Dataset Information

Complex two-component signaling regulates the general stress response in Alphaproteobacteria.

ABSTRACT: The general stress response (GSR) in Alphaproteobacteria was recently shown to be controlled by a partner-switching mechanism that is triggered by phosphorylation of the response regulator PhyR. Activation of PhyR ultimately results in release of the alternative extracytoplasmic function sigma factor ?(EcfG), which redirects transcription toward the GSR. Little is known about the signal transduction pathway(s) controlling PhyR phosphorylation. Here, we identified the single-domain response regulator (SDRR) SdrG and seven histidine kinases, PakA to PakG, belonging to the HWE/HisKA2 family as positive modulators of the GSR in Sphingomonas melonis Fr1. Phenotypic analyses, epistasis experiments, and in vitro phosphorylation assays indicate that Paks directly phosphorylate PhyR and SdrG, and that SdrG acts upstream of or in concert with PhyR, modulating its activity in a nonlinear pathway. Furthermore, we found that additional SDRRs negatively affect the GSR in a way that strictly requires PhyR and SdrG. Finally, analysis of GSR activation by thermal, osmotic, and oxidative stress indicates that Paks display different degrees of redundancy and that a specific kinase can sense multiple stresses, suggesting that the GSR senses a particular condition as a combination of, rather than individual, molecular cues. This study thus establishes the alphaproteobacterial GSR as a complex and interlinked network of two-component systems, in which multiple histidine kinases converge to PhyR, the phosphorylation of which is, in addition, subject to regulation by several SDRRs. Our finding that most HWE/HisKA2 kinases contribute to the GSR in S. melonis Fr1 opens the possibility that this notion might also be true for other Alphaproteobacteria.

SUBMITTER: Kaczmarczyk A

PROVIDER: S-EPMC4260549 | biostudies-literature | 2014 Dec

REPOSITORIES: biostudies-literature

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Complex two-component signaling regulates the general stress response in Alphaproteobacteria.

Kaczmarczyk Andreas A Hochstrasser Ramon R Vorholt Julia A JA Francez-Charlot Anne A

Proceedings of the National Academy of Sciences of the United States of America 20141117 48

The general stress response (GSR) in Alphaproteobacteria was recently shown to be controlled by a partner-switching mechanism that is triggered by phosphorylation of the response regulator PhyR. Activation of PhyR ultimately results in release of the alternative extracytoplasmic function sigma factor σ(EcfG), which redirects transcription toward the GSR. Little is known about the signal transduction pathway(s) controlling PhyR phosphorylation. Here, we identified the single-domain response regul ...[more]

PMID: 25404331

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Project description:Two common signal transduction mechanisms used by bacteria to sense and respond to changing environments are two-component systems (TCSs) and eukaryote-like Ser/Thr kinases and phosphatases (eSTK/Ps). Enterococcus faecalis is a Gram-positive bacterium and a serious opportunistic pathogen that relies on both a TCS and an eSTK/P pathway for intrinsic resistance to cell wall-targeting antibiotics. The TCS consists of a histidine kinase (CroS) and a response regulator (CroR) that become activated upon exposure of cells to cell wall-targeting antibiotics, leading to a modulation of gene expression. The eSTK/P pathway consists of a transmembrane kinase (IreK) and its cognate phosphatase (IreP), which act antagonistically to mediate antibiotic resistance through an unknown mechanism. Because both CroS/R and IreK/P contribute to enterococcal resistance toward cell wall-targeting antibiotics, we hypothesized that these signaling systems are intertwined. To test this hypothesis, we analyzed CroR phosphorylation and CroS/R-dependent gene expression to probe the influence of IreK and IreP on CroS/R signaling. In addition, we analyzed the phosphorylation state of CroS, which revealed the IreK-dependent phosphorylation of a Thr residue important for CroS function. Our results are consistent with a model in which IreK positively influences CroR-dependent gene expression through the phosphorylation of CroS to promote antimicrobial resistance in E. faecalis IMPORTANCE Two-component signaling systems (TCSs) and eukaryote-like Ser/Thr kinases (eSTKs) are used by bacteria to sense and adapt to changing environments. Understanding how these pathways are regulated to promote bacterial survival is critical for a more complete understanding of bacterial stress responses and physiology. The opportunistic pathogen Enterococcus faecalis relies on both a TCS (CroS/R) and an eSTK (IreK) for intrinsic resistance to cell wall-targeting antibiotics. We probed the relationship between CroS/R and IreK, revealing the convergence of IreK and the sensor kinase CroS to enhance signaling through CroS/R and increase antimicrobial resistance in E. faecalis This newly described example of eSTK/TCS convergence adds to our understanding of the signaling networks mediating antimicrobial resistance in E. faecalis.

Project description:For successful infection to occur, a pathogen must be able to evade or tolerate the host's defense systems. This requires the pathogen to first recognize the host environment and then signal this response to elicit a complex adaptive program in order to activate its own defense strategies. In both prokaryotes and eukaryotes, two-component signaling systems are utilized to sense and respond to changes in the external environment. The hybrid histidine kinases (HHKs) at the start of the two-component signaling pathway have been well characterized in human pathogens. However, how these HHKs regulate processes downstream currently remains unclear. This study describes the role of a response regulator downstream of these HHKs, sskA, in Talaromyces marneffei, a dimorphic human pathogen. sskA is required for asexual reproduction, hyphal morphogenesis, cell wall integrity, osmotic adaptation, and the morphogenesis of yeast cells both in vitro at 37°C and during macrophage infection, but not during dimorphic switching. Comparison of the ?sskA mutant with a strain in which the mitogen-activated protein kinase (MAPK) of the high-osmolarity glycerol pathway (SakA) has been deleted suggests that SskA acts upstream of this pathway in T. marneffei to regulate these morphogenetic processes. This was confirmed by assessing the amount of phosphorylated SakA in the ?sskA mutant, antifungal resistance due to a lack of SakA activation, and the ability of a constitutively active sakA allele (sakA(F316L) ) to suppress the ?sskA mutant phenotypes. We conclude that SskA regulates morphogenesis and osmotic stress adaptation in T. marneffei via phosphorylation of the SakA MAPK of the high-osmolarity glycerol pathway. IMPORTANCE This is the first study in a dimorphic fungal pathogen to investigate the role of a response regulator downstream of two-component signaling systems and its connection to the high-osmolarity glycerol pathway. This study will inspire further research into the downstream components of two-component signaling systems and their role during pathogenic growth.

Dataset Information

Complex two-component signaling regulates the general stress response in Alphaproteobacteria.

Publications

Complex two-component signaling regulates the general stress response in Alphaproteobacteria.

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