High stimulus unmasks positive feedback in an autoregulated bacterial signaling circuit

Miyashiro, Tim; Goulian, Mark

doi:10.1073/pnas.0807278105

Cited by 76 publications

(156 citation statements)

References 37 publications

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“…For typical TCSs, as long as the total RR concentration remains within the saturating range, variations of RR concentration cause little change in RR phosphorylation. It appears that the activated wild-type expression levels of the PhoR/PhoB, PhoQ/PhoP, and EnvZ/OmpR systems are all within, or close to, the saturating range (11,12). For a different system, with proteins expressed below saturating concentrations, phosphorylation robustness may not be anticipated.…”

Section: Discussionmentioning

confidence: 99%

“…To fit the concentration-dependent phosphorylation saturation with minimal sets of parameters, we adopted a simple kinetic model that has been successful in describing the behaviors of other TCSs (11,12). The model uses two Michaelis-Menten kinetic reactions to describe PhoR-catalyzed phosphotransfer and dephosphorylation, respectively (Fig.…”

Section: Characterization Of In Vivo Phob Phosphorylation At Pi-depletedmentioning

confidence: 99%

“…It has been shown that the steady-state output of the Escherichia coli chemotaxis system is dependent on concentrations of chemotaxis proteins (10). Even for a few TCSs in which concentration robustness has been modeled and demonstrated, only within a certain range of concentrations do the output responses become relatively insensitive to TCS protein levels (4,11,12). For a wider range of protein levels, the steady-state level of phosphorylated RR (RR∼P) is described as a function of total RR concentrations defined by kinetic parameters unique to individual TCSs (11-13).…”

mentioning

confidence: 99%

“…Even for a few TCSs in which concentration robustness has been modeled and demonstrated, only within a certain range of concentrations do the output responses become relatively insensitive to TCS protein levels (4,11,12). For a wider range of protein levels, the steady-state level of phosphorylated RR (RR∼P) is described as a function of total RR concentrations defined by kinetic parameters unique to individual TCSs (11)(12)(13). Therefore, robust or not, the relationship of RR∼P and total RR amount still provides rich information for in vivo characterization of these unique kinetic parameters.…”

mentioning

confidence: 99%

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Probing kinase and phosphatase activities of two-component systems in vivo with concentration-dependent phosphorylation profiling

Gao¹,

Stock²

2012

Proc. Natl. Acad. Sci. U.S.A.

142

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Quantitative analyses of protein concentrations, modifications and activities in their native environments are playing an increasingly vital role in unraveling the general principles underlying signal transduction pathways. The prevalent bacterial two-component systems (TCSs) use a central phosphotransfer for signaling; however, in vivo characterization of the kinase and phosphatase activities of TCS proteins is often limited by traditional transcriptional reporter assays and complicated by simultaneous actions of multiple TCS activities. Here, we report a strategy that combines concentration-dependent phosphorylation profiling and mathematical modeling to characterize the cellular activities of the archetype Escherichia coli PhoR/PhoB system. Phosphorylation of the response regulator (RR) PhoB has been found to be dependent on the total concentrations of PhoB/PhoR and saturated at high concentrations. The relationship between RR phosphorylation and total concentrations has been defined by the modeling of the kinase and phosphatase reactions and quantified to derive the biochemical parameters of the PhoR/PhoB system in vivo. In a further test of this approach on a PhoB mutant, PhoB , it proved highly effective in exploring the mechanistic differences of TCSs that are not revealed by traditional reporter assays. Measurement of biochemical parameters for PhoB F20D led to the discovery that a weaker interaction between the histidine sensor kinase and RR could result in a higher and nonrobust phosphorylation due to diminished phosphatase activities.in vivo phosphorylation | two-component signal transduction T wo-component systems (TCSs), one of the predominant signaling schemes in bacteria, connect input stimuli and output responses with a core phosphotransfer between a histidine sensor kinase (HSK) and a cognate response regulator (RR) (1, 2). The signaling pathway is often simply described as a series of steps that include autophosphorylation of HSKs, phosphotransfer to cognate RRs, and output modulation, usually via transcription regulation, mediated by phosphorylated RRs. Far from a simple on/off switch, phosphorylation levels of many TCS proteins are under sophisticated control by multiple enzymatic activities. One of the fundamental questions in TCS studies is what percentage of protein molecules are phosphorylated in the presence or absence of the stimuli, but the exact phosphorylation levels have not been well quantified in vivo. Without the quantification of phosphorylation, it is extremely difficult to characterize TCS kinase and phosphatase activities in their native cellular environments. This has been identified as one of the key questions outstanding in TCS research (3, 4).RR phosphorylation levels are commonly inferred from transcriptional reporter activities even though gene transcription is downstream of RR phosphorylation and often complicated by additional regulatory factors. A change in transcription could result from alteration of RR phosphorylation levels as well as effects of additional uniden...

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Section: Discussionmentioning

confidence: 99%

Section: Characterization Of In Vivo Phob Phosphorylation At Pi-depletedmentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

See 2 more Smart Citations

Probing kinase and phosphatase activities of two-component systems in vivo with concentration-dependent phosphorylation profiling

Gao¹,

Stock²

2012

Proc. Natl. Acad. Sci. U.S.A.

142

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“…For most two-component pathways, abundance of the response regulator likely exceeds that of the cognate kinase. The well-characterized E. coli kinase EnvZ and its partner OmpR are found at a ratio of about ~1:35, and other pathways are reported to have similar ratios [22,23]. The higher abundance of the response regulators creates a scenario in which a given regulator effectively outcompetes non-cognate regulators for binding to a cognate kinase, further preventing unwanted phosphotransfer events.…”

Section: Mechanisms Ensuring Specificity In Two-component Signaling Pmentioning

confidence: 99%

Determinants of specificity in two-component signal transduction

Podgornaia

Laub

2013

Current Opinion in Microbiology

128

133

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Maintaining the faithful flow of information through signal transduction pathways is critical to the survival and proliferation of organisms. This problem is particularly challenging as many signaling proteins are part of large, paralogous families that are highly similar at the sequence and structural levels, increasing the risk of unwanted cross-talk. To detect environmental signals and process information, bacteria rely heavily on two-component signaling systems comprised of sensor histidine kinases and their cognate response regulators. Although most species encode dozens of these signaling pathways, there is relatively little cross-talk, indicating that individual pathways are well insulated and highly specific. Here, we review the molecular mechanisms that enforce this specificity. Further, we highlight recent studies that have revealed how these mechanisms evolve to accommodate the introduction of new pathways by gene duplication.3

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References

2020

Structure and Function of the Bacterial Genome

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High stimulus unmasks positive feedback in an autoregulated bacterial signaling circuit

Cited by 76 publications

References 37 publications

Probing kinase and phosphatase activities of two-component systems in vivo with concentration-dependent phosphorylation profiling

Probing kinase and phosphatase activities of two-component systems in vivo with concentration-dependent phosphorylation profiling

Determinants of specificity in two-component signal transduction

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