Signal integration in bacterial two-component regulatory systems

Abstract

Two-component systems (TCSs) and phosphorelays are key mediators of bacterial signal transduction. The signals activating these systems promote the phosphorylated state of a response regulator, which is generally the form that carries out specific functions such as binding to DNA and catalysis of biochemical reactions. An emerging class of proteins-termed TCS connectors-modulate the output of TCSs by affecting the phosphorylation state of response regulators. TCS connectors use different mechanisms of action for signal integration, as well as in the coordination and fine-tuning of cellular processes. Present in both Gram-positive and Gram-negative bacteria, TCS connectors are critical for a variety of physiological functions including sporulation, competence, antibiotic resistance, and the transition to stationary phase.

Figure 1.

Schematics of the proteins and domains that constitute TCSs (left) and phosphorelays (right). The input domain of a sensor kinase responds to its signal by activating the autokinase domain, which autophosphorylates from ATP at a conserved histidine residue. The phosphorylated sensor kinase interacts with the receiver domain of the response regulator, which catalyzes the phosphoryl transfer to a conserved aspartate residue. Phosphorylation of the response regulator activates its output domain, which performs a specific biochemical function such as transcriptional regulation. A phosphorelay contains, besides the sensor kinase and the terminal response regulator, an intermediate response regulator lacking an output domain and a His-containing phosphotransfer protein. In some phosphorelays, the phosphotransfer protein and/or the intermediate response regulator is fused with the sensor kinase in a single polypeptide.

Figure 2.

TCS connectors can inhibit sensor autophosphorylation or promote dephosphorylation of phosphorylated response regulators. In the B. subtilis phosphorelay, the sensor kinase KinA is activated by an unknown signal, which results in autophosphorylation from ATP and subsequent phosphotransfer to the response regulator Spo0F. Spo0F passes on the phosphoryl group to the His-containing protein Spo0B, which in turn transfers it to the terminal acceptor, the response regulator Spo0A. The phosphorylated form of Spo0A acts as a transcription factor, being the key activator of sporulation genes. The connectors Sda and KipI block activation of the phosphorelay by inhibiting KinA autophosphorylation. The connectors RapA, RapB, RapE, and RapH promote dephosphorylation of the response regulator Spo0F-P; the connectors Spo0E, YisI, and YnzD act in a similar way on Spo0A-P. The expression of connectors is controlled by factors such as growth conditions, status of the DNA replication machinery, and development of competence (through the action of ComA and ComK—the key regulators of competence genes).

Figure 3.

TCS connectors can promote activation of response regulators and sensor kinases. (A) The connector-mediated pathway from S. enterica. The low Mg²⁺ signal activates the PhoP/PhoQ TCS, which triggers the expression of the connector PmrD. PmrD binds to the phosphorylated form of the response regulator PmrA, thereby protecting it from dephosphorylation. PmrA-P binds to DNA and regulates its target promoters. PmrA-P represses transcription of the pmrD gene, thus establishing a negative feedback loop controlling PmrD expression. The PmrA/PmrB TCS can be activated directly by the Fe³⁺ signal. (B) The B1500 protein from E. coli connects the EvgA/EvgS and PhoP/PhoQ TCSs. In response to an unknown signal, the EvgA/EvgS system triggers the expression of B1500, which interacts with the sensor kinase PhoQ, thereby promoting activation of PhoP and resulting in transcription of PhoP-activated genes.

Figure 4.

TCS connectors can inhibit binding of activated response regulators to DNA or prevent RNA polymerase from interacting with a response regulator. The ComA/ComP TCS from B. subtilis responds to the extracellular quorum-sensing signal, the peptide ComX. Upon activation, ComA promotes transcription of the srf operon, which leads to development of the competent state. Binding of ComA to DNA is inhibited by the connectors RapC, RapF, and RapH. These connectors, in turn, are deactivated upon binding to the corresponding Phr peptides. The action of ComA is also inhibited by the connector Spx, which disrupts the interaction between ComA and RNA polymerase.