The Sinorhizobium meliloti sensor histidine kinase CbrA contributes to free-living cell cycle regulation

Abstract

Sinorhizobium meliloti is alternately capable of colonizing the soil as a free-living bacterium or establishing a chronic intracellular infection with its legume host for the purpose of nitrogen fixation. We previously identified the S. meliloti two-component sensor histidine kinase CbrA as playing an important role in regulating exopolysaccharide production, flagellar motility and symbiosis. Phylogenetic analysis of CbrA has highlighted its evolutionary relatedness to the Caulobacter crescentus sensor histidine kinases PleC and DivJ, which are involved in CtrAdependent cell cycle regulation through the shared response regulator DivK. We therefore became interested in testing whether CbrA plays a role in regulating S. meliloti cell cycle processes. We find the loss of cbrA results in filamentous cell growth accompanied by cells that contain an aberrant genome complement, indicating CbrA plays a role in regulating cell division and possibly DNA segregation. S. meliloti DivK localizes to the old cell pole during distinct phases of the cell cycle in a phosphorylation-dependent manner. Loss of cbrA results in a significantly decreased rate of DivK polar localization when compared with the wild-type, suggesting CbrA helps regulate cell cycle processes by modulating DivK phosphorylation status as a kinase. Consistent with a presumptive decrease in DivK phosphorylation and activity, we also find the steady-state level of CtrA increased in cbrA mutants. Our data therefore demonstrate that CbrA contributes to free-living cell cycle regulation, which in light of its requirement for symbiosis, points to the potential importance of cell cycle regulation for establishing an effective host interaction.

Sinorhizobium meliloti CtrA Stability Is Regulated in a CbrA-Dependent Manner That Is Influenced by CpdR1

“…Previous studies observed increased levels of CtrA in cbrA mutants (9,10). Here we show that the increased level of CtrA in a ⌬cbrA mutant is due, in part, to a significant increase in its stability.…”

“…1). For example, the DivJ/PleC homolog CbrA is necessary for proper cell cycle progression such that a null mutant displays aberrant morphologies indicative of cell division defects (9,42). CbrA promotes DivK phosphorylation, and this is likely the reason for increased CtrA levels and phosphorylation in cbrA mutants (9,10).…”

“…Known and putative S. meliloti cell cycle regulators are conserved among other members of the Alphaproteobacteria that also specialize in chronic colonization of eukaryotic hosts, in particular rhizobial species in the genera Agrobacterium, Bartonella, and Brucella (2, 4). Thus, a deeper understanding of S. meliloti cell cycle regulation will provide insight into processes that are broadly important to both cell cycle progression and host-microbe interaction (5,6,7,8,9,10,11,12).…”

Global analysis of cell cycle gene expression of the legume symbiontSinorhizobium meliloti

“…The transcription of these factors (which include ctrA, pleC, divK, divJ, cpdR, chpT, and sciP) was mostly upregulated coincident with the timing of their prescribed function in C. crescentus. This observation, along with previous molecular studies, strongly suggests that the basic functions of these cell cycle proteins are well conserved between S. meliloti and C. crescentus (16,24,25,40). Interestingly, many of the S. meliloti genes with cell cycle-regulated transcripts that are either not conserved or not cell cycle regulated in C. crescentus, are required for the symbiotic relationship between S. meliloti and its legume host-including the repABC genes controlling megaplasmid replication and maintenance, as well as genes involved in cyclic glucan production.…”

“…For example, it has been shown that altering the expression of genes central to S. meliloti cell cycle processes (i.e., ftsZ, dnaA, minE, and ccrM) produces bacteroid-like polyploid cells (10)(11)(12)(13) and that mutation of conserved cell cycle regulators (cbrA, cpdR1, and divJ) blocks bacteroid formation and symbiosis (14)(15)(16). Furthermore, in the α-proteobacterium Caulobacter crescentus, the cellular differentiation program governing morphological and replicative asymmetry in progeny cells is genetically integrated with the cell cycle (17).…”