A large family of anti‐activators accompanying XylS/AraC family regulatory proteins

Abstract

Summary AraC Negative Regulators (ANR) suppress virulence genes by directly down-regulating AraC/XylS members in Gram-negative bacteria. In this study we sought to investigate the distribution and molecular mechanisms of regulatory function for ANRs among different bacterial pathogens. We identified more than 200 ANRs distributed in diverse clinically important gram negative pathogens, including Vibrio spp., Salmonella spp., Shigella spp., Yersinia spp., Citrobacter spp., enterotoxigenic (ETEC) and enteroaggregative E. coli (EAEC), and members of the Pasteurellaceae. By employing a bacterial two hybrid system, pull down assays, and surface plasmon resonance (SPR) analysis, we demonstrate that Aar (AggR-activated regulator), a prototype member of the ANR family in EAEC, binds with high affinity to the central linker domain of AraC-like member AggR. ANR-AggR binding disrupted AggR dimerization and prevented AggR-DNA binding. ANR homologs of Vibrio cholera, Citrobacter rodentium, Salmonella enterica and ETEC were capable of complementing Aar activity by repressing aggR expression in EAEC strain 042. ANR homologs of ETEC and Vibrio cholerae bound to AggR as well as to other members of the AraC family, including Rns and ToxT. The predicted proteins of all ANR members exhibit three highly conserved predicted α-helices. Site-directed mutagenesis studies suggest that at least predicted α-helices 2 and 3 are required for Aar activity. In sum, our data strongly suggest that members of the novel ANR family act by directly binding to their cognate AraC partners.

The Hcp-like protein HilE inhibits homodimerization and DNA binding of the virulence-associated transcriptional regulator HilD in Salmonella

“…Consistently, the HilD-binding sites on different genes have two direct repeat sequences (21,27,51). Several other AraC-like transcriptional regulators act as dimers, such as ToxT, ExsA, UreR and AggR, which control expression of virulence genes in Vibrio cholerae, Pseudomonas aeruginosa, Proteus mirabilis and E. coli (45,48,49,61,62). HilE interacts with the central region of HilD and thus inhibits its dimerization, which indirectly would affect the DNA binding activity of this regulator.…”

“…This double interaction and effect supports a tight control of the HilD activity by HilE. In enteroagregative E. coli (EAEC), the Aar protein interacts with the central region of AggR, containing the dimerization domain, inhibiting the dimerization and the DNA binding of this AraClike regulator (62). Similarly, in P. aeruginosa, the ExsD protein interacts with the N-terminal region of ExsA, containing the dimerization domain, thus preventing the dimerization and DNA binding of this regulator (45,61,63).…”

Genetic heterogeneity of the Spy1336/R28—Spy1337 virulence axis in Streptococcus pyogenes and effect on gene transcript levels and pathogenesis

“…Recently, we proposed that the Spy1337 protein is a positive transcriptional regulator of both Spy1336/R28 and Spy1337, and that by regulating expression of Spy1336/R28 and other genes, Spy1337 is involved in emm28 GAS virulence [38]. To positively regulate virulence gene expression, AraC-like transcriptional regulators usually either bind to chemical effectors present at the site of infection to cause a conformational change favoring DNA-binding to their cognate gene targets [39,40], or they bind to AraC negative regulators (ANRs) that inhibits such interactions [41]. In addition, they can regulate their own expression [42][43][44][45][46].…”

A novel mouse model ofCampylobacter jejunienteropathy and diarrhea

“…Thus we can now explore whether zinc or key repair nutrients might ameliorate these disease outcomes. For example, zinc can suppress expression of not only such host genes as the CFTR chloride channel that is responsible for secretory diarrhea with cholera and the muc-2 gene, but also microbial genes such as the aggR -regulated virulence gene expression in enteroaggregative E. coli (EAEC) in vitro or in vivo [33], and homologous genes have been reported in several other enteric bacterial pathogens like Vibrio cholerae , Shigella and Citrobacter, and possibly Campylobacter [49, 50]. Effects of zinc on enteric infections are complex.…”