SummaryThe Kdp system of Escherichia coli is composed of the high-affinity K + transporter KdpFABC and the two regulatory proteins KdpD (sensor kinase) and KdpE (response regulator), which constitute a typical twocomponent system. The kdpFABC operon is induced under K + -limiting conditions and, to a lesser extent, under high osmolality in the medium. In search for the stimulus sensed by KdpD, we studied the inhibitory effect of extracellular K + on the Kdp system at pH 6.0, which is masked by unspecific K + transport at higher pH values. Based on KdpD derivatives carrying single aspartate replacements in the periplasmic loops which are part of the input domain, we concluded that the inhibition of the Kdp system at extracellular K + concentrations above 5 mM is mediated via KdpD/ KdpE and not due to inhibition of the K + -transporting KdpFABC complex. Furthermore, time-course analyses of kdpFABC expression revealed that a decline in the extracellular K + concentration efficiently stimulates KdpD/KdpE-mediated signal transduction. In this report we provide evidence that the extracellular K + concentration serves as one of the stimuli sensed by KdpD.

Two‐component systems (TCS) play a pivotal role for bacteria in stress regulation and adaptation. However, it is not well understood how these systems are modulated to meet bacterial demands. Especially, for those TCS using an accessory protein to integrate additional signals, no data concerning the role of the accessory proteins within the coordination of the response is available. The Cpx envelope stress two‐component system, composed of the sensor kinase CpxA and the response regulator CpxR, is orchestrated by the periplasmic protein CpxP which detects misfolded envelope proteins and inhibits the Cpx system in unstressed cells. Using selected reaction monitoring, we observed that the amount of CpxA and CpxR, as well as their stoichiometry, are only marginally affected, but that a 10‐fold excess of CpxP over CpxA is needed to switch off the Cpx system. Moreover, the relative quantification of the proteome identified not only acid stress response as a new indirect target of the Cpx system, but also suggests a general function of the Cpx system for cell wall stability.

BackgroundThe aminoglycoside antibiotic gentamicin was supposed to induce a crosstalk between the Cpx- and the Arc-two-component systems (TCS). Here, we investigated the physical interaction of the respective TCS components and compared the results with their respective gene expression and protein abundance. The findings were interpreted in relation to the global proteome profile upon gentamicin treatment.ResultsWe observed specific interaction between CpxA and ArcA upon treatment with the aminoglycoside gentamicin using Membrane-Strep-tagged protein interaction experiments (mSPINE). This interaction was neither accompanied by detectable phosphorylation of ArcA nor by activation of the Arc system via CpxA. Furthermore, no changes in absolute amounts of the Cpx- and Arc-TCS could be determined with the sensitive single reaction monitoring (SRM) in presence of gentamicin. Nevertheless, upon applying shotgun mass spectrometry analysis after treatment with gentamicin, we observed a reduction of ArcA ~ P-dependent protein synthesis and a significant Cpx-dependent alteration in the global proteome profile of E. coli.ConclusionsThis study points to the importance of the Cpx-TCS within the complex regulatory network in the E. coli response to aminoglycoside-caused stress.Electronic supplementary materialThe online version of this article (10.1186/s12866-017-1100-9) contains supplementary material, which is available to authorized users.