RitR is an archetype for a novel family of redox sensors in the streptococci that has evolved from two-component response regulators and is required for pneumococcal colonization

Abstract

To survive diverse host environments, the human pathogen Streptococcus pneumoniae must prevent its self-produced, extremely high levels of peroxide from reacting with intracellular iron. However, the regulatory mechanism(s) by which the pneumococcus accomplishes this balance remains largely enigmatic, as this pathogen and other related streptococci lack all known redox-sensing transcription factors. Here we describe a two-component-derived response regulator, RitR, as the archetype for a novel family of redox sensors in a subset of streptococcal species. We show that RitR works to both repress iron transport and enable nasopharyngeal colonization through a mechanism that exploits a single cysteine (Cys128) redox switch located within its linker domain. Biochemical experiments and phylogenetics reveal that RitR has diverged from the canonical two-component virulence regulator CovR to instead dimerize and bind DNA only upon Cys128 oxidation in air-rich environments. Atomic structures show that Cys128 oxidation initiates a “helical unravelling” of the RitR linker region, suggesting a mechanism by which the DNA-binding domain is then released to interact with its cognate regulatory DNA. Expanded computational studies indicate this mechanism could be shared by many microbial species outside the streptococcus genus.

A High-Throughput Method for Identifying Novel Genes That Influence Metabolic Pathways Reveals New Iron and Heme Regulation in Pseudomonas aeruginosa

“…Expression and purification of PA0709 and PA3390 were carried out as described in reference 147 . His 6 -tagged PA0709 and PA3390 proteins were expressed in E. coli T7 Express lysY i/q cells (New England BioLabs Inc., Ipswich, MA) carrying pET15DG1-PA0709 or pET15DG1-PA3390.…”

TprA/PhrA Quorum Sensing System Has a Major Effect on Pneumococcal Survival in Respiratory Tract and Blood, and Its Activity Is Controlled by CcpA and GlnR

“…Female, 8–10 week old MF1 mice weighing ~30–35 g (Charles River, UK), were anesthetized with 2.5% v/v isoflurane (Astra Zeneca, Macclesfield, UK) over oxygen (2–2.5 liter/min). For the pneumonia model, ~1.5 × 10 6 pneumococci, in 50 μL PBS (Gaspar et al, 2014; Glanville et al, 2018) were administered dropwise into the nostrils of the anesthetized mouse, held vertically. After infections, CFU were determined to ensure that the intended number of pneumococci had been administered.…”

Regulation of pneumococcal epigenetic and colony phases by multiple two-component regulatory systems

“…TCS02 (WalR/K), the only TCS essential for pneumococcal viability, is associated with cell wall peptidoglycan metabolism [28,29]; TCS03 (LiaR/S) senses cell wall damage and inhibits autolysis [30], TCS04 (PnpR/S) regulates uptake of inorganic phosphate [31,32]; TCS06 (CbpR/S) activates expression of choline-binding protein A or PspC [33,34]; TCS08 is involved in the cellobiose uptake [35], production of pilus-1 [16] and surface protein PavB [16]; TCS13 (BlpR/H) regulates the production of bacteriocins (pneumocins) [36][37][38]. The orphan response regulator RR14 represses the transcription of the pneumococcal iron uptake (Piu) and other nutrient uptake loci in response to oxidative stress [39,40]. The cellular functions of the remaining five TCSs (1, 7, 9, 10 and 11) are poorly characterized.…”

“…RR14 represses the transcription of the pneumococcal iron uptake (Piu) and nutrient uptake loci [39]. A recent study has shown that RR14 serves as a principal regulator in response to oxidative stress via the cysteine-mediated dimerization, which enhances pneumococcal tolerance to hydrogen peroxide and other oxidants [40]. S. pneumoniae is famous for its production of millimolar range of hydrogen peroxide as a metabolic byproduct of pyruvate oxidase SpxB [67,68].…”