“…It is well established that under optimal conditions CckA drives oscillations of the master cell cycle regulator CtrA through dynamically switching between its kinase and phosphatase activities [16,22,34,37]. Our new data suggest that environmental stress locks CckA in its phosphatase mode leading to the rapid inactivation of CtrA, its elimination through the protease ClpXP and consequently a block of CtrA regulated functions including cell division (Fig 8B).…”
Section: Discussionmentioning
confidence: 55%
“…Although our data show that CckA phosphatase activity is critical for the stress-dependent inactivation of CtrA, the detailed molecular process by which stress signals modulate CckA function remains to be elucidated. Our results rule out the involvement of the small signaling molecule c-di-GMP (Fig 7E and 7F), which promotes CckA phosphatase activity at the G1-to-S transition and promotes CtrA degradation by ClpXP under non-stress conditions [21,24,37]. Similarly, the stress-dependent regulation does not appear to be mediated by the upstream regulators DivL and DivK (Fig 7), which are critical for the cell cycle-dependent regulation of CckA in the absence of stress [22].…”
The bacterial cell cycle has been extensively studied under standard growth conditions. How it is modulated in response to environmental changes remains poorly understood. Here, we demonstrate that the freshwater bacterium Caulobacter crescentus blocks cell division and grows to filamentous cells in response to stress conditions affecting the cell membrane. Our data suggest that stress switches the membrane-bound cell cycle kinase CckA to its phosphatase mode, leading to the rapid dephosphorylation, inactivation and proteolysis of the master cell cycle regulator CtrA. The clearance of CtrA results in downregulation of division and morphogenesis genes and consequently a cell division block. Upon shift to non-stress conditions, cells quickly restart cell division and return to normal cell size. Our data indicate that the temporary inhibition of cell division through the regulated inactivation of CtrA constitutes a growth advantage under stress. Taken together, our work reveals a new mechanism that allows bacteria to alter their mode of proliferation in response to environmental cues by controlling the activity of a master cell cycle transcription factor. Furthermore, our results highlight the role of a bifunctional kinase in this process that integrates the cell cycle with environmental information.
“…It is well established that under optimal conditions CckA drives oscillations of the master cell cycle regulator CtrA through dynamically switching between its kinase and phosphatase activities [16,22,34,37]. Our new data suggest that environmental stress locks CckA in its phosphatase mode leading to the rapid inactivation of CtrA, its elimination through the protease ClpXP and consequently a block of CtrA regulated functions including cell division (Fig 8B).…”
Section: Discussionmentioning
confidence: 55%
“…Although our data show that CckA phosphatase activity is critical for the stress-dependent inactivation of CtrA, the detailed molecular process by which stress signals modulate CckA function remains to be elucidated. Our results rule out the involvement of the small signaling molecule c-di-GMP (Fig 7E and 7F), which promotes CckA phosphatase activity at the G1-to-S transition and promotes CtrA degradation by ClpXP under non-stress conditions [21,24,37]. Similarly, the stress-dependent regulation does not appear to be mediated by the upstream regulators DivL and DivK (Fig 7), which are critical for the cell cycle-dependent regulation of CckA in the absence of stress [22].…”
The bacterial cell cycle has been extensively studied under standard growth conditions. How it is modulated in response to environmental changes remains poorly understood. Here, we demonstrate that the freshwater bacterium Caulobacter crescentus blocks cell division and grows to filamentous cells in response to stress conditions affecting the cell membrane. Our data suggest that stress switches the membrane-bound cell cycle kinase CckA to its phosphatase mode, leading to the rapid dephosphorylation, inactivation and proteolysis of the master cell cycle regulator CtrA. The clearance of CtrA results in downregulation of division and morphogenesis genes and consequently a cell division block. Upon shift to non-stress conditions, cells quickly restart cell division and return to normal cell size. Our data indicate that the temporary inhibition of cell division through the regulated inactivation of CtrA constitutes a growth advantage under stress. Taken together, our work reveals a new mechanism that allows bacteria to alter their mode of proliferation in response to environmental cues by controlling the activity of a master cell cycle transcription factor. Furthermore, our results highlight the role of a bifunctional kinase in this process that integrates the cell cycle with environmental information.
