Evolution of a Bacterial Regulon Controlling Virulence and Mg2+ Homeostasis

Related organisms typically rely on orthologous regulatory proteins to respond to a given signal. However, the extent to which (or even if) the targets of shared regulatory proteins are maintained across species has remained largely unknown. This question is of particular significance in bacteria due to the widespread effects of horizontal gene transfer. Here, we address this question by investigating the regulons controlled by the DNA-binding PhoP protein, which governs virulence and Mg2+ homeostasis in several bacterial species. We establish that the ancestral PhoP protein directs largely different gene sets in ten analyzed species of the family Enterobacteriaceae, reflecting both regulation of species-specific targets and transcriptional rewiring of shared genes. The two targets directly activated by PhoP in all ten species (the most distant of which diverged >200 million years ago), and coding for the most conserved proteins are the phoPQ operon itself and the lipoprotein-encoding slyB gene, which decreases PhoP protein activity. The Mg2+-responsive PhoP protein dictates expression of Mg2+ transporters and of enzymes that modify Mg2+-binding sites in the cell envelope in most analyzed species. In contrast to the core PhoP regulon, which determines the amount of active PhoP and copes with the low Mg2+ stress, the variable members of the regulon contribute species-specific traits, a property shared with regulons controlled by dissimilar regulatory proteins and responding to different signals

Evolution of a Bacterial Regulon Controlling Virulence and Mg2+ Homeostasis

Related organisms typically rely on orthologous regulatory proteins to respond to a given signal. However, the extent to which (or even if) the targets of shared regulatory proteins are maintained across species has remained largely unknown. This question is of particular significance in bacteria due to the widespread effects of horizontal gene transfer. Here, we address this question by investigating the regulons controlled by the DNA-binding PhoP protein, which governs virulence and Mg2+ homeostasis in several bacterial species. We establish that the ancestral PhoP protein directs largely different gene sets in ten analyzed species of the family Enterobacteriaceae, reflecting both regulation of species-specific targets and transcriptional rewiring of shared genes. The two targets directly activated by PhoP in all ten species (the most distant of which diverged >200 million years ago), and coding for the most conserved proteins are the phoPQ operon itself and the lipoprotein-encoding slyB gene, which decreases PhoP protein activity. The Mg2+-responsive PhoP protein dictates expression of Mg2+ transporters and of enzymes that modify Mg2+-binding sites in the cell envelope in most analyzed species. In contrast to the core PhoP regulon, which determines the amount of active PhoP and copes with the low Mg2+ stress, the variable members of the regulon contribute species-specific traits, a property shared with regulons controlled by dissimilar regulatory proteins and responding to different signals

The impact of antiretroviral therapy on HPV and cervical intraepithelial neoplasia: current evidence and directions for future research

Increasing numbers of human immunodeficiency virus (HIV)-infected women are now accessing life-prolonging highly active antiretroviral therapy (HAART) in developing countries. There is a need for better understanding of interactions of human papillomavirus (HPV) and HIV, especially in the context of increasing life expectancy due to HAART. The data regarding the impact of HAART on reducing the incidence and progression and facilitating the regression of HPV infection and cervical abnormalities is largely inconsistent. Published studies differ in their study designs (prospective or retrospective cohorts or record linkage studies), screening and diagnostic protocols, duration and type of HAART use, recruitment and referral strategies, and definitions of screening test and disease positivity. Due to the ethical and resource limitations in conducting randomized trials of the impact of HAART on incidence of HPV, CIN, and cervical cancer among HIV-infected women, it is important to consider innovative study designs, including quasi-experimental trials and operations research in sentinel populations to answer the critical research questions in this area

An Italian prospective multicenter study on colonoscopy practice and quality: What has changed in the last 10 years

Background: A relevant number of adenomas can be missed during colonoscopy. Aims: Assess the current status of colonoscopy procedures in Italian centers. Methods: A prospective observational study involving 17 hospitals (34 endoscopists) included consecutive patients undergoing standard colonoscopy. In the first phase, endoscopists performed consecutive colonoscopies. In the second phase, retraining via an online learning platform was planned, while in the third phase data were collected analogously to phase 1. Results: A total of 3,504 patients were enrolled. Overall, a BBPS score ≥6 was obtained in 95.6% of cases (94.8% and 96.9% in the pre- and post-training phases, respectively). 88.4% of colonoscopies had a withdrawal time ≥6 min (88.2% and 88.7% in the pre- and post-training phases). Median adenoma detection rate (ADR) was 39.1%, with no significant differences between the pre- and post-training phases (40.1% vs 36.9%; P = 0.83). In total, 81% of endoscopists had a ADR performance above the 25% threshold. Conclusion: High colonoscopy quality standards are achieved by the Italian hospitals involved. Quality improvement initiatives and repeated module-based colonoscopy-training have been promoted in Italy during the last decade, which appear to have had a significant impact on quality colonoscopy metrics together with the activation of colorectal cancer screening programs

