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A novel histidine kinase inhibitor regulating development in Bacillus subtilis.

ABSTRACT: Kinase A is the sensor histidine kinase responsible for processing postexponential phase information and providing phosphate input to the phosphorelay that activates developmental transcription via phosphorylated Spo0A. A protein inhibitor, KipI, of kinase A was discovered encoded in an operon of genes of unknown function but regulated by the availability of fixed nitrogen. KipI is a potent inhibitor of the autophosphorylation reaction of kinase A but does not inhibit phosphate transfer to the Spo0F response regulator once kinase A is phosphorylated. KipI is an inhibitor of the catalytic domain of kinase A affecting the ATP/ADP reactions and not the phosphotransferase functions of this domain. The inhibitory activity of KipI is counteracted by the product of another gene in the operon, KipA. This protein may bind to KipI, preventing its function as an inhibitor of kinase A. KipI may be the first representative of a new class of signal transduction inhibitors that function by direct interaction with the catalytic domain of histidine kinases to counteract signals influencing the "sensor" domain of such kinases. This inhibitor represents yet another way by which the phosphorelay signal transduction system is affected by negative regulators under the control of metabolic, environmental, or cell cycle influences antithetical to the initiation of developmental transcription.

SUBMITTER: Wang L

PROVIDER: S-EPMC316564 | biostudies-literature | 1997 Oct

REPOSITORIES: biostudies-literature

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A novel histidine kinase inhibitor regulating development in Bacillus subtilis.

Wang L L Grau R R Perego M M Hoch J A JA

Genes & development 19971001 19

Kinase A is the sensor histidine kinase responsible for processing postexponential phase information and providing phosphate input to the phosphorelay that activates developmental transcription via phosphorylated Spo0A. A protein inhibitor, KipI, of kinase A was discovered encoded in an operon of genes of unknown function but regulated by the availability of fixed nitrogen. KipI is a potent inhibitor of the autophosphorylation reaction of kinase A but does not inhibit phosphate transfer to the S ...[more]

PMID: 9334321

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Project description:Environmental awareness is an essential attribute of all organisms. The homeoviscous adaptation system of Bacillus subtilis provides a powerful experimental model for the investigation of stimulus detection and signaling mechanisms at the molecular level. These bacteria sense the order of membrane lipids with the transmembrane (TM) protein DesK, which has an N-terminal sensor domain and an intracellular catalytic effector domain. DesK exhibits autokinase activity as well as phosphotransferase and phosphatase activities toward a cognate response regulator, DesR, that controls the expression of an enzyme that remodels membrane fluidity when the temperature drops below ∼30°C. Membrane fluidity signals are transmitted from the DesK sensor domain to the effector domain via rotational movements of a connecting 2-helix coiled coil (2-HCC). Previous molecular dynamic simulations suggested important roles for TM prolines in transducing the initial signals of membrane fluidity status to the 2-HCC. Here, we report that individual replacement of prolines in DesKs TM1 and TM5 helices by alanine (DesKPA) locked DesK in a phosphatase-ON state, abrogating membrane fluidity responses. An unbiased mutagenic screen identified the L174P replacement in the internal side of the repeated heptad of the 2-HCC structure that alleviated the signaling defects of every transmembrane DesKPA substitution. Moreover, substitutions by proline in other internal positions of the 2-HCC reestablished the kinase-ON state of the DesKPA mutants. These results imply that TM prolines are essential for finely tuned signal generation by the N-terminal sensor helices, facilitating a conformational control by the metastable 2-HCC domain of the DesK signaling state.IMPORTANCE Signal sensing and transduction is an essential biological process for cell adaptation and survival. Histidine kinases (HK) are the sensory proteins of two-component systems that control many bacterial responses to different stimuli, like environmental changes. Here, we focused on the HK DesK from Bacillus subtilis, a paradigmatic example of a transmembrane thermosensor suited to remodel membrane fluidity when the temperature drops below 30°C. DesK provides a tractable system for investigating the mechanism of transmembrane signaling, one of the majors interrogates in biology to date. Our studies demonstrate that transmembrane proline residues modulate the conformational switch of a 2-helix coiled-coil (2-HCC) structural motif that controls input-output in a variety of HK. Our results highlight the relevance of proline residues within sensor domains and could inspire investigations of their role in different signaling proteins.

Project description:In Bacillus subtilis, biofilm and sporulation pathways are both controlled by a master regulator, Spo0A, which is activated by phosphorylation via a phosphorelay-a cascade of phosphotransfer reactions commencing with autophosphorylation of histidine kinases KinA, KinB, KinC, KinD, and KinE. However, it is unclear how the kinases, despite acting via the same regulator, Spo0A, differentially regulate downstream pathways, i.e., how KinA mainly activates sporulation genes and KinC mainly activates biofilm genes. In this work, we found that KinC also downregulates sporulation genes, suggesting that KinC has a negative effect on Spo0A activity. To explain this effect, with a mathematical model of the phosphorelay, we revealed that unlike KinA, which always activates Spo0A, KinC has distinct effects on Spo0A at different growth stages: during fast growth, KinC acts as a phosphate source and activates Spo0A, whereas during slow growth, KinC becomes a phosphate sink and contributes to decreasing Spo0A activity. However, under these conditions, KinC can still increase the population-mean biofilm matrix production activity. In a population, individual cells grow at different rates, and KinC would increase the Spo0A activity in the fast-growing cells but reduce the Spo0A activity in the slow-growing cells. This mechanism reduces single-cell heterogeneity of Spo0A activity, thereby increasing the fraction of cells that activate biofilm matrix production. Thus, KinC activates biofilm formation by controlling the fraction of cells activating biofilm gene expression. IMPORTANCE In many bacterial and eukaryotic systems, multiple cell fate decisions are activated by a single master regulator. Typically, the activities of the regulators are controlled posttranslationally in response to different environmental stimuli. The mechanisms underlying the ability of these regulators to control multiple outcomes are not understood in many systems. By investigating the regulation of Bacillus subtilis master regulator Spo0A, we show that sensor kinases can use a novel mechanism to control cell fate decisions. By acting as a phosphate source or sink, kinases can interact with one another and provide accurate regulation of the phosphorylation level. Moreover, this mechanism affects the cell-to-cell heterogeneity of the transcription factor activity and eventually determines the fraction of different cell types in the population. These results demonstrate the importance of intercellular heterogeneity for understanding the effects of genetic perturbations on cell fate decisions. Such effects can be applicable to a wide range of cellular systems.

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A novel histidine kinase inhibitor regulating development in Bacillus subtilis.

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A novel histidine kinase inhibitor regulating development in Bacillus subtilis.

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