“…Although sequence divergence makes the DHp rotational shift unlikely in HisKA HKs (Marina et al, 2005), DHp kinking by means of a conserved proline (Casino et al, 2014; Mechaly et al, 2014) could regulate the separation of the phosphorylatable His from the reaction center, analogously to the HisKA_3 rotational switch. Additional mechanisms that stabilize an ‘open’ state of HisKA HKs via extensive CA-DHp interfaces, have been reported to favor the phosphatase reaction (Dubey et al, 2016) further supporting our hypotheses.10.7554/eLife.21422.020Figure 6.Conceptual model of TCS unidirectional signaling.( A ) P~DesR dephosphorylation catalysis is favored by the opening of the RR phosphorylation lid, via Phe 82(RR) insertion into an HK hydrophobic pocket, and an ionic interaction between Asp 189(HK) and Arg 84(RR) . The Phe 82(RR) movement allows for entry of a nucleophilic water (yellow sphere) positioned by Gln 193(HK) .…”
Two-component systems (TCS) are protein machineries that enable cells to respond to input signals. Histidine kinases (HK) are the sensory component, transferring information toward downstream response regulators (RR). HKs transfer phosphoryl groups to their specific RRs, but also dephosphorylate them, overall ensuring proper signaling. The mechanisms by which HKs discriminate between such disparate directions, are yet unknown. We now disclose crystal structures of the HK:RR complex DesK:DesR from Bacillus subtilis, comprising snapshots of the phosphotransfer and the dephosphorylation reactions. The HK dictates the reactional outcome through conformational rearrangements that include the reactive histidine. The phosphotransfer center is asymmetric, poised for dissociative nucleophilic substitution. The structural bases of HK phosphatase/phosphotransferase control are uncovered, and the unexpected discovery of a dissociative reactional center, sheds light on the evolution of TCS phosphotransfer reversibility. Our findings should be applicable to a broad range of signaling systems and instrumental in synthetic TCS rewiring.DOI:
http://dx.doi.org/10.7554/eLife.21422.001
“…Interestingly, the interaction between the DHp and CA domains in the T116 structure differs from the interaction observed in all previously solved structures of histidine kinases . The CA domains are swapped between two protein chains (Figure C).…”
Section: Resultsmentioning
confidence: 57%
“…Apparently, this particular domain orientation observed in T116 could not be modeled using the template‐based techniques. Even so, the biological significance of this domain swap is unclear as it might well be a consequence of crystal contacts . Moreover, the orientation between CA and DHp domains in histidine kinases has been shown to be highly flexible (Figure B) and may depend on ligand binding, histidine phosphorylation states, and so forth .…”
We participated in Round 37 of the Critical Assessment of PRediction of Interactions (CAPRI), held jointly with the 12th edition of the Critical Assessment of protein Structure Prediction (CASP12), having two major objectives. First, we intended to test the utility of our PPI3D web server in finding and selecting templates for comparative modeling of structures of protein complexes. Our second aim was to evaluate the ability of our model accuracy estimation method VoroMQA to score and rank structural models for protein-protein interactions. Using sequence search in PPI3D and HHpred servers we identified multimeric templates for 7 of 11 CAPRI targets, and models of at least acceptable quality were constructed for 6 of them. The clustering and visual analysis features implemented in the PPI3D software were instrumental in detecting alternative protein-protein interaction interfaces among the identified templates. When a single binding mode was observed for homologous proteins, the structural modeling of the protein complex was fairly straightforward, whereas choosing the correct interaction template from several alternatives turned out to be a difficult task requiring manual intervention. The combination of full structure and interaction interface VoroMQA scores effectively ranked structural models of protein complexes and selected models of better quality from the CAPRI Scoring sets. The overall results show possible uses of PPI3D and VoroMQA in structural modeling of protein-protein interactions and suggest ways for further improvements of both methods.
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