Comparative Proteomic Analysis of the PhoP Regulon in Salmonella enterica Serovar Typhi Versus Typhimurium

Background: S. Typhi, a human-restricted Salmonella enterica serovar, causes a systemic intracellular infection in humans (typhoid fever). In comparison, S. Typhimurium causes gastroenteritis in humans, but causes a systemic typhoidal illness in mice. The PhoP regulon is a well studied two component (PhoP/Q) coordinately regulated network of genes whose expression is required for intracellular survival of S. enterica. Methodology/Principal Findings: Using high performance liquid chromatography mass spectrometry (HPLC-MS/MS), we examined the protein expression profiles of three sequenced S. enterica strains: S. Typhimurium LT2, S. Typhi CT18, and S. Typhi Ty2 in PhoP-inducing and non-inducing conditions in vitro and compared these results to profiles of $phoP^−/Q^−$ mutants derived from S. Typhimurium LT2 and S. Typhi Ty2. Our analysis identified 53 proteins in S. Typhimurium LT2 and 56 proteins in S. Typhi that were regulated in a PhoP-dependent manner. As expected, many proteins identified in S. Typhi demonstrated concordant differential expression with a homologous protein in S. Typhimurium. However, three proteins (HlyE, STY1499, and CdtB) had no homolog in S. Typhimurium. HlyE is a pore-forming toxin. STY1499 encodes a stably expressed protein of unknown function transcribed in the same operon as HlyE. CdtB is a cytolethal distending toxin associated with DNA damage, cell cycle arrest, and cellular distension. Gene expression studies confirmed up-regulation of mRNA of HlyE, STY1499, and CdtB in S. Typhi in PhoP-inducing conditions. Conclusions/Significance: This study is the first protein expression study of the PhoP virulence associated regulon using strains of Salmonella mutant in PhoP, has identified three Typhi-unique proteins (CdtB, HlyE and STY1499) that are not present in the genome of the wide host-range Typhimurium, and includes the first protein expression profiling of a live attenuated bacterial vaccine studied in humans (Ty800)

Speed, Sensitivity, and Bistability in Auto-activating Signaling Circuits

Cells employ a myriad of signaling circuits to detect environmental signals and drive specific gene expression responses. A common motif in these circuits is inducible auto-activation: a transcription factor that activates its own transcription upon activation by a ligand or by post-transcriptional modification. Examples range from the two-component signaling systems in bacteria and plants to the genetic circuits of animal viruses such as HIV. We here present a theoretical study of such circuits, based on analytical calculations, numerical computations, and simulation. Our results reveal several surprising characteristics. They show that auto-activation can drastically enhance the sensitivity of the circuit's response to input signals: even without molecular cooperativity, an ultra-sensitive threshold response can be obtained. However, the increased sensitivity comes at a cost: auto-activation tends to severely slow down the speed of induction, a stochastic effect that was strongly underestimated by earlier deterministic models. This slow-induction effect again requires no molecular cooperativity and is intimately related to the bimodality recently observed in non-cooperative auto-activation circuits. These phenomena pose strong constraints on the use of auto-activation in signaling networks. To achieve both a high sensitivity and a rapid induction, an inducible auto-activation circuit is predicted to acquire low cooperativity and low fold-induction. Examples from Escherichia coli's two-component signaling systems support these predictions

Molecular Mechanics of the α-Actinin Rod Domain: Bending, Torsional, and Extensional Behavior

α-Actinin is an actin crosslinking molecule that can serve as a scaffold and maintain dynamic actin filament networks. As a crosslinker in the stressed cytoskeleton, α-actinin can retain conformation, function, and strength. α-Actinin has an actin binding domain and a calmodulin homology domain separated by a long rod domain. Using molecular dynamics and normal mode analysis, we suggest that the α-actinin rod domain has flexible terminal regions which can twist and extend under mechanical stress, yet has a highly rigid interior region stabilized by aromatic packing within each spectrin repeat, by electrostatic interactions between the spectrin repeats, and by strong salt bridges between its two anti-parallel monomers. By exploring the natural vibrations of the α-actinin rod domain and by conducting bending molecular dynamics simulations we also predict that bending of the rod domain is possible with minimal force. We introduce computational methods for analyzing the torsional strain of molecules using rotating constraints. Molecular dynamics extension of the α-actinin rod is also performed, demonstrating transduction of the unfolding forces across salt bridges to the associated monomer of the α-actinin rod domain

