Identification of Small Molecule Inhibitors of Pseudomonas aeruginosa Exoenzyme S Using a Yeast Phenotypic Screen

Pseudomonas aeruginosa is an opportunistic human pathogen that is a key factor in the mortality of cystic fibrosis patients, and infection represents an increased threat for human health worldwide. Because resistance of Pseudomonas aeruginosa to antibiotics is increasing, new inhibitors of pharmacologically validated targets of this bacterium are needed. Here we demonstrate that a cell-based yeast phenotypic assay, combined with a large-scale inhibitor screen, identified small molecule inhibitors that can suppress the toxicity caused by heterologous expression of selected Pseudomonas aeruginosa ORFs. We identified the first small molecule inhibitor of Exoenzyme S (ExoS), a toxin involved in Type III secretion. We show that this inhibitor, exosin, modulates ExoS ADP-ribosyltransferase activity in vitro, suggesting the inhibition is direct. Moreover, exosin and two of its analogues display a significant protective effect against Pseudomonas infection in vivo. Furthermore, because the assay was performed in yeast, we were able to demonstrate that several yeast homologues of the known human ExoS targets are likely ADP-ribosylated by the toxin. For example, using an in vitro enzymatic assay, we demonstrate that yeast Ras2p is directly modified by ExoS. Lastly, by surveying a collection of yeast deletion mutants, we identified Bmh1p, a yeast homologue of the human FAS, as an ExoS cofactor, revealing that portions of the bacterial toxin mode of action are conserved from yeast to human. Taken together, our integrated cell-based, chemical-genetic approach demonstrates that such screens can augment traditional drug screening approaches and facilitate the discovery of new compounds against a broad range of human pathogens

Sex specific alterations in preterm brain

Background: The literature on brain imaging in premature infants, is mostly made up of studies that evaluate neonates, yet the most dynamic time of brain development happens from birth to one year of age. This study was designed to obtain quantitative brain measures from Magnetic Resonance Imaging (MRI) scans of infants born prematurely at 12 months of age. Methods: The subject group was designed to capture a wide range of gestational age (GA) from premature to full term infants. An age-specific atlas generated quantitative brain measures. A regression model was used to predict effects of gestational age, sex, on brain measures. Results: There was a primary effect of sex on: 1) intracranial volume (ICV), males > females; 2) proportional cerebral cortical gray matter (females > males) and 3) cerebral white matter (males> females). GA predicted cerebral volume and cerebral spinal fluid (CSF). GA also predicted cortical gray matter in a sex specific manner with GA having a significant effect on cortical volume in the males, but not in females. Conclusions and Relevance: Sex differences in brain structure are large early in life. GA had sex specific effects highlighting the importance evaluating sex effects in neurodevelopmental outcomes of premature infants