the cold-sensitive cell division phenotype of divJ mutants. The relationship between these signal transduction components and their target was further defined by demonstrating that the purified DivJ kinase phosphorylates CtrA, as well as DivK. Our studies also showed that phospho-CtrA activates transcription in vitro from the class II f lagellar genes and that their promoters are recognized by the principal C. crescentus sigma factor 73 . We propose that an essential signal transduction pathway mediated by DivJ, DivK, and CtrA coordinates cell cycle and developmental events in C. crescentus by regulating the level of CtrA phosphorylation and transcription from 73 -dependent class II gene promoters. Our results suggest that an unidentified phosphotransfer protein or kinase (X) is responsible for phosphoryl group transfer to CtrA in the proposed DivJ f DivK f X f CtrA phosphorelay pathway.Bacterial two-component signal transduction systems control a wide array of physiological processes in response to a variety of environmental conditions. These systems typically contain a sensor kinase, which is autophosphorylated on a conserved histidine residue, and a cognate response regulator containing a conserved aspartate residue to which the phosphoryl group is transferred (1, 2). This same protein family functions in multistep phosphorelay pathways (3). Recent results have provided evidence that sensor kinases and response regulators also play essential roles in the coordination of cell cycle and developmental events in the aquatic bacterium Caulobacter crescentus (4). The histidine kinase DivJ (5) and response regulator DivK (6) have been implicated in a signal transduction pathway required for cell division initiation. Another response regulator, CtrA, has been identified as a transcription factor responsible for the expression of multiple cell cycleregulated genes (7). Our results now indicate that DivJ and DivK proteins function as part of a multicomponent signal transduction pathway to control the transcriptional activity of CtrA during the cell cycle.Members of two-component signal transduction pathways regulating cell cycle events in C. crescentus were originally identified in a pseudoreversion analysis of pleC, a pleiotropic developmental gene required for motility and polar morphogenesis (8). Several of the pleC...