Nitric Oxide Antagonizes the Acid Tolerance Response that Protects Salmonella against Innate Gastric Defenses

Reactive nitrogen species (RNS) derived from dietary and salivary inorganic nitrogen oxides foment innate host defenses associated with the acidity of the stomach. The mechanisms by which these reactive species exert antimicrobial activity in the gastric lumen are, however, poorly understood.The genetically tractable acid tolerance response (ATR) that enables enteropathogens to survive harsh acidity was screened for signaling pathways responsive to RNS. The nitric oxide (NO) donor spermine NONOate derepressed the Fur regulon that controls secondary lines of resistance against organic acids. Despite inducing a Fur-mediated adaptive response, acidified RNS largely repressed oral virulence as demonstrated by the fact that Salmonella bacteria exposed to NO donors during mildly acidic conditions were shed in low amounts in feces and exhibited ameliorated oral virulence. NO prevented Salmonella from mounting a de novo ATR, but was unable to suppress an already functional protective response, suggesting that RNS target regulatory cascades but not their effectors. Transcriptional and translational analyses revealed that the PhoPQ signaling cascade is a critical ATR target of NO in rapidly growing Salmonella. Inhibition of PhoPQ signaling appears to contribute to most of the NO-mediated abrogation of the ATR in log phase bacteria, because the augmented acid sensitivity of phoQ-deficient Salmonella was not further enhanced after RNS treatment.Since PhoPQ-regulated acid resistance is widespread in enteric pathogens, the RNS-mediated inhibition of the Salmonella ATR described herein may represent a common component of innate host defenses

Early Hypothalamic FTO Overexpression in Response to Maternal Obesity – Potential Contribution to Postweaning Hyperphagia

Intrauterine and postnatal overnutrition program hyperphagia, adiposity and glucose intolerance in offspring. Single-nucleotide polymorphisms (SNPs) of the fat mass and obesity associated (FTO) gene have been linked to increased risk of obesity. FTO is highly expressed in hypothalamic regions critical for energy balance and hyperphagic phenotypes were linked with FTO SNPs. As nutrition during fetal development can influence the expression of genes involved in metabolic function, we investigated the impact of maternal obesity on FTO.Female Sprague Dawley rats were exposed to chow or high fat diet (HFD) for 5 weeks before mating, throughout gestation and lactation. On postnatal day 1 (PND1), some litters were adjusted to 3 pups (vs. 12 control) to induce postnatal overnutrition. At PND20, rats were weaned onto chow or HFD for 15 weeks. FTO mRNA expression in the hypothalamus and liver, as well as hepatic markers of lipid metabolism were measured.At weaning, hypothalamic FTO mRNA expression was increased significantly in offspring of obese mothers and FTO was correlated with both visceral and epididymal fat mass (P<0.05); body weight approached significance (P = 0.07). Hepatic FTO and Fatty Acid Synthase mRNA expression were decreased by maternal obesity. At 18 weeks, FTO mRNA expression did not differ between groups; however body weight was significantly correlated with hypothalamic FTO. Postnatal HFD feeding significantly reduced hepatic Carnitine Palmitoyltransferase-1a but did not affect the expression of other hepatic markers investigated. FTO was not affected by chronic HFD feeding.Maternal obesity significantly impacted FTO expression in both hypothalamus and liver at weaning. Early overexpression of hypothalamic FTO correlated with increased adiposity and later food intake of siblings exposed to HFD suggesting upregulation of FTO may contribute to subsequent hyperphagia, in line with some human data. No effect of maternal obesity was observed on FTO in adulthood

Different populations and sources of human mesenchymal stem cells (MSC): A comparison of adult and neonatal tissue-derived MSC

The mesenchymal stroma harbors an important population of cells that possess stem cell-like characteristics including self renewal and differentiation capacities and can be derived from a variety of different sources. These multipotent mesenchymal stem cells (MSC) can be found in nearly all tissues and are mostly located in perivascular niches. MSC have migratory abilities and can secrete protective factors and act as a primary matrix for tissue regeneration during inflammation, tissue injuries and certain cancers