Construction of a bacterial autoinducer detection system in mammalian cells

Quorum sensing (QS) is a cell density-dependent signaling system used by bacteria to coordinate gene expression within a population. QS systems in Gram negative bacteria consist of transcription factors of the LuxR family and their acyl homoserine lactone (AHL) ligands. We describe here a method for examining QS signaling systems in mammalian cells that uses engineered LuxR-type proteins from the opportunistic pathogen, Pseudomonas aeruginosa, which can function as AHL-dependent transcription factors. The engineered proteins respond to their cognate ligands and display sequence specific DNA binding properties. This system has several potential biotechnological and biological applications. It may be used to characterize any LuxR-type protein, screen animal and plant cell extracts or exudates for compounds that mimic or interfere with AHL signaling or to screen different cell types for AHL inactivating activities

Peroxisome Proliferator-Activated Receptors Mediate Host Cell Proinflammatory Responses to Pseudomonas aeruginosa Autoinducer▿

The pathogenic bacterium Pseudomonas aeruginosa utilizes the 3-oxododecanoyl homoserine lactone (3OC12-HSL) autoinducer as a signaling molecule to coordinate the expression of virulence genes through quorum sensing. 3OC12-HSL also affects responses in host cells, including the upregulation of genes encoding inflammatory cytokines. This proinflammatory response may exacerbate underlying disease during P. aeruginosa infections. The specific mechanism(s) through which 3OC12-HSL influences host responses is unclear, and no mammalian receptors for 3OC12-HSL have been identified to date. Here, we report that 3OC12-HSL increases mRNA levels for a common panel of proinflammatory genes in murine fibroblasts and human lung epithelial cells. To identify putative 3OC12-HSL receptors, we examined the expression patterns of a panel of nuclear hormone receptors in these two cell lines and determined that both peroxisome proliferator-activated receptor beta/delta (PPARβ/δ) and PPARγ were expressed. 3OC12-HSL functioned as an agonist of PPARβ/δ transcriptional activity and an antagonist of PPARγ transcriptional activity and inhibited the DNA binding ability of PPARγ. The proinflammatory effect of 3OC12-HSL in lung epithelial cells was blocked by the PPARγ agonist rosiglitazone, suggesting that 3OC12-HSL and rosiglitazone are mutually antagonistic negative and positive regulators of PPARγ activity, respectively. These data identify PPARβ/δ and PPARγ as putative mammalian 3OC12-HSL receptors and suggest that PPARγ agonists may be employed as anti-inflammatory therapeutics for P. aeruginosa infections

Practical Singly and Doubly Electrophilic Aminating Agents: A New, More Sustainable Platform for Carbon–Nitrogen Bond Formation

Given the importance of amines in a large number of biologically active natural products, active pharmaceutical ingredients, agro­chemicals, and functional materials, the development of efficient C–N bond-forming methods with wide substrate scope continues to be at the frontier of research in synthetic organic chemistry. Here, we present a general and fundamentally new synthetic approach for the direct, transition-metal-free preparation of symmetrical and unsymmetrical diaryl-, aryl­alkyl-, and dialkyl­amines that relies on the facile single or double addition of readily available <i>C</i>-nucleo­philes to the nitrogen atom of bench-stable electro­philic aminating agents. Practical single and double polarity reversal (i.e., umpolung) of the nitrogen atom is achieved using sterically and electronically tunable keto­malonate-derived imines and oximes. Overall, this novel approach represents an operationally simple, scalable, and environmentally friendly alternative to transition-metal-catalyzed C–N cross-coupling methods that are currently used to access structurally diverse secondary amines

Role of birds of prey as carriers and spreaders of Cryptococcus neoformans and other zoonotic yeasts

In the last 20 years, cases of human cryptococcosis, have increased in immunocompromised patients. In several instances, the cases have been associated with the exposure of the patients to bird droppings. In order to investigate birds of prey as potential carriers and spreaders of Cryptococcus neoformans and other yeasts of importance in human infections, 182 swab samples were collected from the cloacae of several species of birds of prey (Group I) and 32 faecal samples from aviaries in which the birds were housed (Group II). Samples were also taken from digestive tract of 60 dead birds (Group III). A total of 454 samples were cultured from which 215 colonies of yeastlike fungi were recovered and identified. Cryptococcusneoformans var. grubii was isolated from three cloacae samples (4.8%) collected from Falco tinnunculus and from one sample (3.1%) obtained from Buteo buteo, as well as from samples collected at the aviaries in which these birds were kept. Overall, 18 samples (9.9%) from Group I, 13 (40.6%) from Group II, 12 crops (20%), three proventriculi (5%) and 12 cloacae (20%) from Group III yielded positive cultures for yeasts. The results indicate that birds of prey and in particular, F. tinnunculus and B. buteo, may act as carriers and spreaders of C. neoformans and other zoonotic yeasts