A single-domain response regulator functions as integrating hub to coordinate general stress response and development in alpha-proteobacteria

Abstract

The alpha-proteobacterial general stress response is governed by a conserved partner-switching mechanism that is triggered by phosphorylation of the response regulator PhyR. In the model organism Caulobacter crescentus PhyR was proposed to be phosphorylated by the histidine kinase PhyK, but biochemical evidence in support of such a role of PhyK is missing. Here, we identify a single-domain response regulator, MrrA, that is essential for general stress response activation in C. crescentus. We demonstrate that PhyK does not function as a kinase but accepts phosphoryl groups from MrrA and passes them on to PhyR, thereby adopting the role of a histidine phosphotransferase. MrrA is phosphorylated by at least six histidine kinases that likely serve as stress sensors. MrrA also transfers phosphate to LovK, a histidine kinase involved in C. crescentus holdfast production and attachment, that also negatively regulates the general stress response. We show that LovK together with the response regulator LovR acts as a phosphate sink to redirect phosphate flux away from the PhyKR branch. In agreement with the biochemical data, a mrrA mutant is unable to activate the general stress response and shows a hyper-attachment phenotype, which is linked to decreased expression of the major holdfast inhibitory protein HfiA. We propose that MrrA serves as a central phosphorylation hub that coordinates the general stress response with C. crescentus development and other adaptive behaviors. The characteristic bow-tie architecture of this phosphorylation network with MrrA as the central knot may expedite the evolvability and species-specific niche adaptation of this group of bacteria.\n\nImportanceTwo-component systems (TCSs) consisting of a histidine kinase and a cognate response regulator are predominant signal transduction systems in bacteria. To avoid cross-talk, TCS are generally thought to be highly insulated from each other. However, this notion is based largely on studies of the HisKA subfamily of histidine kinases, while little information is available for the HWE and HisKA2 subfamilies. The latter have been implicated in the alpha-proteobacterial general stress response. Here, we show that in the model organism Caulobacter crescentus an atypical FATGUY-type single-domain response regulator (SDRR), MrrA, is highly promiscuous both in accepting and transferring phosphoryl groups from and to a number of up- and downstream kinases, challenging the current view of strictly insulated TCSs. Instead, we propose that FATGUY response regulators have evolved in alpha-proteobacteria to serve as central phosphorylation hubs to broadly sample information and distribute phosphoryl groups between the general stress response pathway and other TCSs, thereby coordinating multiple cellular behaviors. This signaling cascade includes presumable histidine kinases harboring intact catalytic and ATP-binding (CA) domains that, however, do not function as kinases, but instead have adopted a role as histidine phosphotransferases. Our work highlights a complex phosphorylation network in alpha-proteobacteria that coordinates the general stress response with changes in other cellular behaviors and development.